New liposome compositions

FIELD: medicine, pharmaceutics.

SUBSTANCE: group of inventions refers to medicine and is applicable in the form of liposome-containing compounds for cancer therapy. A liposome contains one or more phosphatidylcholines, a first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, an orientation modified derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, an encapsulated therapeutic agent and at least one additional lipid which represents cholesterol or a cholesterol derivative. The orientation modified derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine contains an orientation ligand attached to a second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine. The first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine is presented by formula 1,

and the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine is presented by formula 3, . The orientation ligand preferentially represents transferring, while the encapsulated therapeutic agent is oxalyplatine. The liposome is free from non-modified phosphatidylethanolamine, egg phosphatidylcholine or hydrophilic polymer used to prolong a half lifetime of the liposome in a circulatory channel, and the orientation ligand is other than an intact antibody. What is also described is a method of producing the liposome and a method of treating cancer with using it.

EFFECT: group of inventions provides better target delivery of the therapeutic substance in the tumour cells.

113 cl, 25 dwg, 4 tbl, 30 ex

 

Cross-reference to related applications

In this application claims the priority of patent application of Japan No. 2005-67469 filed March 10, 2005, the contents of which are incorporated here by reference in its entirety.

The level of technology

The effectiveness of the treatment of many diseases, including cancer, has improved significantly over the last few decades, however, many treatment regimens require the use of drugs with dangerous side effects, including, for example, alopecia, nausea, vomiting, weakness, etc. In some schemes the introduction can also be assigned to the use of drugs that are not stable under physiological conditions, such as biological therapeutic agent (e.g., genes or gene products) and/or other medicinal substances, which easily disintegrate or otherwise changed with the introduction and thus lose their effectiveness to achieve the desired therapeutic result. Such instability also makes it more difficult and expensive storage of medicinal substances and the production of drugs for injection.

There are several classes of anticancer agents, covering about 100 specific drugs, as well as numerous schemes of combined drug therapy, methods of delivery and treatment. The contradictory, the same agents can be classified according to several criteria, such as the connection class and the disease being treated. Specific agents that have advantages in the rapid division of cancer cells and directly acting on specific phases of the cell cycle, presenting a different classification method. Agents can be grouped according to the type and severity of their side effects or delivery method. However, the most generally accepted classification of anticancer agents based on biotherapy is the classification according to the class of chemical compounds that are widely covers the mechanism of action of these compounds.

Depending on the source of information, there are small differences in the classification of anticancer agents. Classes of compounds are described in Physician''s Desk Reference as follows: alkaloids; alkylating agents; antitumor antibiotics; antimetabolites; hormones and analogues hormones; immunomodulators; photosensitizing agents; and other heterogeneous agents.

Compounds of the class of alkaloids can also be identified as inhibitors of mitosis, as they are specific to a particular phase of the cell cycle and are designed to inhibition of mitosis or inhibit enzymes required for mitosis. They occur usually from plant alkaloids and other natural products and are the during M-phase of the cell cycle. This class of compounds commonly used for the treatment of neoplasms, such as acute lymphoblastic leukemia, Hodgkin's and non-Hodgkin's lymphoma; neuroblastoma and lung cancer, mammary gland and the testes.

Alkylating agents represent a large class of chemotherapeutic agents, including the following subclasses, each of which represents the number of individual drug substances: alkyl sulphonates; aziridine; ethylenimine and methylmelamine; derivatives of mustard gas derivative nitrosoanatabine; and others, including platinum compounds. Alkylating agents attack the neoplastic cells by direct alkylation of the DNA of the cells and, consequently, cause the malfunction of DNA replication. This class of compounds typically used to treat a variety of diseases, including chronic leukemia, lymphoma Nahodkinskogo type, Hodgkin's lymphoma, multiple myeloma and certain types of lung cancer, breast cancer and ovarian cancer.

Derivative nitrosoanatabine often included in the category of alkylating agents, and they have a similar mechanism of action, but instead direct alkylation of DNA they inhibit the DNA repair enzymes, causing a violation of reparation. These compounds have the advantages of being able to cross the blood-brain barrier and, therefore, can be used in the us for the treatment of brain tumors.

Antitumor antibiotics possess antimicrobial and cytotoxic activity and also affect the DNA by chemical inhibition of enzymes and mitosis or by changing cell membranes. They are not specific phases of the cell cycle and are widely used for the treatment of cancer.

Antimetabolite class of anticancer agents affects the growth of DNA and RNA and is specific for the S phase of the cell cycle. They can be further classified by the type of connection that includes analogs of folic acid, purine analogues, and pyrimidine analogues. They are often used in the treatment of chronic leukemias, tumors of the breast, ovary, gastrointestinal tract.

There are two classes of hormones or analogues hormones, used as an anticancer agent, a corticosteroid hormones and sex hormones. While some of corticosteroid hormones can kill cancer cells and slow tumor growth, and they are used to treat lymphoma, leukemia, etc., sex hormones operate primarily by reducing the growth of cancerous tumors of the breast, prostate, and endometrium. There are numerous subclasses of hormones and hormonal analogues, including androgen antagonists hormone of the adrenal cortex, antiandrogens, antiestrogens, inhibitors arene is tazy, the estrogen analogues releasing factor, luteinizing hormone (LHRH) and progestins.

Additional small class of anticancer agents classified as immunotherapy. There are agents that are designed to stimulate the immune system to more effectively attack on neoplastic (cancerous) cells. This therapy is often used in combination with other therapies.

There are also a number of compounds, such as campothecine, which are usually listed as "other" anti-cancer agents, and they can be applied for the treatment of various neoplasms.

The combination anticancer agents is also used to treat some types of cancer. For example, Sanofi Syntholabo sells ELOXATIN™ (oxaliplatin for injection) for the treatment of colorectal cancer for use in combination with 5-fluorouracil and leukocoria. This combination of drugs is often used in conjunction with surgical intervention in the treatment of colorectal cancer. Oxaliplatin is an alkylating agent, which is thought to be acts by inhibiting both replication and transcription of DNA. Unlike other agents, containing platinum, oxaliplatin demonstrated reduced the likelihood of sustainability. Oxaliplatin is additionally described in U.S. patents Nos. 4169846; 5338874; 5298642; 595913; 5420319; 5716988; 5290961; and Wilkes GM. "New therapeutic options in colon cancer: focus on oxaliplatin " Clin J Oncol Nurs. (2002) 6:131-137.

Although there are many anti-cancer agents, the advantages of these compounds are often outweighed by the severity of side effects provided by the agent. This ratio is often referred to as therapeutic index, which describes the balance between the required dose to achieve the destruction of cancer cells compared to the dose at which the substance unacceptably toxic to the individual. Obstacle for most anticancer agents is relatively low range of therapeutic index (i.e. a narrow range of doses in which cancer cells are destroyed without unacceptable toxicity for the individual). This characteristic limits the frequency and dosage, when the agent is useful, and often the side effects become unbearable before cancer can be eradicated.

Serious side effects that occur with most types of cancer chemotherapy, are the result of a nonspecific nature of these drugs, which do not distinguish between healthy and cancer cells, and instead of destroying them. With the help of certain medicinal substances, specific phases of the cell cycle, trying to reduce these effects, directionally acting on the phase of the cell C is KLA, involved in replication and cell division. These drugs, however, do not distinguish between cancer cells and healthy cells that are found in normal cell division. Cells that suffer the most from these types of chemotherapy, are those who often undergo cell division, including blood cells, cells of hair follicles and cells in the sexual and digestive tracts.

The most common side effects of anticancer agents are nausea and vomiting. The majority of individuals suffering from myelosuppressive or suppress the function of the bone marrow, which produces red blood cells, white blood cells and platelets. These and other side effects are further compounded by the suppression of the immune system, accompanied by the destruction and lack of production of white blood cells and the associated risk of opportunistic infections.

Other side effects common to a broad spectrum of anticancer agents include: hair loss (alopecia); weight loss; changes in taste; stomatitis and esophagitis (inflammation and ulcers); constipation; diarrhea; weakness; heart damage; changes of the nervous system; lung damage; damage to reproductive tissues; liver damage; kidney damage and urinary system.

Wide dipsophobia effects associated with most anti-cancer agents and their severity in individuals who are already weakened by disease and immunodeficient, has forced researchers to look for the mechanisms through which they can relieve some of the side effects, while maintaining at the same time, the effectiveness of the treatment. There have been several approaches to this problem. These include concomitant chemotherapy, when many anti-cancer agents are administered together; adjuvant therapy, when additional agents are prescribed together with an anticancer agent to deal with the side effects of anticancer agent; the combination of different treatment modalities, when chemotherapy is combined with radiation and/or surgical intervention; alternative media for delivery to the introduction of anti-cancer agents, liposomes with encapsulated anticancer agents.

Liposomes are formed when phospholipids and their derivatives are dispersed in water. When dispersion in water, phospholipids form closed vesicles called liposomes, which are characterized by a lipid bilayer encapsulating water-based. Different liposomes were used as carriers for making them therapeutic agents such as drugs, enzymes and genetic sequences for use in honey is medical science, in pharmaceutical science and biochemistry.

Examples of liposomal compositions include U.S. patents Nos. 4983397; 6476068; 5834012; 5756069; 6387397; 5534241; 4789633; 4925661; 6153596; 6057299; 5648478; 6723338; 6627218; patent applications U.S. Nos: 2003/0224037; 2004/0022842; 2001/0033860; 2003/0072794; 2003/0082228; 2003/0212031; 2003/0203865; 2004/0142025; 2004/0071768; international patent application WO 00/74646; WO 96/13250; WO 98/33481; Papahadjopolulos D, Allen TM, Gbizon A, et al., "Sterically stabilized liposomes: Improvements in pharmacokinetics and antitumor therapeutic efficacy" Proc Natl Acad Sci USA (1991) 88: 11460-11464; Allen TM, Martin FJ. "Advantages of liposomal delivery systems for anthracyclines" Semin Oncol (2004) 31: 5-15 (suppl 13). Weissig et al. Pharm. Res. (1998) 15: 1552-1556.

At earlier stages in the development of liposomes used existing in the nature of the phospholipids of cell membranes, such as the phospholipids of egg yolk and soybean phospholipids. In the case of intravenous administration, however, liposomes using these phospholipids was probably involved in the reticuloendothelial system of the liver or spleen, a short stay in the blood resulting in reducing the effectiveness of the drug. After this, in the form of a solution to this problem as part of the membrane of liposomes used synthetic phospholipids, whose lipid part contains only saturated connection, in order to make a more solid membrane of liposomes.

When you try to increase the half-life in the circulatory direction and avoid capture reticuloendothelial the social system researchers have developed liposomes which has been modified by inclusion of peg or other hydrophilic polymers (such as PEG a liposome, in which one or more of the components of lipids has been modified by attaching PEG). Modified PEG-liposomes was also frequently described as "shielded" liposomes. Doxil™ (liposomes with doxorubicin HCl for injection) is enclosed in the liposome doxorubicin with the addition of polyethylene glycol (PEG), used to avoid the reticuloendothelial system (RES) and prolongation of the circulation time of the drug. See Vail DM, Amantea MA, Colbern GT, et al., "Pegylated Liposomal Doxorubicin: Proof of Principle Using Preclinical Animal Models and Pharmacokinetic Studies." Semin Oncol. (2004) 31 (Suppl 13): 16-35. However, adverse effects were also called prolonged retention in the blood (for example, the syndrome of the hands and feet, the adverse effect of Doxil™ peripheral system and so on), which became a recognized problem.

Examples of liposomes include U.S. patents Nos. 4983397; 5013556; 6316024; 6056973; 5945122; 5891468; 6126966; 5593622, 5676971; 6586559; and 5846458, the publication of patent applications U.S. Nos. 2003/0224037; 2004/0022842; 2003/0113262; 2002/0136707; international patent application WO 99/30686; WO 02/41870 Aliminana et al., Prep. Biochem. Biotech. (2004) 34(1): 77-96. Liposomes are described in U.S. patents Nos. 6228391; 6197333; 6046225; 5292524; and in the publications US patents Nos. 20050271588; 20040213833; 20040029210; 20030175205; 20030162748; 20030130190; 20030059461; and 2002003453.

In addition to liposomes, modified PEG, researchers have developed many other derivatives of lipids. These derived lipids may also be included in liposomes. See, for example, international patent application WO 93/01828; Park YS, Maruyama K, Huang L. "Some negatively charged phospholipids derivatives prolong the liposome circulation in vivo." Biochimica et Biophysica Acta (1992) 1108: 257-260; Ahl et al., Biochimica Biophys. Acta (1997) 1329: 370-382.

Additional lipid compositions described in U.S. patent Nos. 6936272; 6897196; 6077834; and in the publications US patents Nos. 20050136064; 20040234588; 20030215490; 20030166601; and 20010038851.

In addition to the modification of liposomes with PEG and other hydrophilic polymers, researchers have also developed liposomes, which are designed for specific target delivery to specific cell types by including guiding factors (also referred to as guides ligands for specific cell types. Examples of guiding factors/ligands include asialoglycoprotein, folate, transferrin, antibodies, etc. In some cases, one or more components of the lipids can be modified by attaching the guiding factor.

Examples of lipid compositions, including guiding factors include the U.S. patents Nos. 5049390; 5780052; 5786214; 6316024; 6056973; 6245427; 6524613; 6749863; 6177059; 6530944; patent applications U.S. Nos. 2004/0022842; 2003/0224037; 2003/143742; 2003/0228285; 2002/0198164; 2003/0220284; 2003/0165934; 2003/007779; international patent application Nos. WO 95/33841; WO 95/19434; WO 2001037807; WO 96/33698; WO 2001/49266; WO 9940789; WO 9925320; WO 9104014; WO 92/07959; EP 1369132; JP 2001002592; Iinuma H, Maruyama K, et al., "Intracellular targeting therapy of cisplatin-encapsulated transferrin-polyethylene glycol liposome on peritoneal dissemination of gastric cancer" Int J Cancer (2002) 99 130-137; Ishida O, Maruyama K, Tanahashi H, Iwatsuru M, Sasaki K, et al., "Liposomes bearing polyethylene glycol-coupled transferrin with intracellular targeting property to the solid tumors in vivo." Pharmaceutical Research (2001) 18: 1042-1048; Holmberg et al., Biochem. Biophys. Res. Comm. (1989) 165(3): 1272-1278; Nam et al., J. Biochem. Mol Biol (1998) 31(1): 95-100; Nag et al., J. Drug Target. (1999) 6(6): 427-438.

In particular, Iinuma et al. developed Tf-PEG-liposome with transferrin (Tf), attached to the surface of liposomes. Iinuma et al. showed that a greater number of liposomes was contacted with the surface of tumor cells and observed a higher capture of liposomes tumor cells in the case of Tf-PEG-liposomes compared to PEG-liposomes (Inuma et al., ibid; Ishida et al., ibid.).

However, despite recent advances made in the field of drug delivery and labeled compounds, including the use of formulations of liposomes, there remains a need for improved lipid compositions for drug delivery and labeled compounds to specific cells and/or tissues, thus achieving a therapeutic or diagnostic effect. In particular, in the field of Oncology medicinal compositions with improved specificity and reduced t what xinou necessary for certain therapeutic success without the adverse effects on healthy cells and also without the induction of dangerous side effects in the individual, being treated. Similar, labeled compounds, which can be used to diagnose conditions, especially life-threatening conditions, at an early stage (for example, with high specificity and/or high sensitivity) and also for accurate monitoring of the severity/extent of condition (e.g., progression or regression with or without treatment), should also significantly improve the quality and success of therapy.

Summary of the invention

The present invention relates to a new lipid containing compositions (including liposomes (for example, liposomes directional unloaded liposomes), a lipid mixture containing liposomes of the composition), which may not necessarily include drug or labeled compound or can be applied to obtain compositions that include a drug or labeled compound, where lipid containing composition creates benefits in terms of reducing side effects of drug or labeled compound, and/or prevention of the destruction and/or loss of effectiveness of the drug or labeled compound. The invention also includes methods of preparation and use are described here lipid containing compositions. In certain aspects of the invention containing the lipid composition can b shall be applied for the treatment or diagnosis of cancer (for example, breast cancer, gastric cancer, colorectal cancer, colon cancer, pancreatic cancer, not small cell lung cancer, small cell lung cancer, brain cancer, liver cancer, kidney cancer, prostate cancer, bladder cancer, ovarian cancer or hematologic malignancies (e.g., leukemia, lymphoma, multiple myeloma etc).

In certain embodiments provides liposomes directed action, including one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine encapsulated drug or labeled compound, and optionally at least one additional lipid, where a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine guide includes a ligand that is attached to the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and where the derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 1,

and the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 3,

where R1, R2, R5and R6 each independently represents an acyl group, and m and p independently represent an integer from 1 to 10; and where the liposome does not include non-modified phosphatidylethanolamine or polyethylene glycol and guide where the ligand is not an intact antibody.

In other embodiments offers an unloaded liposomes comprising one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and optionally at least one additional lipid, where a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine guide includes a ligand that is attached to the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine; and where the derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by the formula 1

and the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 3

where R1, R2, R5and R6each independently represents an acyl group, and m and p independently represent an integer from 1 to 10; and where the liposome does not include the unmodified phosphatide is ethanolamine, the polyethylene glycol, drug or labeled compound and guide where the ligand is not an intact antibody.

In certain embodiments of liposomes directional and non-loaded liposomes R1, R2, R5and R6are oleoyl or stearoyl, and m and p is 3. In certain embodiments guides ligand is transferrin. In specific embodiments, one or more phospholipids are a DMPC or DSPC, and there is at least one additional lipid representing cholesterol. In certain embodiments of liposomes directional and non-loaded liposomes R1, R2, R5and R6are oleoyl, m, and p is 3, one or more phospholipids are DMPC and additional lipid is a cholesterol.

In certain embodiments of liposomes directional and non-loaded liposomes m and p each independently is an integer from 2 to 4. In some embodiments, m and p are equal and are an integer from 2 to 4. In specific embodiments, m and p equal 3. In certain embodiments, R1, R2, R5and R6each independently represents oleoyl, stearoyl, Palmitoyl or myristoyl. In some embodiments, R1and R2are the same and R and R6are the same. In other embodiments, R1, R2, R5and R6are the same. In specific embodiments, R1, R2, R5and R6are oleoyl or stearoyl. In certain embodiments, R1, R2, R5and R6are oleoyl.

In additional embodiments offers a lipid mixture comprising a mixture of one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and optionally at least one additional lipid, where the derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 1

and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 2

and where R1, R2, R3and R4each independently represents an acyl group, m and n independently represent an integer from 1 to 10; and where the mixture includes a non-modified phosphatidylethanolamine or polyethylene glycol.

In certain embodiments the lipid mixtures m and n each independently is an integer from 2 to 4. In some embodiments m and n are equal and are an integer is 2 to 4. In specific embodiments m and n equal 3.

In certain embodiments the lipid mixtures R1, R2, R3and R4each independently represents oleoyl, stearoyl, Palmitoyl or myristoyl. In some embodiments, R1and R2are the same and R3and R4are the same. In specific embodiments, R1, R2, R3and R4are the same. In some embodiments, R1, R2, R3and R4are oleoyl or stearoyl. In certain embodiments m and n is equal to 3, where one or more phospholipids are a DMPC or DSPC, and there is at least one additional lipid, representing cholesterol. In certain embodiments the lipid mixtures R1, R2, R3and R4are oleoyl, m and n is 3, one or more phospholipids are DMPC and additional lipid is a cholesterol.

In additional embodiments offers a lipid mixture comprising a mixture of one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and optionally at least one additional lipid, where the derived N-(ω)-dicarbonate the acid and phosphatidylethanolamine represented by formula 1

and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine guide includes a ligand which is connected with the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine; and where the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 3

where R1, R2, R5and R6each independently represents an acyl group, m and p independently represent an integer from 1 to 10; and where the mixture includes a non-modified phosphatidylethanolamine or polyethylene glycol, and guide where the ligand is not an intact antibody.

In certain embodiments the lipid mixtures, where there is a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, m and p each independently is an integer from 2 to 4. In specific embodiments, m and p are equal and are an integer from 2 to 4. In specific embodiments, m and p equal to 3.

In certain embodiments the lipid mixtures, where there is a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, R1, R2, R5and R6each independently represents oleoyl, stearo is, the Palmitoyl or myristoyl. In some embodiments, where R1and R2are the same, and R5and R6are the same. In additional embodiments, R1, R2, R5and R6are the same. In some embodiments, R1, R2, R5and R6are oleoyl or stearoyl. In some embodiments, R1, R2, R5and R6are oleoyl or stearoyl, m, and p is 3, one or more phospholipids are a DMPC or DSPC, at least one additional lipid is a cholesterol, and directing ligand is transferrin. In certain embodiments the lipid mixtures R1, R2, R5and R6are oleoyl, m, and p is 3, one or more phospholipids are a DMPC, additional lipid is a cholesterol, and directing ligand is transferrin.

In certain embodiments of the offers containing liposome composition comprising a liposome comprising one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and optionally at least one additional lipid, where the derived N-(ω)-dicarboxylic acid and phosphatidylserine the mine is represented by formula 1

and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 2

where R1, R2, R3and R4each independently represents an acyl group, m and n independently represent an integer from 1 to 10; and where the composition does not include non-modified phosphatidylethanolamine or polyethylene glycol.

In some embodiments containing liposomes compositions m and n each independently is an integer from 2 to 4. In certain embodiments m and n are equal and are an integer from 2 to 4. In specific embodiments m and n equal 3.

In some embodiments containing liposomes compositions R1, R2, R3and R4each independently represents oleoyl, stearoyl, Palmitoyl or myristoyl. In specific embodiments, R1and R2are the same, and R3and R4are the same. In some embodiments, R1, R2, R3and R4are the same. In certain embodiments, R1, R2, R3and R4are oleoyl or stearoyl. In some embodiments, R1, R2, R3and R4are oleoyl or stearoyl, and m and n equal to 3, and one or more phospholipids are DMC, DSPC, POPC or DPP. In certain embodiments, R1, R2, R3and R4are oleoyl, m and n is 3, one or more phospholipids are a DMPC, and additional lipid is a cholesterol.

In additional embodiments containing liposome compositions are available containing liposome composition comprising a liposome comprising one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and optionally at least one additional lipid, where the derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 1

and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine guide includes a ligand that is attached to the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine; and where the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 3

and where R1, R2, R5and R6each independently represents an acyl group, m and p independently represent an integer from 1 to 10; and where the composition does not include named fitsirovannye the phosphatidylethanolamine or polyethylene glycol, and guide where the ligand is not an intact antibody.

In certain embodiments containing liposomes compositions where there is a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, m and p each independently is an integer from 2 to 4. In specific embodiments, m and p are equal and are an integer from 2 to 4. In some embodiments, m and p equal to 3.

In certain embodiments containing liposomes compositions where there is a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, R1, R2, R5and R6each independently represents oleoyl, stearoyl, Palmitoyl or myristoyl. In specific embodiments, R1and R2are the same, and R5and R6are the same. In certain embodiments, R1, R2, R5and R6are the same. In some embodiments, R1, R2, R5and R6are oleoyl or stearoyl. In specific embodiments, R1, R2, R5and R6are oleoyl or stearoyl, one or more phospholipids are a DMPC or DSPC, at least one additional lipid is a cholesterol, and directing ligand is TRANS is Erin. In certain embodiments, R1, R2, R5and R6are oleoyl, m, and p is 3, one or more phospholipids are a DMPC, additional lipid is a cholesterol, and directing ligand is transferrin.

In additional embodiments containing liposome compositions include medicinal substance. In certain embodiments, one or more phospholipids are a DMPC or DSPC, R1, R2and, when present, R5and R6are oleoyl or stearoyl, m, and, when present, p is 3, at least one additional lipid, when present, is a cholesterol drug is oxaliplatin, and guide the ligand, if present, represents transferrin. In certain embodiments, the composition further includes a sugar at a concentration of from about 1 to about 20% sugar (about./vol.). In certain embodiments, one or more phospholipids are a DMPC or DSPC, R1, R2and, when present, R5and R6are oleoyl, m, and, when present, p is 3, at least one additional lipid, when present, is a cholesterol drug substance is oxlip is Athyn, and guide the ligand, if present, represents transferrin.

In additional embodiments containing liposome compositions include labeled compound. In certain embodiments of the labeled compound comprises a radioisotope balance.

In certain embodiments of liposomes directional unloaded liposomes, lipid mixtures containing liposomes of the present compositions, at least one additional lipid. In specific embodiments, at least one additional lipid is a cholesterol or a derivative of cholesterol.

In specific embodiments of liposomes directional unloaded liposomes, lipid mixtures containing liposome compositions of one or more phospholipids are phosphatidylcholine, phosphatidic acid, phosphatidylserine or phosphatidylglycerol. In specific embodiments, one or more phospholipids are neutral phospholipids. In some embodiments, one or more phospholipids are phosphatidylcholine. In specific embodiments, the phosphatidylcholine comprises the residue of a saturated fatty acid. In certain embodiments, one or more phospholipids are a DMPC, DSPC, POPC or DPPC. In some of these embodiments there is at least one additional lipid. And is definitely one of them, at least one additional lipid is a cholesterol or a derivative of cholesterol. In specific embodiments included DMPC and cholesterol, DSPC and cholesterol, POPC and cholesterol or DPPC and cholesterol. In certain embodiments included DMPC and cholesterol.

In specific embodiments of liposomes directional unloaded liposomes, lipid mixtures containing liposomes compositions guide ligand when present, is aimed at the target cell. In some embodiments the guide ligand directed to a receptor on the cell surface of target cells. In specific embodiments the guide ligand is a transferrin, folic acid, hyaluronic acid, a sugar chain or a fragment of monoclonal antibodies. In other embodiments the guide ligand is a transferrin, folic acid, hyaluronic acid or a sugar chain. In specific embodiments the guide ligand is a transferrin, folic acid, hyaluronic acid or a sugar chain. In some embodiments the guide ligand is a transferrin. In specific embodiments, the transferrin is in a holo-form, but not in the APO-form. In some embodiments, the transferrin is in a holo-form.

In certain embodiments of liposomes directional and ringrose is related to the liposome mean diameter of the liposome is from about 50 nm to about 250 nm. In others, the average diameter of the liposomes is from about 90 nm to about 200 nm.

In specific embodiments of liposomes directional and non-loaded liposomes Zeta-potential of liposomes is negative. In certain embodiments, the Zeta potential is from about -75 mV to approximately -90 mV. In others, the Zeta-potential is from about -80 mV to about -85 mV.

In certain embodiments containing liposome compositions, liposomes directional and non-loaded liposomes compositions additionally include the solution.

In specific embodiments of liposomes directional and containing liposome compositions of the present medicinal substance.

In specific embodiments of liposomes directional and containing liposome compositions of the drug is oxaliplatin. In certain embodiments, when the drug is oxaliplatin, guide the ligand is a transferrin. In certain embodiments, at least one additional lipid is present and is a cholesterol.

In certain embodiments of the medicinal substance is an anticancer agent. In specific embodiments of the medicinal substance is cytotoxic is some of the medicinal substance. In some embodiments of the medicinal substance is an inhibitor of topoisomerase I. In specific embodiments, the inhibitor of topoisomerase I is topotecan or irinotecan. In other embodiments the medicinal substance is a Vinca alkaloid. In specific embodiments, the Vinca alkaloid is vincristine, vinblastine, Finlepsin, windowsin, vinorelbine or vindesine. In some embodiments of the medicinal substance is a nucleic acid. In certain embodiments the nucleic acid is an antisense oligonucleotide or a ribozyme. In some embodiments of the medicinal substance is an alkylating agent. In specific embodiments of the medicinal substance is an taxanes. In other embodiments the medicinal substance is an antagonist of metabolism. In certain embodiments of the medicinal substance is an antitumor antibiotic. In some embodiments of the medicinal substance is a medicinal substance for the hormonal therapy. In some embodiments of the medicinal substance is a medicinal substance with a molecular target.

In some embodiments of liposomes directional and containing liposome compositions, in addition to the public the substance is a compound of platinum. In specific embodiments, the platinum compound is biltin, cisplatin, carboplatin, ormaplatin, oxaliplatin, triplatin, angloplats, lobaplatin or spiroplatin. In some embodiments, the platinum compound is oxaliplatin.

In some embodiments, where the platinum compound is a oxaliplatin, R1, R2, R5and R6are oleoyl or stearoyl, m and p is equal to 3, the guide ligand is transferrin, one or more phospholipids are a DMPC or DSPC and at least one additional lipid is present and is a cholesterol. In certain embodiments, where the drug is oxaliplatin, R1, R2, R5and R6are oleoyl, m and p is equal to 3, the guide ligand is transferrin, one or more phospholipids are DMPC and at least one additional lipid is present and is a cholesterol. In specific embodiments the liposomes directional and containing liposome composition is free of other lipid components.

In some embodiments, where the drug is oxaliplatin, oxaliplatin dissolved in an aqueous solution of sugar selected from the group consisting of trehalose, maltose is, sucrose, mannose, lactose, mannitol, glycerin and dextrose. In certain embodiments sugar is in a concentration of from about 1 to about 20% sugar (about./vol.). In specific embodiments, the concentration of oxaliplatin is from about 0.1 mg/ml to about 25 mg/ml inside the liposomes. In other embodiments, the concentration of oxaliplatin is from about 0.5 mg/ml to about 10 mg/ml inside the liposomes. In some embodiments, the concentration of oxaliplatin is from about 0.5 mg/ml to about 3 mg/ml

In specific embodiments of liposomes directional and containing liposomes of the present compositions labeled compound. In certain embodiments of the labeled compound comprises a radioisotope balance. In specific embodiments radioisotope balance includes125I.

In specific embodiments of liposomes directional and containing liposomes compositions the concentration guide ligand included in the liposome is from about 1.0 mg/ml to about 3.0 mg/ml In the other concentration guide ligand included in the liposome is from about 1.0 mg/ml to about 2.5 mg/ml

In specific embodiments of liposomes directional and containing liposome compositions, where the drug is TES substance is present and represents oxaliplatin, guide the ligand is a transferrin. In specific embodiments, the transferrin is in a holo-form. In some embodiments, iron ion is in a concentration of from approximately 0.4 to approximately 3.0 µg/ml In other embodiments, iron ion is in a concentration of from about 0.4 to about 1.5 mg/ml

In specific embodiments of liposomes directional unloaded liposomes, lipid mixtures containing liposome compositions, liposomes, lipid mixtures or containing liposome composition does not include a cationic lipid. In specific embodiments the liposomes, lipid mixtures or containing liposome composition does not include anionic lipid. In some embodiments the liposomes, lipid, liposome, lipid mixtures or containing liposome composition does not include any anionic lipid or cationic lipid.

In specific embodiments of liposomes directional unloaded liposomes, lipid mixtures containing liposomes compositions compositions additionally include the solution. In certain embodiments the lipid mixture-free solution. In specific embodiments, the solution is an aqueous solution or a mixture of an aqueous solution and mixing with the water solvent.

In specific embodiments of liposomes directional unloaded liposomes, lipid mixtures ImageReady liposome compositions compositions additionally include sucrose.

In an additional aspect of the invention offers the pharmaceutical compositions described herein containing liposome compositions. Specific embodiments containing liposome compositions, liposomes directional and non-loaded liposomes include containing liposome compositions, liposomes directional or non-loaded liposomes, as described here, and one or more pharmaceutically suitable carriers, excipients, diluents, stabilizers or preservatives.

In another aspect of the invention provides kits comprising one described here contains the lipid composition. Certain embodiments containing liposome compositions, liposomes directional and non-loaded liposomes include containing liposome compositions, liposomes directional or non-loaded liposomes in the packaging and instructions for use.

In certain embodiments of the kits containing liposome compositions, liposomes directional or non-loaded liposomes, as described here, are contained in the first container, and one or more pharmaceutically acceptable carriers, excipients, diluents, stabilizers or preservatives contained in the second container.

In specific embodiments of the available sets, described here includes pharmaceutical formulations, packaging and toolbar the options for the application.

In another aspect of the invention provides methods of obtaining described herein lipid containing compositions.

In specific embodiments provides methods of obtaining liposomes directed action, as here described, which includes stages:

a) mixing one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and optionally at least one additional lipid from the lipid mixture;

b) adding a drug or labeled compound to the lipid mixture formed in stage (a);

c) formation of liposomes.

In additional embodiments of the method proposed stage (d) purification of liposomes from the stage (c). In specific embodiments of the medicinal substance in stage (b) before mixing is in aqueous solution. In certain embodiments of stage (c) comprises sonication or stirring. In some embodiments of stage (c) comprises extrusion.

In other embodiments provides methods of obtaining liposomes directed action, as here described, which includes stages:

a) mixing one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, operations ether is a derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and optionally at least one additional lipid from the lipid mixture;

where operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 2

where R3and R4each independently represents an acyl group, and n independently represents an integer from 1 to 10;

b) adding a drug or labeled compound to the lipid mixture formed in stage (a);

c) formation of liposomes; and

d) attaching a guide ligand to operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

In certain embodiments of the above method, the method also includes a step (e) purification of liposomes from the stage (d).

In specific embodiments of the medicinal substance in stage (b) before mixing is in aqueous solution. In some embodiments of stage (c) comprises extrusion. In specific embodiments of stage (c) includes stirring.

In certain embodiments provides methods of obtaining unloaded liposomes, as described here, which includes stages:

a) mixing one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, modified guiding factor derived N-(ω)-dicarboxylic what Islami and phosphatidylethanolamine, and optionally at least one additional lipid from the lipid mixture; and

b) the formation of liposomes.

In certain embodiments of the methods of obtaining the unloaded liposomes, the method further includes a step (c) purification of liposomes from stage (b).

In specific embodiments of stage (b) includes processing the ulrazvuk or mixing. In some embodiments of stage (b) comprises extrusion.

In other embodiments provides methods of obtaining unloaded liposomes, as described here, which includes stages:

a) mixing one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and optionally at least one additional lipid from the lipid mixture,

where operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 2

where R3and R4each independently represents an acyl group, and n independently represents an integer from 1 to 10;

b) formation of liposomes; and

c) attaching a guide ligand to operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine education modified upravlyayuschim factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

In certain embodiments of the methods of obtaining the unloaded liposomes, methods further include stage (d) purification of liposomes from the stage (c).

In specific embodiments of stage (b) includes processing the ulrazvuk or mixing. In some embodiments of stage (b) comprises extrusion.

In other embodiments provides methods of obtaining lipid containing compositions as described herein comprising a stage of mixing one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

In additional embodiments provides methods of obtaining lipid containing compositions, where there is at least one additional lipid, as here described, comprising a stage of mixing one or more phospholipids, at least one additional lipid-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

In other embodiments provides methods of obtaining lipid containing compositions as described herein comprising a stage of mixing one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified towards the shining factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

In additional embodiments provides methods of obtaining lipid containing compositions, where there is at least one additional lipid, as here described, comprising a stage of mixing one or more phospholipids, at least one additional lipid-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

There are also ways of getting described herein containing liposome compositions, incorporating the following stages:

a) mixing one or more phospholipids and its derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and, when present, operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine or modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and optional, when present, at least one additional lipid from the lipid mixture; and

b) adding a drug lipid mixture formed in stage (a); and

c) formation of liposomes.

In certain embodiments of the methods of obtaining various lipid containing compositions (liposomes directional unloaded liposomes containing liposomes comp is dispositions), where there are drug substance, drug substance is an aqueous solution. In certain embodiments of stage a) is carried out in the presence of an organic solvent. In some embodiments the aqueous solution additionally contains sugar. In certain embodiments the aqueous solution may also include miscible with water, an organic solvent.

In other embodiments provides methods of obtaining containing liposome compositions, incorporating the following stages:

a) mixing one or more phospholipids and its derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine and,

when present, operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine or modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and optional, when present, at least one additional lipid from the lipid mixture; and

b) adding a labeled compound to the lipid mixture formed in stage (a);

c) formation of liposomes.

In certain embodiments of the methods of obtaining various lipid containing compositions (liposomes directional unloaded liposomes containing liposomes compositions), which is labeled the connection, the connection is labeled in water rest the re. In certain embodiments of stage a) is carried out in the presence of an organic solvent. In certain embodiments the aqueous solution may also include miscible with water, an organic solvent.

In certain embodiments also provides methods of obtaining containing liposome compositions, as described here, where containing liposome compositions include modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, which includes stages

a) mixing one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and optionally at least one additional lipid from the lipid mixture; and

b) adding a solvent to the mixture formed in stage (a) with the formation containing liposomes of the composition.

In specific embodiments of phase mixing (a) is carried out in the presence of an organic solvent. In specific embodiments the solvent at the stage (b) is an aqueous solution or a mixture of an aqueous solution and mixed with water, an organic solvent.

In certain embodiments of stage (b) includes processing the ulrazvuk or mixing. In some wapl is the use stage (b) comprises extrusion.

In specific embodiments of the methods of obtaining lipid containing compositions on the stage (a) is present at least one additional lipid.

In some embodiments of the methods of obtaining lipid containing compositions on the stage (a) is present operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

In certain embodiments of the methods of obtaining lipid containing compositions on the stage (a) is a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

In an additional aspect of the invention provides methods of treatment or diagnosis with application described herein lipid containing compositions.

In specific embodiments provides methods of treatment of cancer, comprising a) introduction liposomes directed action, as described here, the needy in this individual in amounts effective for the treatment of cancer, where the liposome directional incorporates a medicinal substance and the medicinal substance is an anticancer agent.

In certain embodiments of the method of treatment or diagnosis of the individual is a mammal. In specific embodiments the individual is the man.

In certain embodiments of methods of treating cancer is a breast W is Lesa stomach, colon, colorectal cancer, pancreatic cancer, not small cell lung cancer, small cell lung cancer, brain cancer, liver cancer, kidney cancer, prostate cancer, bladder cancer, ovarian cancer or hematological malignant diseases.

In some embodiments of the methods of treatment stage (a) is carried out before, simultaneously with or after combined therapy for cancer of different modalities. In specific embodiments, the combination treatment of cancer of different modalities include chemotherapy, radiation or surgery.

In specific embodiments of the methods of treatment stage (a) is carried out before, simultaneously with or after adjuvant therapy of cancer. In specific embodiments adjuvant cancer therapy involves the introduction of one or more agents to reduce hair loss, vomiting, immunosuppression, nausea, diarrhea, rash, sensory disorders, anemia, weakness, disease or syndrome hands and feet. In some embodiments of stage (a) is performed before, simultaneously with or after administration of one or more additional anti-cancer agents. In certain embodiments one or more additional anti-cancer agents include 5-fluorouracil, leucovorin, capecitabine, UFT/LV (tegafur-uracil and leucovorin), irinotecan, antibody against EGFR, anti-VEGF antibody, tyrosine kinase inhibitor, or their combination is.

In some embodiments of the methods of treatment of the liposomes directional administered by parenteral administration. In specific embodiments parenteral administration is by injection or intravenous infusion.

There are also methods of diagnosis, which includes stages:

a) the introduction of liposomes directed action, as described here, the needy in this individual in the amount effective for detection, where the liposome directed action includes labeled compound; and

b) detection of labeled compounds.

In additional embodiments of the diagnostic methods, the methods further include stage (c) comparing the level of detected labeled compounds with the number of labeled compounds defined in the previous point in time.

In additional embodiments of methods of diagnosis of stage (b) includes determining using a gamma-counter.

In an additional aspect of the invention offers a modified transferrin derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, where the derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 3

where R5and R6each independently represents an acyl group, and p represents an integer from 1 to 10, and transferrin attached to the derived N-ω)-dicarboxylic acid and phosphatidylethanolamine.

In certain embodiments of the modified transferrin-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine p is an integer from 2 to 4. In specific embodiments, p is 3.

In some embodiments of the modified transferrin-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine R5and R6each independently represents oleoyl, stearoyl, Palmitoyl or myristoyl. In specific embodiments, R5and R6are the same. In some embodiments, R5and R6are oleoyl or stearoyl. In certain embodiments, R5and R6are oleoyl and p is 3.

In an additional aspect also provides pharmaceutical compositions comprising modified transferrin derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, as here described, and one or more pharmaceutically acceptable carriers, excipients, diluents, stabilizers or preservatives.

In certain embodiments offers a lipid mixture comprising a mixture of at least two different neutral lipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine, where the derived N-(ω)-dicarboxylic acid and FOSFA is tadalafilonline represented by formula 1

and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 2

where R1, R2, R3and R4each independently represents an acyl group, m and n independently represent an integer from 1 to 10; and where the mixture includes a non-modified phosphatidylethanolamine or polyethylene glycol.

In certain embodiments of the offers containing liposome composition comprising at least two different neutral lipid-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine and encapsulated drug or labeled compound, where the derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 1

and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 2

where R1, R2, R3and R4each independently represents an acyl group; m and n independently represent an integer from 1 to 10; and where the mixture includes a non-modified phosphatidylethanolamine or polyethylene glycol.

In concrete the x incarnations containing liposomes compositions and lipid mixtures m and n each independently is an integer from 2 to 4. In certain embodiments m and n are equal and are an integer from 2 to 4. In other embodiments m and n equal 3.

In some embodiments containing liposomes compositions and lipid mixtures R1, R2, R3and R4each independently represents oleoyl, stearoyl, Palmitoyl or myristoyl. In some embodiments, R1and R2are the same, and R3and R4are the same. In specific embodiments, R1, R2, R3and R4are the same. In some embodiments, R1, R2, R3and R4are oleoyl.

In specific embodiments containing liposomes compositions and lipid mixtures the molar ratio of neutral lipids:derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine:operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine is approximately 95:4:1.

In some embodiments containing liposomes compositions and lipid mixtures, where neutral lipids represent DMPC and cholesterol, the molar ratio of DMPC:cholesterol:(a derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine+operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine) is 50:45:5. In certain of these embodiments derived N-(ω)-dicarboxylic acid and is fosfatidiletanolamina is a NG-DOPE and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an NHS-NG-DOPE.

In certain embodiments provides liposomes directed action, including at least two different neutral lipid-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and encapsulated drug or labeled compound, where a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine guide includes a ligand that is attached to the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine; and where the derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 1

and the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 3

where R1, R2, R5and R6each independently represents an acyl group, and m and p independently represent an integer from 1 to 10; and where the liposome does not include non-modified phosphatidylethanolamine or polyethylene glycol, and guide where the ligand is not intact antibody.

In specific embodiments provides liposomes directed action, including neutral phosphatidylcholine, cholesterol sludge is derived cholesterol, the derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine modified transferrin-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and encapsulated oxaliplatin, where the modified transferrin-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine includes transferrin, attached through amide linkages carboxylic acid to the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine; and where the derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 1

and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 2, and the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 3

where R1, R2, R5and R6each independently represents an acyl group, and m and p independently represent an integer from 1 to 10; and where the liposome does not include non-modified phosphatidylethanolamine or polyethylene glycol. In certain embodiments, the liposome directional essentially free from EDC and/or DCC.

In certain embodiments of liposomes directional m and p each independently is an integer from 2 to 4. In some embodiments m and of the equal and are an integer from 2 to 4. In specific embodiments, m and p equal to 3.

In some embodiments of liposomes directional R1, R2, R5and R6each independently represents oleoyl, stearoyl, Palmitoyl or myristoyl. In certain embodiments, R1and R2are the same and R5and R6are the same. In specific embodiments, R1, R2, R5and R6are the same. In some embodiments, R1, R2, R3and R4are oleoyl or stearoyl. In certain embodiments, R1, R2, R3and R4are oleoyl.

In a specific embodiment of liposomes directional guide ligand directed to the target cell. In specific embodiments the guide ligand directed to a receptor on the cell surface of target cells. In some embodiments the guide ligand is a transferrin, folic acid, hyaluronic acid, a sugar chain or a fragment of monoclonal antibodies. In certain embodiments the guide ligand is a transferrin, folic acid, hyaluronic acid or a sugar chain. In specific embodiments the guide ligand is a transferrin. In some of these embodiments, the transferrin is in a holo-form, but not up in the form. In other embodiments, the transferrin is in the APO-form.

In certain embodiments the lipid mixtures containing liposomes compositions and liposomes directed action, the compositions do not include anionic lipid. In some embodiments, the compositions do not include a cationic lipid. In some embodiments, the compositions do not include a cationic lipid or anionic lipid. In specific embodiments, the composition does not include phosphatidylglycerol or its derivative. In specific embodiments, the compositions do not include egg phosphatidylcholine.

In some embodiments the lipid mixtures containing liposomes compositions and liposomes, which are focused on at least two different neutral lipid is one or more phospholipids and cholesterol or a derivative of cholesterol. In some embodiments, at least one of the at least two different neutral lipid is a phospholipid. In certain embodiments the lipid mixtures containing liposomes compositions and liposomes, which are focused on at least two different neutral lipid are phosphatidylcholine and cholesterol. In specific embodiments, one of at least two different neutral lipids is a DMPC, DSPC or DPPC. In some embodiments one of the at least two different neutral lipids p is ecstasy a cholesterol or a derivative of cholesterol. In specific embodiments, at least two different neutral lipid represent DMPC and cholesterol, DSPC and cholesterol or DPPC and cholesterol. In specific embodiments, at least two different neutral lipid represent DMPC and cholesterol.

In certain embodiments of liposomes directional average diameter of the liposomes is from approximately 50 nm to approximately 250 nm. In certain embodiments, the average diameter of the liposomes is from about 90 nm to about 200 nm. In specific embodiments, the average diameter of the liposomes is from about 100 nm to about 140 nm.

In certain embodiments of liposomes directional Zeta-potential of liposomes is negative. In specific embodiments, the Zeta potential is from about -75 mV to approximately -90 mV. In some embodiments, the Zeta potential is from about -80 mV to about -85 mV.

In some embodiments the lipid mixtures containing liposomes compositions and liposomes directional derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an NG-DOPE (where NG-DOPE is equivalent to the fact that R1and R2are OleOle, and m is 3) and, when present, operations ether derivative N-(ω)-dicarboxylic key is lots and phosphatidylethanolamine is an NHS-NG-DOPE (where NHS-NG-DOPE is equivalent in what R3and R4are OleOle, and n equals 3) or, when present, a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a TF-NG-DOPE (where TF-NG-DOPE is equivalent to the fact that R5and R6are OleOle, and p is equal to 3).

In some embodiments containing liposomes compositions and liposomes directional compositions additionally include the solution.

In certain embodiments of liposomes directional molar ratio of neutral lipids:derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine:a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is approximately 95:4:1.

In certain embodiments of liposomes directional molar ratio of DMPC:cholesterol:(a derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine + modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine) is approximately 50:45:5.

In specific embodiments containing liposomes compositions and liposomes directed action is labeled compound. In certain embodiments of the labeled compound comprises a radioisotope balance. In specific embodiments of the labeled compound include125/sup> I.

In some embodiments containing liposomes compositions and liposomes directed action is the medicinal substance. In specific embodiments of the medicinal substance is an anticancer agent. In some embodiments of the medicinal substance is a cytotoxic agent. In certain embodiments of the medicinal substance is an inhibitor of topoisomerase I. In specific embodiments, the inhibitor of topoisomerase I is topotecan or irinotecan. In other embodiments the medicinal substance is a Vinca alkaloid. In some embodiments, the Vinca alkaloid is vincristine, vinblastine, Finlepsin, windowsin, vinorelbine or vindesine. In other embodiments the medicinal substance is a nucleic acid. In some of these embodiments the nucleic acid is an antisense oligonucleotide or a ribozyme. In specific embodiments of the medicinal substance is a compound of platinum. In certain embodiments, the platinum compound is biltin, cisplatin, carboplatin, ormaplatin, oxaliplatin, triplatin, angloplats, lobaplatin or spiroplatin. In specific embodiments, the platinum compound is oxaliplatin. In some embodiments l is the drug substance is an alkylating agent. In specific embodiments of the medicinal substance is an taxanes. In other embodiments the medicinal substance is an antagonist of metabolism. In certain embodiments of the medicinal substance is an antitumor antibiotic. In some embodiments of the medicinal substance is a medicinal substance for the hormonal therapy. In specific embodiments of the medicinal substance is a medicinal substance with a molecular target.

In some embodiments, where there is oxaliplatin, oxaliplatin dissolved in an aqueous solution of sugar selected from the group consisting of trehalose, maltose, sucrose, lactose, mannose, mannitol, glycerin and dextrose. In certain embodiments sugar is in a concentration of from about 1 to about 20% sugar (about./vol.). In some embodiments, the concentration of oxaliplatin is from about 0.1 mg/ml to about 25 mg/ml inside the liposomes. In other embodiments, the concentration of oxaliplatin is from about 0.5 mg/ml to about 10 mg/ml inside the liposomes. In some embodiments, the concentration of oxaliplatin is from about 0.5 mg/ml to about 3 mg/ml

In specific embodiments where the modified healthy lifestyles is the missing factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, concentration guide ligand included in the liposome is from about 1.0 mg/ml to about 3.0 mg/ml In certain embodiments, the concentration of the guide ligand included in the liposome is from about 1.0 mg/ml to about 2.5 mg/ml

In certain embodiments where the modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, guide the ligand is a transferrin. In some embodiments, the transferrin is in a holo-form, but not in the APO-form. In certain embodiments, the transferrin is in a holo-form. In some embodiments, iron ion is in a concentration of from approximately 0.4 to approximately 3.0 µg/ml In other embodiments, iron ion is in a concentration of from about 0.4 to about 1.5 mg/ml

In some embodiments the lipid mixtures containing liposomes compositions and liposomes directional compositions free from lipid components, other than two different neutral lipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine. In other embodiments, the compositions are free from lipid components other than phosphatidyl is Lina, cholesterol or a derivative of cholesterol-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified with transferrin-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

In another aspect offers pharmaceutical compositions comprising liposomes directional or containing liposome compositions, as described herein and one or more pharmaceutically acceptable carriers, excipients, diluents, stabilizers or preservatives.

In another aspect of the invention provides kits containing one or more described herein lipid mixtures containing lipid compositions or liposomes directional, packaging and instructions for use.

In certain embodiments, the kit includes liposomes directed action. In specific embodiments the liposomes directed action are contained in the first container, and one or more pharmaceutically acceptable carriers, excipients, diluents, stabilizers or preservatives contained in the second container.

If not noted otherwise, lipid containing compositions as described herein are intended for use in the methods of treatment and diagnosis, as described here, and can be described here is included in the pharmaceutical compositions and kits. Described herein lipid containing compositions (VK is UCA lipid mixture, containing liposomes of the composition), liposomes (including liposomes targeted action, unloaded liposomes and so on) may, unless otherwise stated, be obtained using the methods described here received.

In another aspect provides methods of obtaining described herein lipid containing compositions, comprising a stage of mixing at least two different neutral lipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

In certain embodiments provides methods of obtaining described herein containing liposome compositions, incorporating the following stages:

a) mixing at least two different neutral lipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine with the formation of lipid mixtures;

b) adding a drug lipid mixture formed in stage (a); and

c) formation of liposomes.

In certain embodiments, the stage of mixing (a) is performed in the presence of an organic solvent.

In some embodiments of the medicinal substance in stage (b) before mixing is in aqueous solution.

In certain embodiments of stage (c) including the otherwise sonication or stirring. In specific embodiments of stage (c) comprises extrusion.

In additional embodiments provides methods of obtaining described here liposomes directed action, which includes stages:

a) mixing at least two different neutral lipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine with the formation of lipid mixtures;

b) adding a drug or labeled compound to the lipid mixture formed in stage (a);

c) formation of liposomes; and

d) attaching a guide ligand to operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

In certain embodiments of the above method, the method further includes a step (e) purification of liposomes from the stage (d). In specific embodiments of the medicinal substance in stage (b) before mixing is in aqueous solution. In certain embodiments of stage (c) comprises sonication or stirring. In some embodiments of stage (c) comprises extrusion.

There are also additional ways to obtain liposomes directed action, which includes stages:

a) mixing of phosphatidylcholine, cholesterol or a derivative of cholesterol-derived N-(ω)-dick is oil acid and phosphatidylethanolamine with the formation of the lipid mixture;

b) adding oxaliplatin to the lipid mixture formed in stage (a);

c) formation of liposomes; and

d) functionalization part derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine education operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine; and

e) attaching transferrin to operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

In certain embodiments the method further includes a step (f) purification of liposomes from the stage (e).

In specific embodiments of the method of the medicinal substance in stage (b) before mixing is in aqueous solution.

In some embodiments of the method stage (c) comprises sonication or stirring.

In an additional aspect of the invention features the use of lipid containing compositions (including liposomes targeted actions) and their compounds, as described here, to get drugs. In particular, for obtaining a medicinal product for use in the treatment or diagnosis of conditions as described here. Further, their pharmaceutical compositions described herein in various ways, also intended for use in preparation of medicines for use in the treatment and diagnosis of conditions and in accordance with the description of the different ways here, if not stated otherwise.

In an additional aspect of the invention provides methods of treating cancer, including stage a) introduction liposomes directed action, as described here, the needy in this individual in amounts effective for the treatment of cancer, where the medicinal substance is an anticancer agent.

In some embodiments the individual is a mammal. In specific embodiments the individual is the man.

In certain embodiments the cancer is a cancer of the breast, stomach, colon, colorectal cancer, pancreatic cancer, not small cell lung cancer, small cell lung cancer, brain cancer, liver cancer, kidney cancer, prostate cancer, bladder cancer, ovarian cancer or hematological malignant diseases.

In some embodiments of the methods of treatment stage (a) is carried out before, simultaneously with or after combined therapy for cancer of different modalities. In specific embodiments, the combination treatment of cancer of different modalities include chemotherapy, radiation or surgery.

In some embodiments of the methods of treatment stage (a) is carried out before, simultaneously with or after adjuvant therapy of cancer. In specific embodiments adjuvant cancer therapy involves the introduction of one or more agents to reduce the Oia hair loss, vomiting, immunosuppression, nausea, diarrhea, rash, sensory disorders, anemia, weakness, disease or syndrome hands and feet. In certain embodiments of stage (a) is performed before, simultaneously with or after administration of one or more additional anti-cancer agents. In specific embodiments, one or more additional anti-cancer agents include 5-fluorouracil, leucovorin, capecitabine, UFT/LV (tegafur-uracil and leucovorin), irinotecan, antibody against EGFR, anti-VEGF antibody, tyrosine kinase inhibitor, or combinations thereof.

In certain embodiments of the treatment methods liposomes directional administered by parenteral administration. In specific embodiments parenteral administration is by injection or intravenous infusion.

Brief description of figures

Figure 1 is a given schematic illustration of liposomes directed action.

Figure 2 is given a schematic representation of the active targeting of drug substances on tumor cells using liposomes directed action.

Figure 3 is given a schematic representation of the proposed mechanism of action of liposomes directional containing oxaliplatin.

Figure 4 presents a schematic representation of a method of obtaining A liposomes directed action.

Figure 5 presents a schematic of pic is BA obtain B liposomes directed action.

Figure 6 presents the cytotoxicity oxaliplatin on AsPC-1 cells at various concentrations oxaliplatin.

7 shows the number of transferrin receptors present on the cell surface of normal cells and cell lines of tumor origin.

On Fig presents the distribution by size mixtures containing liposomes obtained in example 6 were obtained using QELS; a) 1, B) Input 2, C) Input 3, D) 4, E) Input 5, F) Input 6.

Figure 9 presents the concentration of liposomes in the blood, where (♦) shows liposomes Tf-PEG, obtained in example 6, (■) shows liposomes Tf-NG-cells of the dspe:NG-cells of the dspe:DSPC:CH, obtained in example 5, and (▲) indicates liposomes Tf/PEG-NG-cells of the dspe, obtained in example 6.

Figure 10 presents the concentration of liposomes in cancer tissues, where (♦) shows liposomes Tf-PEG, obtained in example 6, (■) shows liposomes Tf-NG-cells of the dspe:NG-cells of the dspe:DSPC:CH, obtained in example 5, and (▲) indicates liposomes Tf/PEG-NG-cells of the dspe, obtained in example 6.

Figure 11 shows the accumulation in tumor tissue liposomes NG-PE obtained in example 13, after intravenous injection, where liposomes NGPE, encapsulated termineonline labeled125I was injected into mice native tumor colon 26. Data are presented as mean ± STD. off. (n=5). (□) 0 mol% (-);(<) 1 mol% (+) Tf-NG-DOPE. * The significance of differences from 0 mol% (-).

The piano is g presents the inhibitory effects of liposomes on tumor growth using sediment data about the growth of the tumor to the days after the initial impact, where (♦) shows liposomes Tf-PEG, obtained in example 9, (<) displays PEG-liposomes obtained in example 6 without Tf; (▲) indicates liposomes Tf-NG-cells of the dspe:NG-cells of the dspe:DSPC:CH, obtained in example 8; (○) shows liposomes NG-cells of the dspe:NG-cells of the dspe:DSPC:CH, obtained in example 8 without Tf; (*) indicates liposomes Tf/PEG-NG-cells of the dspe obtained in example 9; (●shows liposomes PEG-NG-cells of the dspe obtained in example 9 without Tf; (+) indicates the l-OHP solution; and (-) indicates no effect.

On Fig presents the effect of different concentrations of NG-PT(NG-cells of the dspe) percentage dose of medicinal substance found in the blood, where the concentration (% of total lipid) NG-cells of the dspe is following: (♦) 0%,(■)1%; (▲) 3%; (x) 6%; (○) 12%; and with the following lipids (●) MPB 6%; (+) PDP 6%.

On Fig presents a delay in the blood of liposomes with different linkers dicarboxylic acids with and without Tf; where (♦) Tf-NGPE; (<) Tf-NSPE; (▲) Tf-MPB; (x) NGPE (without Tf card); (*) MPB (without Tf card).

On Fig presents experimental analysis of liposomes using electrophoresis: lane 6 shows the (liposome transferrin-N-glutamyltranspeptidase (liposome Tf-NG-cells of the dspe-)); track 5 (liposome transferrin-polyethylene glycol-distearoylphosphatidylcholine (liposome Tf-PEG-cells of the dspe-)). Lanes 1-4 contain h-APO-Tf (240 ng), h-APO-Tf (120 ng), h-APO-Tf (60 ng), and h-APO-Tf (30 ng), respectively.

On Fig presents the number of transferrin associated with liposomes Tf-NGDSPE with (lane 5) or without (lane 4) non-NG-cells of the dspe, included in the liposome. Lanes 1-3 contain h-APO-Tf (400 ng), h-APO-Tf (200 ng) and h-APO-Tf (50 ng), respectively.

On Fig presents accumulation oxaliplatin in the blood after administration (■)liposomes NG-DOPe:tf-NG-DOPE:DMPC:CH at 5 mg/kg (●) liposomes Tf-PEG at 5 mg/kg

On Fig presents accumulation oxaliplatin in tumors of the colon 26 after injection (■) liposomes NG-DOPe:tf-NG-DOPE:DMPC:CH at 5 mg/kg (●) liposomes Tf-PEG at 5 mg/kg mice with tumors of the colon 26.

On Fig presents the antitumor effect of liposomes NG-DOPe:tf-NG-DOPE:DMPC:CH mice-media tumors of the colon 26 where (●) indicates the control media (9% sucrose); (▲) indicates the l-OHP solution, 5 mg/kg; (♦)indicates liposomes NG-DOPe:tf-NG-DOPE:DMPC:CH, 5 mg/kg (■) indicates liposomes Tf-PEG , 5 mg/kg

On Fig presents the antitumor effect of liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH mice carrier xenograft tumor HCT-116 colon of a person, where (●) indicates the control medium (300 mm (10,27%) sucrose); (○) indicates unloaded the liposome (without medicinal substances), (▲) indicates the l-OHP solution, 15 mg/kg; (♦) indicates liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH, 10 mg/kg (■) indicates liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH, 15 mg/kg All liposomes were injected on the exact body weight from the volume of injection 0,103 ml/10 g body weight.

On Fig presents the antitumor effect of liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH mice, nose and the residents of xenograft tumor HT-29 colon of man, where (●) indicates the control medium (300 mm (10,27%) sucrose; (▲) indicates liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH, 15 mg/kg (■) indicates liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH, 10 mg/kg (♦) indicates liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH, 6,7 mg/kg

On Fig presents the antitumor effect of liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH mice carrier xenograft tumor MKN45 human gastric, where (●) indicates the control medium (300 mm (10,27%) sucrose; (▲) indicates liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH, 15 mg/kg; (■) indicates liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH, 10 mg/kg and (♦indicates liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH, 6,7 mg/kg

On Fig presents the antitumor effect of liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH mice carrier xenograft tumor COLO 205 human (●) indicates the control media (9% sucrose); (▲) indicates the l-OHP solution, 5 mg/kg q4d × 3 (16 day), 10 mg/kg q2d × 2 (day 47), 2 mg/kg q2d × 6 (51 days); (♦)indicates liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH, 5 mg/kg, q4d × 3 (16 day), 10 mg/kg q2d × 2 (day 47), 2 mg/kg q2d × 6 (51 days); and (■) indicates liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH, 10 mg/kg q4d × 3 (16 day), 15 mg/kg q2d × 2 (day 47), 4 mg/kg q2d × 6 (51 days).

On Fig presents the pattern of Na-DDS-PAGE after recovery 2-mercaptoethanol, where lane 1 represents molecular weight markers, lane 2 is a holo-transferrin, lanes 3-5 represent liposomes NG-DOPe:Tf-NG-DOPE:DMPC:CH, and track 6 is a Tf-NG-DOE.

On Fig presents an example of an HPLC chromatogram of a suitable system.

Detailed description of the invention

Here are lipid containing compositions (including liposomes targeted action, unloaded liposomes containing liposomes of the composition, lipid mixtures and so on), and methods for their preparation and use are described here lipid containing compositions. Lipid containing compositions and liposomes, in particular, those offered here, suitable for the production of pharmaceutical compositions for use in the treatment or diagnosis of various conditions, including cancer. Compositions, including pharmaceutical compositions, provide a more effective diagnostics and treatment schemes with reduced side effects associated with the drug or labeled compound to be delivered to the individual. Increased efficiency and reduced adverse effects should increase therapeutic index of the drug compound and to provide the ability to successfully treat a variety of conditions, including cancer, and should also increase the efficiency and to reduce adverse effects associated with the diagnosis. The increased specificity of drug formulations when accompanied by the reduction of side effects should give therapeutic benefits more and dia is the azone being treated individuals, thus, saving or prolonging life and improving the quality of life of individuals in need of treatment. Increased the specificity of the compounds of labelled compounds with a parallel reduction of side effects should increase the number of individuals who can be successful diagnosis, for example, can be resistant to a diagnostic composition, and also to improve accuracy (e.g., sensitivity, etc.) diagnosis, including the possibility of earlier diagnosis of conditions and more effective monitoring of disease severity (e.g., progression or regression with or without therapy).

Included in the compositions described herein are pharmaceutical compositions containing the lipid compositions. Described herein lipid containing compositions include, but are not limited to, liposomes that encapsulate a drug and labeled compounds, and can be used for treatment or diagnosis of disease or other conditions requiring treatment or diagnosis, including, for example, cancer (e.g. breast cancer, gastric, colorectal cancer and cancer of the colon).

When traditional anticancer agents (including cytotoxic) is injected, the entire body is subjected to the action of medicinal substance, not selectively. This can be R the d adverse reactions, cancer is not a target, and/or the effect of the medicinal substance can be lost in the process of its circulation. Encapsulation of drug substances in the composition of the liposomes before the introduction can lead to one or more advantages, including the reduction of side(s) of effect(s) of medicinal substance in a normal cell, the protection of medicinal substance before it reaches pathological target cells when the medicinal substance can be unstable, the prolonged presence of the drug in the circulation system for delivery to pathological cells, and/or acceleration of drug delivery to pathological the target cell. A more specific target of a drug and a reduction in the loss of medicinal substance for capturing the RES is also an advantage in reducing the amount of drug that must be entered and thereby reduces the cost of therapy and also has other benefits described here.

Similar, many labeled compounds have side effects and/or destroyed in the time interval between introduction and diagnosis (e.g., the time during which the diagnosis is carried out technically, for example, detection of radioisotopes, magnetic resonance imaging, ultrasound, and so on). The inclusion m the black of the compounds described herein contain lipid composition should increase the efficiency of labeled compounds such as, for example, threshold detection, which can be achieved with smaller doses labeled compounds, the reduction of side effects of the agent, and/or expansion of the time window, which can be carried out diagnostics.

Lipid containing compositions also include lipids, modified guide ligands (for example, liposomes directional unloaded liposomes containing liposomes of the composition, the lipid mixture) or other derivatives. For example, liposomal composition comprising a guide factor (for example, transferrin etc) and modified lipids (for example, derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine), designed to improve security effectiveness of anticancer agents (e.g., oxaliplatin) due to the prolongation of the circulation time of the drug substance in plasma (compared with the medicinal substance is injected only in solution) and due to tumor cell receptors specific for guiding factor. This improved bioavailability and focus on the tumor should lead to enhanced security and increased antitumor activity and, consequently, a higher probability of effective treatment needs of individuals while reducing side effects associated with many of Lech is the only substances especially with the severe side effects associated with most anti-cancer agents. Similar such modification and guides the factors can also be applied effectively in a specific place (e.g., types of tumors, organs, tissue, etc. to deliver the labeled compounds.

In the invention are also modified with transferrin derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, which can be applied in this lipid containing compositions and their compositions.

Lipid containing compositions described here including liposomes, can be obtained using the methods described here, as well as ways of obtaining liposomes, well-known specialist in the art and suitable from the point of view of the guidance offered in the present description. Unless otherwise specified herein described liposomes containing liposome compositions can be included without limitation in the pharmaceutical compositions and/or kits, including pharmaceutical compositions and/or kits as described here, and additionally, those that should be obvious to a person skilled in the art and suitable from the point of view of the guidance offered in the present description. Similar to liposomes containing liposomes compositions and pharmaceutical compositions comprising liposomes containing lipo the ohms of the composition, can be applied without limitations, unless otherwise specified, the methods of treatment or diagnosis that is compatible with the description offered in this specification, and in accordance with the practice of professionals in the art and suitable from the point of view offered here are instructions.

Example liposomes directed action, including drugs (oxaliplatin)is presented schematically in Fig. 1. Suggested mechanisms of capture and mode of action of liposomes directional offered in Fig. 2 and 3. Used here, the term "liposome directional" refers generally to the liposome components, including at least one or more phospholipid(s), NωPE, TF-NωPE, and which also includes a drug or labeled compound, as described here. Each of these components, as described in the description, without limitation, may be incorporated into liposomes directed action of the present invention without leaving the scope of the presented instructions. It is noted that the unloaded liposomes described herein in great detail, can also be "designed"so that they can include modified Tf NωPEs, but they usually do not include drug or labeled compound.

Derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine

About vannie here lipid containing compositions include, at least one derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine in accordance with formula 1, below:

where R1and R2independently represent an acyl group, and m represents an integer from 1 to 10.

Used here, the term "derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine and related compounds are derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine covered by formula 1, as presented here. Similar abbreviation NωPE is used here to denote derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine covered by the formula 1 (e.g., Nω-DOPE, NωDSPE, NG-DOPE and so on) and, for example, NG-DOPE refers to N-getarilformazanami(s) of formula 1, unless otherwise specified.

It is implied that only described here phosphatidylethanolamine(s)included in the lipid containing compositions (including liposomes targeted action), are derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 1 or operations esters of derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 2 or a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 3, as described here in more detail below. As a result, the initial herein, the term "non-modified phosphatidylethanolamine and related compounds are phosphatidylethanolamine, semi-synthetic(Kim) to phosphatidylethanolamine(us), synthetic(Kim) to phosphatidylethanolamine(us) and/or their derivatives, which are not covered by formula 1, formula 2 or formula 3.

A variety of acyl groups represented by R1and R2can be used in the formula 1, as is well known to a person skilled in the technical field.

In some embodiments the acyl group of a saturated or unsaturated aliphatic carboxylic acids having 12-22 carbon atoms. Examples of acyl groups include, but are not limited to, acyl groups derived from lauric acid, tridecanoic acid, myristic acid, pentadecanoic acid, palmitic acid, margaric acid, stearic acid, nonadecanoic acid, arachnid acid, heneicosanol acid, beganovi acid, 2-lauraleedavis acid, 5-lauraleedavis acid, 11-lauraleedavis acid, 5-myristoleate acid, myristoleate acid, 2-palmitoleic acid, 7-palmitoleic acid, CIS-9-palmitoleic acid, TRANS-9-palmitoleic acid, petroselinic acid, Petroselinum acid, oleic acid, elaidic acid, vaccinology acid, gondolas acid, TRANS-gondolas acid, erucic acid, linoleic acid, linoleinovoy acid, α-e is stearinovoi acid, β-oleostearin acid, linolenic acid, pseudoallescheria acid, arachidonic acid, eicosapentaenoic acid or docosahexaenoic acid.

In certain embodiments the acyl group of a saturated or unsaturated aliphatic carboxylic acids having 14-18 carbon atoms. Examples of acyl groups of this type include, but are not limited to, those which are derived from oleic acid (18 carbons), palmitic acid (16 carbons), stearic acid (18 carbons) or myristic acid (14 carbons). As is known to experts in the art, the appropriate acyl groups are oleoyl, Palmitoyl, stearoyl and myristoyl respectively.

In other embodiments acyl group come from saturated or unsaturated aliphatic carboxylic acids having 14-18, 14-20, 14-22, 16-18, 16-20, 16-22, 18-20, 18-22, 12, 14, 16, 18, 20 or 22 carbon. In certain embodiments acyl group come from saturated or unsaturated aliphatic carboxylic acids having even more carbon.

In some embodiments acyl group come from oleic acid (oleoyl), stearic acid (stearoyl), palmitic acid (Palmitoyl), linoleic acid (linoleoyl, 18 carbons) or ministerului acid (myristoyl). In other UXO is ameneh acyl group come from oleic acid (oleoyl). In certain embodiments acyl group come from stearic acid (stearoyl). In some embodiments acyl group come from palmitic acid (Palmitoyl). In other embodiments acyl group come from ministerului acid (myristoyl).

In some embodiments acyl group come from saturated aliphatic carboxylic acids, such as, but not limited to, palmitic acid (16 carbons), stearic acid (18 carbons) or maistrova acid (14 carbons).

In other embodiments acyl group come from unsaturated aliphatic carboxylic acids, such as, but not limited to, oleic acid (oleoyl, 18 carbons), linoleic acid (linoleoyl, 18 carbons) or linolenic acid (linolenic, 18 carbons). In some embodiments the acyl group is from linoleic acid.

In certain embodiments m is 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In other embodiments m is an integer from 1-8, 1-6, 1-5, 1-7, 1-3, 1-2, 2-8, 2-6, 2-5, 2-4, 2-3, 3-4, 3-5, or 3-6. In some embodiments m is an integer from 2-4. In other embodiments m is 1, 2 or 3.

In certain embodiments m is an integer from 1 to 4. As known to a person skilled in the technical field, m=1 corresponds to a derivative of malonic acid phosphatidylethanolamine (PE), while m=2, 3, and 4 corresponds to a derivative of succinic acid, glutaric acid and adipic acid PE, respectively. In some embodiments m=3 (glutaric acid).

In certain embodiments, R1and R2are the same acyl group. In other embodiments, R1and R2represent different acyl groups. In certain embodiments, R1and R2are oleoyl, stearoyl, Palmitoyl or myristoyl. In some embodiments, R1and R2are oleoyl. In other embodiments, R1and R2represent stearoyl. In specific embodiments, R1and R2represent the Palmitoyl. In other embodiments, R1and R2are myristoyl.

In some embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 1 represents the N-patriciaarobertsaharris (NG-DOPE (i.e., where R1and R2are oleoyl and m is 3)). In other embodiments of this N-glutamyltranspeptidase (NG-cells of the dspe (i.e., where R1and R2represent stearoyl and m is 3)). In other embodiments of this N-glutamylcysteinylglycine (NG-DMPE (i.e., where R1and R2are myristoyl and m is 3)). In other embodiments of this N-glutamylcysteinylglycine (NG-DPPE (i.e., where R1and R2represent the Palmitoyl and m is 3)). In other embodiments of this N-succinylsulfathiazole (NS-cells of the dspe (i.e., where R1and R2represent stearoyl and m is 2)). In other embodiments of this N-administrativeregulatory (NA-cells of the dspe (i.e., where R1and R2represent stearoyl and m is equal to 4)). In certain embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 1 is a NG-DOPE or NG-cells of the dspe.

Preparation of derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine

Described here are derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine can be obtained by linking dicarboxylic acid with the amino group of phosphatidylethanolamine.

Phospholipids, including phosphatidylethanolamine and their derivatives, used for the purposes described here, should be of high purity and must be perfectly homogeneous. Known methods for producing high purity phospholipids include extraction of lipid from the buffer solution and purification using column chromatography. For example, methods of obtaining N-succiniciproducens described in international patent application, publication WO93/01828 (JPAH7-501316) and U.S. patent Nos. 5804552 and 5554728, the contents of which are incorporated herein in their entirety. These methods include obtaining clearance derived phospholipid from actionnow mixtures by chromatography of the reaction mixture on a column of silica gel 60. Dipalmitoylphosphatidylethanolamine (DPPE) reacts with the anhydride of succinic acid with triethylamine as a catalyst at room temperature under nitrogen gas for 16 hours.

Other methods of obtaining derivatives of N-(ω-carboxy)acylaminopenicillin described in the published patent application Japan JPA2001-261688, which include purification by separation of the liquid layer after the addition of buffer solution pH 3.5 and 7.5 to the reaction mixture, and fully included here by reference. In this case, the PE was introduced into the reaction with the anhydride of dicarboxylic acid with an alkaline triethylamine as catalyst at 4°C for 1 hour. This method may not work well for all derivatives of phosphatidylethanolamine.

DOPE (dioleoylphosphatidylcholine) can also be obtained commercially or obtained by methods known to the person skilled in the art. For example, briefly, lecithin (source substance API) can be chemically hydrolyzed with getting glycerophosphocholine, which is marked by deposition. The lipid is then acelerou using activated oleic acid and DOPC (dioleoylphosphatidylcholine) was isolated using column chromatography, normal phase and passed through an ion-exchange column for purification. DOPE can be obtained from DOPC through interaction with what trolamine using phospholipase D.

Phospholipid derivatives of N-(ω-carboxy)acylaminopenicillin can be also obtained in the same manner as described, for example, in U.S. patent No. 4534899, which is incorporated herein in its entirety. Briefly, the dicarboxylic anhydride is introduced into reaction with a phospholipid, such as phosphatidylethanolamine with obtaining derived dicarboxylic acid and phosphatidylethanolamine.

Operations ester derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine

Operations ester derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, as described here, represented by the following formula 2:

where R3and R4independently represent an acyl group, and n represents an integer from 1 to 10.

Used here, the term "operations esters derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and related compounds refers to the operations of the ester derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine covered by formula 2, as presented here. Similar abbreviation SuccNωPE can be used to denote the operations of ester derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine covered by the formula 2 (for example, SuccNω-DOPE, SuccNω-cells of the dspe, SuccNG-DOPE, etc. and, for example, NHS-NG-PE refers to succ needname ether, N-glutarylhistamine(s), formula 2 formed through reaction with the NHS, unless otherwise specified.

A wide variety of acyl groups, which are represented by R3and R4can be applied, as is well understood specialists in the art and as described above for R1and R2. If this is not specified otherwise, just assume that here the description of the acyl groups in relation to formula 1 (e.g., R1and R2) is applicable to the acyl groups in relation to formula 2 (for example, R3and R4). Including, in particular, the above description in the section entitled "Derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine".

In certain embodiments, R3and R4are the same acyl group. In other embodiments, R3and R4represent different acyl groups. In certain embodiments, R3and R4are oleoyl, stearoyl, Palmitoyl or myristoyl. In some embodiments, R3and R4are oleoyl. In other embodiments, R3and R4represent stearoyl. In certain embodiments, R3and R4represent the Palmitoyl. In other embodiments, R3and R4are myristoyl.

In certain embodiments n is 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In other in which plewniak n is an integer from 1-8, 1-6, 1-5, 1-7, 1-3, 1-2, 2-8, 2-6, 2-5, 2-4, 2-3, 3-4, 3-5, or 3-6. In some embodiments, n is an integer from 2-4. In other embodiments n is 1, 2 or 3.

In certain embodiments, n is an integer from 1 to 4. As known to a person skilled in the technical field, n=1 corresponds to a derivative of malonic acid phosphatidylethanolamine (PE), whereas n=2, 3 or 4 corresponds to a derivative of succinic acid, glutaric acid and adipic acid PE, respectively. In some embodiments, n=3 (glutaric acid).

In some embodiments operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 2 represents operations ether N-patriciaarobertsaharris (NG-DOPE). In other embodiments it operations ether N-glutamylcysteinylglycine (NG-cells of the dspe). In certain embodiments operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 2 represents operations ether NG-DOPE or NG-cells of the dspe.

In some embodiments operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 2 represents operations ether N-patriciaarobertsaharris (SuccNG-DOPE (i.e., where R3and R4are oleoyl and n is 3)). In other embodiments it operations ether N-glutamylcysteine the phosphatidylethanolamine (SuccNG-cells of the dspe (i.e., where R3and R4represent stearoyl and n is 3)). In other embodiments it operations ether N-glutamylcysteinylglycine (SuccNG-DMPE (i.e., where R3and R4are myristoyl and n is 3)). In other embodiments it operations ether N-glutamylcysteinylglycine (SuccNG-DPPE (i.e., where R3and R4represent the Palmitoyl and n is 3)). In other embodiments it operations ether N-succinylsulfathiazole (SuccNS-cells of the dspe (i.e., where R3and R4represent stearoyl and n is 2)). In other embodiments it operations ether N-administrativeregulatory (SuccNA-cells of the dspe (i.e., where R3and R4represent stearoyl and n is 4)). In certain embodiments operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 2 represents SuccNG-DOPE or SuccNG-cells of the dspe.

In some embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an Nω-DOPE and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a SuccNω-DOPE. In other embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an Nω-cells of the dspe and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine is predstavljaet a SuccNω-cells of the dspe. In some embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an Nω-DOPE and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a SuccNω-cells of the dspe. In certain other embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an Nω-cells of the dspe and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a SuccNω-DOPE. In certain embodiments operations ether can represent the NHS (for example, NHS-Nω-DOPE, NHS-Nω-cells of the dspe and so on).

In some embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an NG-DOPE and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a SuccNG-DOPE. In other embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an NG-cells of the dspe and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a SuccNG-cells of the dspe. In some embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an NG-DOPE and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a SuccNG-cells of the dspe. In certain other embodiments derived N-(ω)-dicarboxylic KIS is the notes and phosphatidylethanolamine is an NG-cells of the dspe and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a SuccNG-DOPE. In certain of these embodiments operations ether can represent the NHS (for example, NHS-NG-DOPE, NHS-NG-cells of the dspe and so on).

Receiving operations of the ester derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine

Described herein operations ester derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine can be obtained by modification described herein derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine obtained as is known in the art and described herein. Receiving operations of the ester derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine is also described in more detail below, including examples. From the point of view of the guidance offered in the present description, a person skilled in the field of technology should also be able to modify the methods described here.

The method of obtaining operations of the ester derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine of the present invention is as follows.

To 1 equivalent of the derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 2, as described here, add approximately 0.7 to 1.3 equivalent of the NHS, which is dissolved in an organic solvent, which has no active hydrogen. The mixture is then injected into the reaction with approximately 0.7 to 1.3 equivalent connection is carbodiimide at 0-50°C. for about 1-7 days.

Examples of organic solvents that do not have active hydrogen include, but are not limited to, esters (e.g. ethyl acetate, butyl acetate, etc.), aliphatic hydrocarbons (e.g. hexane, heptane and so on), aromatic hydrocarbons (e.g. toluene, xylene, etc.), halogenated hydrocarbons (e.g. chloroform, dichloromethane, dichloroethane, etc.), ethers (for example, THF, dioxane, diethyl ether, etc.), cyclic hydrocarbons (e.g. cyclohexane, etc.), DMF and DMSO. An organic solvent may also be dehydrated.

A variety of carbodiimide compounds may be used as long as the compounds have a carbodiimide group (-N=C=N-). For example, carbodiimide compounds that can be used include, but are not limited to, carbodiimide groups, such as N,N'-dicyclohexylcarbodiimide (DCC), N,N'-diisopropylcarbodiimide, N-ethyl-N'-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC), etc. In certain embodiments apply DCC. In other use EDC.

The above reaction can also be performed under conditions that minimize or eliminate the obtaining by-products. Undesirable side products include compounds of urea (for example, N,N'-dicyclohexylamine, N-ethyl-N'-(3-dimethylaminopropyl)urea, etc.), N-allir the bathrooms compounds of urea, carboxyanhydride compounds and compounds 5-oxazolone. Conditions and materials that do not contribute to or minimize the formation of by-products include 1) slow dissolving compounds, carbodiimide in an organic solvent, 2) carrying out the reaction at temperatures below 0°C to prevent heat generation in the reaction, etc. Other ways to optimize the reaction and to minimize the receipt of by-products should be clear to experts in the field of technology, especially from the point of view presented here of the instructions.

The organic solvent which can dissolve the compound carbodiimide, is the same as the organic solvent, which has no active hydrogen, as described above. The solvent used for dissolving carbodiimide, and an organic solvent without active hydrogens can be the same or different.

Over the course of the reaction can be monitored using thin-layer chromatography (TLC), high performance liquid chromatography (HPLC) and/or light scattering detectors. Other methods of tracking progress of the reaction must be known to specialists in this field of technology.

Cleaning can be performed using column chromatography on silica gel using a mixture of chloroform and methanol is. Additional purification methods must be known to specialists in this field of technology.

In the fully dehydrated organic solvents and in the absence of strong acidity or strong alkalization operations ester derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, as described here, is usually stable.

Modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine

Modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine include directing the ligand attached to the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, where the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 3

where R5and R6independently represent an acyl group, and p represents an integer from 1 to 10.

Used here, the term "modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and related compounds relates to derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine covered by formula 3 and modified guiding factor, as is offered here. Similar abbreviation Tf-NωPE can be used on the I denote the modified guiding factor derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine (for example, Tf-Nω-DOPE, Tf-Nω-cells of the dspe, Tf-NG-DOPE and so on) and, for example, Tf-NG-PE refers to directing ligand connected to the N-glutarylhistamine(s)of formula 3.

A wide variety of acyl groups, which are represented by R5and R6can be applied, as is well understood specialists in the art and as described above for R1and R2. If this is not specified otherwise, just assume that here the description of the acyl groups in relation to formula 1 (e.g., R1and R2) is applicable to the acyl groups in relation to formula 3 (for example, R5and R6). Including, in particular, the above description in the section entitled "Derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine".

In certain embodiments, R5and R6are the same acyl group. In other embodiments, R5and R6represent different acyl groups. In certain embodiments, R5and R6are oleoyl, stearoyl, Palmitoyl or myristoyl. In some embodiments, R5and R6are oleoyl. In other embodiments, R5and R6represent stearoyl. In certain embodiments, R5and R6represent the Palmitoyl. In other embodiments, R5and R6are meristo the L.

In certain embodiments p is 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In other embodiments, p is an integer from 1-8, 1-6, 1-5, 1-7, 1-3, 1-2, 2-8, 2-6, 2-5, 2-4, 2-3, 3-4, 3-5, or 3-6. In some embodiments p is an integer from 2-4. In other embodiments, p is 1, 2 or 3.

In certain embodiments, p is an integer from 1 to 4. As is well known to specialists in this field of technology, p=1 corresponds to a derivative of malonic acid phosphatidylethanolamine (PE), whereas p=2, 3 or 4 corresponds to a derivative of succinic acid, glutaric acid and adipic acid D, respectively. In some embodiments p=3 (glutaric acid).

In some embodiments, a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 3 represents a modified guiding factor N-patriciaarobertsaharris (TF-NG-DOPE). In other embodiments, a modified guiding factor N-glutamyltranspeptidase is (TF-NG-cells of the dspe). In certain embodiments, a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 3 represents a TF-NG-DOPE or TF-NG-cells of the dspe.

In some embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 3 represents the N-patriciaarobertsaharris the (NG-DOPE (i.e., where R5and R6are oleoyl and p is 3)). In other embodiments is N-glutamyltranspeptidase (NG-cells of the dspe (i.e., where R5and R6represent stearoyl and p is 3)). In other embodiments is N-glutamylcysteinylglycine (NG-DMPE (i.e., where R5and R6are myristoyl and p is 3)). In other embodiments is N-glutamylcysteinylglycine (NG-DPPE (i.e., where R5and R6represent the Palmitoyl and p is 3)). In other embodiments is N-succinylsulfathiazole (NS-cells of the dspe (i.e., where R5and R6represent stearoyl and p is 2)). In other embodiments is N-administrativeregulatory (NA-cells of the dspe (i.e., where R5and R6represent stearoyl and p is equal to 4)). In certain embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 3 represents a NG-DOPE or NG-cells of the dspe.

In certain embodiments the guide ligand is a transferrin (Tf), which is described in more detail below, and its derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine included in the modified transferrin-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine formula 3, as described here.

In some embodiments derived N-(ω)-dicarboxylic acid fosfatidiletanolamina represents Nω-DOPE and guide the ligand is a transferrin (Tf), and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a Tf-Nω-DOPE. In other embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an Nω-cells of the dspe and the guide ligand is a transferrin (Tf), and a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a Tf-Nω-cells of the dspe. In some embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an Nω-DOPE and guide the ligand is a transferrin (Tf), and a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a Tf-Nω-cells of the dspe. In certain other embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an Nω-cells of the dspe and the guide ligand is a transferrin (Tf), and a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a Tf-Nω-DOPE. In certain of these embodiments, m may be equal to 3, and the derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an NG-PE in accordance with formula 1.

In some embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is predstavljaet a NG-DOPE and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a TF-NG-DOPE. In other embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an NG-cells of the dspe and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a TF-NG-cells of the dspe. In some embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an NG-DOPE and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a TF-NG-cells of the dspe. In certain other embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is an NG-cells of the dspe and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is a TF-NG-DOPE.

Modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine can be obtained from SuccNωPE through reaction with guides ligand and other reagents as described here in more detail. TF-NωPE can either be obtained before mixing with other lipid components described herein lipid containing compositions (and optionally purified) or it can be obtained in situ by the interaction with prior SuccNωPE, which is included in the lipid containing composition.

Additional lipid components is you

Lipid containing compositions as described herein can also contain one or more additional lipid components in addition to those described here NωPE, SuccNωPE and/or TF-NωPE. Can be applied to a variety of additional lipid components, however, the term "additional(e) lipid(s) component(s)" is not intended to include unmodified phosphatidylethanolamine (PE) or derivatives of D in formula 1, 2 or 3. In some embodiments one or more additional(s) of the lipid(s) component(s) may be a phospholipid or one or more phospholipids. In certain embodiments one or more additional lipid components may include at least two neutral lipid. In other embodiments may be one or more phospholipids and, optionally, additional lipid" (which is not a phospholipid). Can be applied to different neutral lipids, however, at least two neutral lipid are not intended for inclusion nemodifitsirovannykh of phosphatidylethanolamine (PE) or modified PE of formulas 1, 2 or 3. Does not mean that one used here, the term "phospholipid" should include PE or their derivatives of the formulae 1, 2 or 3. Similar, the term "additional lipid" does not include any PE or their derivatives of the formulae 1, 2, or 3, or other phospholipids.

In specific embodiments, two or more neutral lipids can be applied in this lipid containing compositions and formulations. For example, at least two, at least three, at least four of neutral lipids; or two, three or four neutral lipid. In specific embodiments, there are two neutral lipid. In certain embodiments the lipid components of the compositions is limited to two neutral lipids, NωPE and SuccNωPE (or modified TF NωPE, when the reaction was carried out with a guiding factor).

In some embodiments, when the additional lipid component(s) includes a phospholipid, a phospholipid can be a phosphatidylcholine, including existing in nature, semi-synthetic or synthetic phosphatidylcholine (e.g., DSPC, DMPC, and so on). In some embodiments, the phosphatidylcholine is a failure not of natural phosphatidylcholine (e.g., phosphatidylcholine eggs). In particular what's the embodiments, the phosphatidylcholine is the failure acylphosphatidylethanolamine (for example, DMPC, DPPC, POPC, DSPC and so on). In some embodiments, the phospholipid is cationic. In other embodiments, the phospholipid is anionic. In some embodiments, the phospholipid is neutral. In specific embodiments, one or more phospholipid(s) are not anionic. In other embodiments, one or more phospholipid(s) are not cationic. In certain embodiments, where there is more than one phospholipid, can be included anionic or neutral lipid. Examples of phospholipids include, but are not limited to, phosphatidylcholine (PC), phosphatidic acid, phosphatidylserine, phosphatidylglycerol, etc. In some embodiments lipid containing compositions do not include phosphatidylserine or phosphatidylglycerol.

In certain embodiments, at least one of the at least two neutral lipids may be a phospholipid. In some embodiments, the phospholipid can be a phosphatidylcholine, including existing in nature, semi-synthetic or synthetic phosphatidylcholine (e.g., DSPC, DMPC, and so on). In some embodiments, the phosphatidylcholine is a failure not of natural phosphatidylcholine (e.g., phosphatidylcholine eggs). In specific embodiments, the phosphatidylcholine is acylphosphatidylethanolamine (e.g., DMPC, DPPC, POPC, DSPC and so on).

In some embodiments at least one of at least two neutral lipids can be a cholesterol or a derivative of cholesterol (e.g., related to the cholesterol connection, positively charged cholesterol (for example, DC-Chol), including radioisotope residue (for example,3H,14C,125I131I and so on)that have a functional residue (for example, fluorescent residue, etc.)).

In certain embodiments, when lipid containing compositions include one or more phospholipids, the composition may optionally include additional neutral nepovolite as an additional lipid. For example, cholesterol or a derivative of cholesterol, as described above.

Phospholipids (PC) for the application described here in lipid containing compositions include synthetic, semi-synthetic or existing in nature phospholipids. Examples of phospholipids include, but are not limited to, phosphatidylcholine (PC), phosphatidic acid, phosphatidylserine, phosphatidylglycerol, etc. In other embodiments, one or more phospholipids include phosphatidylcholine (PC) or phosphatidic acid, and do not include phosphatidylserine or phosphatidylglycerol.

In some embodiments, the phospholipid is a phosphatidylcholine. In certain embodiments, the phosphatidylcholine may represent, for example, diste railwastaition (DSPC), dimyristoylphosphatidylcholine (DMPC), dipalmitoylphosphatidylcholine (DPPC), palmitoyloleoylphosphatidylcholine (POPC), egg phosphatidylcholine (EPC), hydrogenated soy phosphatidylcholine (HSPC), etc. In specific embodiments, at least one phospholipid is a phosphatidylcholine. In certain embodiments, the phosphatidylcholine is a DMPC. In other embodiments, the phosphatidylcholine is a DSPC. In other embodiments, the phosphatidylcholine is a DPPC. In other embodiments, the phosphatidylcholine is a POPC. In other embodiments, the phosphatidylcholine is an EPC. In other embodiments, the phosphatidylcholine is an HSPC. In some embodiments includes one phospholipid and it is a DMPC, DSPC, DPPC, POPC, EPC or HSPC. In specific embodiments where the lipid containing composition includes a single phospholipid (PE phospholipid), the phospholipid is a DMPC. In other embodiments where the lipid containing composition includes a single phospholipid (PE phospholipid), the phospholipid is a DSPC. In other embodiments where the lipid containing composition includes a single phospholipid (PE phospholipid), the phospholipid is a DPPC. In other embodiments where the lipid containing composition includes a single phospholipid (PE phospholipid), the phospholipid is the Wallpaper POPC. In other embodiments where the lipid containing composition includes a single phospholipid (PE phospholipid), the phospholipid is an EPC. In other embodiments where the lipid containing composition includes a single phospholipid (PE phospholipid), the phospholipid is a HSPC.

In certain embodiments additional(e) lipid(s) component(s) may include at least one phospholipid, such as cholesterol or a derivative of cholesterol. In specific embodiments additional(e) lipid(s) component(s) are the only phospholipid and cholesterol. In certain embodiments additional(e) lipid(s) component(s) include at least one phosphatidylcholine and cholesterol. In specific embodiments additional(e) lipid(s) component(s) include only the phosphatidylcholine and cholesterol. In certain embodiments, where included cholesterol, phospholipid is a DSPC, DMPC, DPPC, POPC, EPC or HSPC. In some embodiments additional(e) lipid(s) component(s) include cholesterol and DMPC. In other embodiments additional(e) lipid(s) component(s) include cholesterol and DSPC. In certain embodiments additional(e) lipid(s) component(s) are cholesterol and one of DMPC or DSPC. In certain embodiments additional(e) lipid(e) component is(s) represent cholesterol and DMPC. In other embodiments additional(e) lipid(s) component(s) are cholesterol and DSPC. In other embodiments additional(e) lipid(s) component(s) include cholesterol and DPPC. In other embodiments additional(e) lipid(s) component(s) include cholesterol and POPC. In other embodiments additional(e) lipid(s) component(s) include cholesterol and EPC. In other embodiments additional(e) lipid(s) component(s) include cholesterol and HSPC. In certain embodiments additional(e) lipid(s) component(s) are cholesterol and one of DMPC, DSPC, DPPC, POPC, EPC or HSPC. In certain embodiments additional(e) lipid(s) component(s) are cholesterol and DMPC. In other embodiments additional(e) lipid(s) component(s) are cholesterol and DSPC. In other embodiments additional(e) lipid(s) component(s) are cholesterol and DPPC. In other embodiments additional(e) lipid(s) component(s) are cholesterol and POPC. In other embodiments additional(e) lipid(s) component(s) are cholesterol and EPC. In other embodiments additional(e) lipid(s) component(s) are cholesterol and HSPC.

In specific embodiments of the present one phospholipid and phospholipid is not a HSPC or EPC.

In particular the embodiments have one or more phospholipids. In certain embodiments, one or more phospholipids include phosphatidylcholine. In certain embodiments include one or more phospholipids and cholesterol (or derivative of cholesterol). In certain embodiments, the phospholipid is a phosphatidylcholine and composition additionally contains cholesterol (or derivative of cholesterol). In specific embodiments, the phosphatidylcholine is a phosphatidylcholine, which includes the residue of a saturated fatty acid (e.g., DMPC, DSPC or DPPC). In certain embodiments, the phosphatidylcholine is not a phosphatidylcholine eggs. In specific embodiments, the phosphatidylcholine is not a HSPC.

In specific embodiments, there are two neutral lipid. In some embodiments, two neutral lipids are cholesterol (or derivative) cholesterol and phosphatidylcholine. In certain embodiments, the phosphatidylcholine is a phosphatidylcholine, which includes the residue of a saturated fatty acid (e.g., DMPC, DSPC or DPPC). In certain embodiments, the phosphatidylcholine is not a phosphatidylcholine eggs.

Additional(e) lipid(s) component(s), as described here and known to experts in the art, commercially available from several suppliers, including, for example, Avanti Polar Lipids, Inc. (Alabaster, AK), Northern Lpid Inc. (Canada), Lipoid GmbH (Germany), NOF Corporation (Japan), Nippon Fine Chemical Co., Ltd (Japan).

Pharmaceutical substances

Various medicinal substances can be incorporated into lipid containing compositions of the present invention, for example compound or gene. In certain embodiments of the medicinal substance can be an anticancer agent, for example an anti-cancer agent, suitable for encapsulation in a liposome. The amount of drug substance for inclusion in the lipid containing compositions and their compounds, as described here, can be easily determined by a person skilled in the field of technology from the point of view offered here are instructions and depending on selected medicinal substance and intended use of the composition or composition, taking into account factors specific for both drug substance and for the individual being treated, as described hereafter.

In certain embodiments of the medicinal substance can be a nucleic acid, for example, nucleic acid encoding a sequence with anti-cancer properties. For example, but not limited to, antisense oligonucleotides, ribozymes, etc.

In some embodiments the anti-cancer agent may be a cytotoxic drug, including those which of quiet well-known specialists in the field of technology and practitioners. Examples of anticancer agents include inhibitors of topoisomerase I, Vinca alkaloids, alkylating agents (including platinum compounds), taxanes and other well-known specialists in this field of technology.

In some embodiments the anti-cancer drug can be an inhibitor of topoisomerase I, for example, but not limited to, topotecan, irinotecan, etc.

Anticancer drugs may also be a Vinca alkaloid such as vincristine, vinblastine, Finlepsin, windowsin, vinorelbine, vindesine, etc.

Further, the anti-cancer drug substance may also be a platinum compound. Non-limiting examples of platinum compounds include biltin, cisplatin, carboplatin, ormaplatin, oxaliplatin, triplatin, angloplats, lobaplatin, spiroplatin etc.

Oxaliplatin (platinum (II) CIS-oxalato complex TRANS-1-1,2-diaminocyclohexane) is platinum more specifically, organoplatinum, the complex having a structure represented by the following formula, shown below. Oxaliplatin is also known as the following: diaminocyclohexane platinum, DACH-platinum and CIS-[(1R, 2R)-1,2-cyclohexanediamine-N,N'][oxalato(2)-O,O']platinum (C8H14N2O4Pt; Mol. M 397,4 g/mol). As mentioned previously, oxaliplatin is the active pharmaceutical ingredient in Eloxatin™.

Oxaliplatin is suitable as an anti-cancer agent, as it has therapeutic activity, similar to that of cisplatin, and the relatively low nephrotoxicity and emetic effect (stimulation of the vomiting). The methods of production oxaliplatin well known in the art (for example, JP-A-9 - 40685; U.S. patent Nos. 4169846, 5338874; 5959133; 5298642; and 5290961 (the content of which is incorporated herein in its entirety). Oxaliplatin is additionally described in Chaney SG et al. "Recognition and processing of cisplatin - and oxaliplatin-DNA adducts." Crit Rev Oncol Hematol. (2005) 53: 3-11 (included here by reference in its entirety).

In certain embodiments, the concentration of oxaliplatin encapsulated in the liposome is approximately 1 mg/ml, for example, approximately 0.8 mg/ml

Usually liposomal composition of the present invention contains from about 1 to about 50 μg oxaliplatin/mg lipid and from about 1 to about 150 μg TF/mg lipid. For example, from about 10 to about 50 μg oxaliplatin/mg lipid and from about 10 to about 150 μg TF/mg lipid.

In certain embodiments the compositions contain from about 1 to about 45 μg oxaliplatin/mg lipid, from about 1 to about 40 μg oxaliplatin/mg lipid, from about 1 d is approximately 35 µg oxaliplatin/mg lipid, from about 1 to about 30 μg oxaliplatin/mg lipid, from about 1 to about 25 μg oxaliplatin/mg lipid, from about 1 to about 20 μg oxaliplatin/mg lipid, from about 1 to about 15 μg oxaliplatin/mg lipid, from about 1 to about 10 μg oxaliplatin/mg lipid, from about 1 to about 5 μg oxaliplatin/mg lipid, from about 5 to about 50 μg oxaliplatin/mg lipid, from about 5 to about 45 μg oxaliplatin/mg lipid, from about 5 to approximately 35 µg oxaliplatin/mg lipid, from about 5 to about 25 μg oxaliplatin/mg lipid, from about 5 to about 20 mcg oxaliplatin/mg lipid, from about 5 to about 15 μg oxaliplatin/mg lipid, from about 5 to about 10 μg oxaliplatin/mg lipid, about 1 μg oxaliplatin/mg lipid, about 2 μg oxaliplatin/mg lipid, about 4 μg oxaliplatin/mg lipid, about 5 μg oxaliplatin/mg lipid, about 10 μg oxaliplatin/mg lipid, about 15 μg oxaliplatin/mg lipid, about 20 μg oxaliplatin/mg lipid, about 30 μg oxaliplatin/mg lipid, about 40 μg oxaliplatin/mg lipid and the and about 50 μg oxaliplatin/mg lipid.

In certain embodiments the compositions contain from about 1 to about 145 μg TF/mg lipid, from about 1 to about 120 μg TF/mg lipid, from about 1 to about 115 μg TF/mg lipid, from about 1 to about 100 μg TF/mg lipid, from about 1 to about 90 μg TF/mg lipid, from about 1 to about 70 μg TF/mg lipid, from about 1 to about 60 μg TF/mg lipid, from about 1 to about 50 μg TF/mg lipid from approximately 1 to approximately 25 μg TF/mg lipid, from about 10 to about 150 μg TF/mg lipid, from about 10 to about 140 μg TF/mg lipid, from about 10 to about 125 μg TF/mg lipid, from about 10 to about 100 μg TF/mg lipid, from about 10 to about 80 μg TF/mg lipid, from about 10 to about 50 μg TF/mg lipid, from about 10 to about 25 μg TF/mg lipid, about 1 μg TF/mg lipid, about 5 μg TF/mg lipid, about 10 μg TF/mg lipid, about 25 μg TF/mg lipid, about 40 μg TF/mg lipid, about 50 μg TF/mg lipid, about 70 μg TF/mg lipid, about 100 μg TF/mg lipid, about 120 μg TF/mg lipid, about 140 μg TF/mg lipid, or about 150 μg TF lipid.

In some embodiments from about 0.5 to about 50 μg oxaliplatin/mg lipid and from about 1 to about 150 μg TF/mg lipid. In some embodiments from about 5 to about 50 μg oxaliplatin/mg lipid and from about 10 to about 100 μg/mg, In certain embodiments from about 2 to about 50 μg oxaliplatin/mg lipid and from about 5 to about 150 μg TF/mg lipid; from about 3 to about 50 μg oxaliplatin/mg lipid and from about 5 to about 150 μg TF/mg lipid; from about 4 to about 50 μg oxaliplatin/mg lipid and from about 5 to about 150 μg TF/mg lipid; from approximately 2 to approximately 40 μg oxaliplatin/mg lipid and from about 5 to about 150 μg TF/mg lipid; from about 3 to about 40 μg oxaliplatin/mg lipid and from about 5 to about 150 μg TF/mg lipid; from about 4 to about 40 μg oxaliplatin/mg lipid and from about 5 to about 150 μg TF/mg lipid; from about 2 to about 50 μg oxaliplatin/mg lipid and from about 10 to about 150 μg TF/mg lipid; from about 3 to about 50 mcg oxaliplatin/mg lipid and from about 10 to about the part 150 μg TF/mg lipid; from approximately 4 to approximately 50 µg oxaliplatin/mg lipid and from about 10 to about 150 μg TF/mg lipid; from about 5 to about 50 μg oxaliplatin/mg lipid and from about 5 to about 100 μg TF/mg lipid; from about 5 to about 50 μg oxaliplatin/mg lipid and from about 5 to about 100 μg TF/mg lipid; or from about 0.5 to about 50 μg oxaliplatin/mg lipid and from about 5 to about 100 μg TF/mg lipid.

In certain embodiments, the concentration of oxaliplatin in liposomal composition equal to 0.8 ± 10% mg/ml

In certain embodiments, when the drug is oxaliplatin, oxaliplatin can be dissolved in solution (e.g. aqueous solution). In some embodiments, the solution comprises a sugar (e.g., trehalose, maltose, sucrose, lactose, mannose, mannitol, glycerol, dextrose, fructose, etc). The sugar concentration may amount to several percent. For example, the concentration of sugar (about./about.) approximately 0,1-12%; 0,5-12%, 1%-12%, 2%-8%, 2%-6%, 2%-5%, 2%-4%, 2%-5%, 2%-6%, 2%-8%, 2%-9%, 2%-10%, 4%-10%, 4%-9%, 4%-8%, 4%-6%, 3%-4%, approximately 2%, approximately 3%, approximately 4%, approximately 5%, about 6%, about 7%, about 8%, about 9% or about 10%. In certain embodiments, R is the target includes sugar and water is. Assume that the solution in which is dissolved oxaliplatin may also contain additional components, including well-known experts in this field of technology.

In certain embodiments, the sugar concentration is about 5%, about 7%, about 8%, about 9% or about 10%. In other embodiments, the sugar concentration is from about 5% to about 10%. In some embodiments sugar is a dextrose concentration of dextrose in the solution oxaliplatin is approximately 5%. In some embodiments sugar represents dextrose, and the concentration of dextrose in the solution oxaliplatin is approximately 9%. In certain embodiments sugar is a sucrose and the concentration of sucrose in solution oxaliplatin is approximately 9%. In certain embodiments sugar is a sucrose and the concentration of sucrose in solution oxaliplatin is approximately 10%.

In some embodiments, the concentration of sugar in solution may be, for example, from about 50 mg/ml to about 150 mg/ml, from about 50 mg/ml to about 130 mg/ml, from about 50 mg/ml to about 120 mg/ml, from about 50 mg/ml to about 100 mg/ml, from priblizitelen is 80 mg/ml to about 100 mg/ml, from about 90 mg/ml to about 150 mg/ml, from about 90 mg/ml to about 130 mg/ml, about 60 mg/ml, about 80 mg/ml, about 90 mg/ml, about 100 mg/ml, about 110 mg/ml, about 105 mg/ml, about 120 mg/ml, or about 140 mg/ml

The solution may also contain other ingredients known to experts in the art, such as, but not limited to, salts, buffers, polyalcohols, xylytol etc. In certain embodiments, the solution in which is dissolved oxaliplatin, contains sodium phosphate (for example, monobasic and/or dibasic sodium phosphate).

In certain embodiments, the concentration of sodium phosphate may be from approximately 5 to approximately 15 mm. For example, from about 5 to about 12 mm, from about 5 to about 10 mm, from about 5 to about 7 mm, from about 7 to about 12 mm, from about 7 to about 15 mm, from about 9 to about 12 mm, about 5 mm, about 7 mm, about 10 mm, about 12 mm or about 15 mm.

In certain embodiments, the sugar solution may optionally include from about 1.0 to about 1.5 mg/ml of sodium phosphate. For example, from approximately 1.2 to approximately 1.5 mg/ml, from about 1.0 to about 1.7 mg/ml, from approximately 1.0 to approximately 2 mg/ml, from about 1.0 to about 2.5 mg/ml, from about 1.0 to about 3 mg/ml, from about 0.5 to about 3.5 mg/ml of sodium phosphate.

In some embodiments, the pH of the solution should be from about 6.5 to about 7.5, from about 6.7 to about 7.5, from about 7 to about 7.5, about 7, about 7.5, about 6,8 or approximately 6.5.

In some embodiments of the medicinal substance is an oxaliplatin and contained in a solution of approximately 9% sucrose. In some embodiments of the medicinal substance is an oxaliplatin and contained in a solution of approximately 9% sucrose concentration oxaliplatin approximately 1 mg/ml In some embodiments of the medicinal substance is an oxaliplatin and contained in the sucrose solution, about 105 mg/ml In some embodiments of the medicinal substance is an oxaliplatin and contained in the sucrose solution, about 105 mg/ml concentration oxaliplatin in the solution of liposomes of approximately 1 mg/ml In certain embodiments, the solution further comprises sodium phosphate. In certain embodiments oxaliplatin is in a concentration of approximately 0.8 ± 10% mg/ml dissolve the and liposomes.

Labeled compounds

A variety of labeled compounds can also be included in the lipid containing compositions of the present invention. Usually labeled compound can be an agent suitable for the implementation of diagnostic procedures in vivo.

As in the case of the inclusion and application of medicinal substances, as described here, the number of labeled compounds intended for inclusion in the lipid containing compositions and their compounds, as described here, can be easily determined by a person skilled in the field of technology from the point of view presented here of instructions and depending on the selected labeled compounds and intended use of the composition or composition, taking into account factors specific for the labeled compounds, and for the individual subjected to the diagnosis, as described here below.

Examples of labelled compounds include, for example, materials comprising a radioisotope (for example,3H,14C,67Ga111In125I131I133Xe and so on), the material including a fluorescent part (for example, fluorescein, fluoresceinisothiocyanate etc), the material comprising an enzyme (such as peroxidase, alkaline phosphatase etc), as well as additional labeled compounds known to specialists in this field of technology.

As should be clear JV is the expert in this field of technology the choice of labeled compounds and methods of diagnosis must depend on explored organ (e.g. liver, pancreas, prostate etc), cloth (e.g., tumor or non-tumor or tissue type (e.g., breast cancer, etc.)). For example, lipid containing compositions (for example, liposomes directional containing liposomes song, etc), including125I, especially suitable for identification of the presence and determine the severity (e.g., initial, during treatment, after treatment for various types of cancer (eg, breast cancer, gastric cancer, colorectal cancer, colon cancer, etc.) using a gamma counter.

Guiding factors

Unless otherwise specified, the terms "smart factor" and "smart ligand" may be used interchangeably.

Described herein lipid containing compositions are characterized by the inclusion of derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, modified guiding factor (TF-NωPE), aimed at specific target cells. The term "smart factor" refers to a part which can bind to a receptor or surface antigen present on the surface of target cells. In certain embodiments of the guiding factors directed n is the cell surface receptors on specific cell target. The smart factor is often a protein or peptide that can be attached to the lipid component of the lipid containing compositions.

Most effectively guides the factors are chosen so that the receptor or antigen targeted in the factor was present only in those cells that are the target for delivery of drug or labeled compound (e.g., abnormal cells) and was not present in healthy cells. Alternatively, a higher number of receptors or antigens expressed on the target cells (e.g., pathological or diseased cells) compared with cells namereniami (e.g., healthy). Preferably, the receptor or antigen that binds guide factor, or not present, or were present in low numbers in healthy cells, so that the binding guiding factor was not arise often. In other words guiding factors necessary for selective delivery of liposomes, as described here (including encapsulated drug)to target cells (e.g., abnormal, unhealthy, and so on). Selective delivery of the encapsulated drug to the target cells, thus, reduces the possibility of adverse effects due to actions encapsulite the data of drug or labeled compound on cells namesti (for example, healthy), thereby also reducing the side effects known to the individual is administered a composition or its composition.

Examples of guiding factors include, but are not limited to, transferrin, folic acid, folate, hyaluronic acid chains of sugars (e.g., galactose, mannose, etc.), fragments of monoclonal antibodies, asialoglycoprotein etc. and other guiding factors known to specialists in this field of technology.

In specific embodiments the directional factor is a protein or peptide that is directed to the cell surface receptor (e.g., transferrin, folate, folic acid, asialoglycoprotein etc).

In other embodiments the guide factor is directed to an antigen (e.g., fragments of monoclonal antibodies (e.g., Fab, Fab', F(ab')2, Fc, etc)). Do not assume that guides the factors include intact or whole monoclonal antibody. Used here, the term "whole antibody" or "intact antibody" and related compounds generally refers to the IgG antibody or immune globulin. The fragment of the monoclonal antibodies generally refers to the product of the splitting of monoclonal antibodies, for example the fragment obtained using rosalene a protease such as pepsin, etc.

In certain embodiments the directional factor, not voltage, is Allen the antigen (for example, is not a fragment of monoclonal antibodies, for example Fab, Fab', F(ab')2, Fc, etc)).

In certain embodiments the guide factor represents transferrin.

Transferrin (Tf) is an iron binding protein with a molecular weight of 80,000, which is synthesized in hepatocytes and is in the blood. Transferrin provides cells with iron (Fe) via Tf receptors on the surface of each cell. The transferrin receptor is normally expressed in tumor tissues in higher amounts as compared with normal tissues, regardless of the types of tumors. Membranes of tumor cells, as is known, hyperexpression transferrin receptors to maintain cell proliferation. See, Shindelman JE, Ortmeyer AE, Sussman HH. "Demonstration of the transferrin receptor in human breast cancer tissue. Potential marker for identifying dividing cells." Int J Cancer. (1981) 27(3):329-34; Lloyd JM, O Dowd T, Driver M, DE Tee. "Demonstration of an an epitope of the transferrin receptor in human cervical epithelium - a potentially useful cell marker." J Clin Pathol. (1984) 37(2):131-5; and Habeshaw JA, Lister TA, Stansfeld AG, Greaves MF. "Correlation of transferrin receptor expression with histological class and outcome in non-Hodgkin lymphoma." Lancet. (1983) 1(8323): 498-501. The binding of therapeutic agents with transferrin should, therefore, increase the seizure of the drug in tumor cells via the transferrin receptor. Without being bound by mechanism of action, the likely path of the capture of liposomes with transferrin as described here, PR is dstable schematically in Fig. 2 and 3. Transferrin commercially available or can be obtained by recombinant means, as described, for example, in U.S. patent 5026651, incorporated herein fully by reference.

Without being bound by theory, it is believed that the conjugation of transferrin (Tf) with NωPE is carried out using the reaction of primary amine with NωPE that will introduce to the formation of amide linkages carboxylic acid anchored between the lipid and protein.

In certain embodiments the molar ratio of Tf to the total lipid present in the liposomal product directional approximately 0,00014:1 mol/mol (Tf:total lipid) (0,015 mass./mass.). In other embodiments the molar ratio of Tf:total lipid present in the liposomal product directional ranges from approximately 0,016 to approximately 0,029:from about 126 to about 158 mm/mm.

Lipid containing compositions

Described herein lipid containing compositions include liposomes directed action, including derivatives, lipids, additional lipids and encapsulated drug or labeled compound, and intermediate compounds used to produce liposomes directed action, including liposomal mixture containing liposome composition as described here, where lipid containing compositions (in the including liposomes directional) free from unmodified phosphatidylethanolamine and hydrophilic polymers, such as, but not limited to, polyethylene glycol. Lipid containing compositions also include liposomes, which include Tf, but does not include drug or labeled compound (e.g., unloaded liposomes).

Used here, the term "hydrophilic polymer" and related compounds refers to polymers such as polyethylene glycol (PEG) and other polyethoxysiloxane polymers, which are used in the field of liposomes to protect liposomes in an attempt to increase the half-life of liposomes in a circulatory manner. Understood that this term covers available hydrophilic polymers, ecovalence associated with liposomes, as well as hydrophilic polymers, which somehow conjugated or covalently associated with a particular component of liposomes (e.g., modified PEG-lipids etc). Such hydrophilic polymers alternative are identified in this area as "water-soluble" polymers. Additional examples of hydrophilic polymers include, but are not limited to, polyvinyl alcohol, polylactic acid, polyglycolic acid, polyvinylpyrrolidone, polyacrylamide, polyglycerins, polyoxazolines etc.

Used here, the term "lipid mixture and kindred relates to mixtures of lipid components, as described here, where the lipid mixture newslocal solution for example, an aqueous solution (e.g., water, buffer or a mixture of water and miscible with water solvent (e.g., sugar (e.g., trehalose, sucrose, lactose, mannose, dextrose, fructose etc), sugar alcohols (e.g. sorbitol, maldita, lactate, glycerol, mannitol, etc), alcohol (e.g. ethanol, tert-butanol etc) etc) or organic solvent.

The term "containing liposome composition and related refers to mixtures of lipid and, optionally, drug(s) substance(s) or labeled(s) connection(s)in which an aqueous solution (e.g., water, buffer (e.g., acetate buffer, phosphate buffer, citrate buffer, borate buffer, tartrate buffer, etc. or a mixture of water and miscible with water solvent) included with mixing (e.g., one or more mixing, shaking and so on). The aqueous solution may also include additional components such as one or more sugars (e.g. trehalose, maltose, sucrose, lactose, mannose, dextrose, fructose, etc), sugar alcohols (e.g. sorbitol, ▫ maltitol, lactic, lures, glycerin etc), alcohol (e.g. ethanol, tert-butanol etc), etc. and an aqueous solution may also include an organic solvent (for example, esters (e.g. ethyl acetate, butyl acetate, etc.), aliphatic hydrocarbons (e.g. hexane, heptane, etc.) aromatic hydrocarbons (for example, toluene, xylene and so on), halogenated hydrocarbons (e.g. chloroform, dichloromethane, dichloroethane, etc.), ethers (for example, THF, dioxane, diethyl ether, isopropyl ether, etc.), cyclic hydrocarbons (e.g. cyclohexane, etc.), DMF, DMSO, etc. or mixtures thereof. Containing liposome composition should usually include not homogeneous mixture of lipid, aqueous solution and liposomes, which have a wide distribution around 100-10000 nm and the average diameter of 500-2000 nm. Characteristic examples containing liposome compositions are additionally presented in the examples.

In certain embodiments lipid containing compositions do not include hydrophilic polymers. In specific embodiments lipid containing compositions do not include PEG.

In some embodiments the intermediate lipid mixtures include at least two different neutral lipid or one or more phospholipids and its derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine, where lipid components described here in more detail and where the mixture is free from non-modified phosphatidylethanolamine and hydrophilic polymers, such as polyethylene glycol. Optional aqueous solution, as described here, can be mixed with the lipid components with the formation containing liposomes of the composition. In certain embodiments the lipid mixture including the AET drug or labeled compound. In specific embodiments the lipid mixture can be treated with the formation containing liposomes of the liposomal composition or structure.

In some embodiments the intermediate lipid mixtures include at least two different neutral lipid or one or more phospholipids, derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and drug or labeled compound, where lipid components and the medicinal substance/labeled compound described herein in more detail and where the mixture is free from non-modified phosphatidylethanolamine and hydrophilic polymers, such as polyethylene glycol. Optionally, aqueous solution, as described here, can be mixed with the lipid components with the formation containing liposomes of the composition, for example, when you add a drug or labeled compound in the form of an aqueous solution of a medicinal substance/labeled compounds. In certain embodiments containing liposome composition may be processed (e.g., using one or more methods of extrusion, excluding the size of chromatography, etc. or methods known in the art) with the formation of liposomes.

In some embodiments the lipid mixtures include one or more phospholipids or at least two different neutral lipid production is the initial N-(ω)-dicarboxylic acid and phosphatidylethanolamine and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine, where lipid components described here in more detail and where the mixture is free from non-modified phosphatidylethanolamine and hydrophilic polymers, such as polyethylene glycol. The mixture may also be essentially free from non-NHS source materials, by-product and/or product splitting associated with the synthesis operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine (for example, carbodiimides (e.g., DCC, EDC etc), acylated compounds of urea and so on). Optionally, aqueous solution, as described here, can be mixed with the lipid components with the formation containing liposomes of the composition.

In some embodiments the intermediate lipid mixtures include one or more phospholipids or at least two different neutral lipid-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, where lipid components described here in more detail and where the mixture is free from non-modified phosphatidylethanolamine and hydrophilic polymers, such as polyethylene glycol. The mixture may also be essentially free from non-NHS source materials, by-product and/or product splitting associated with the synthesis operations ether of proizvodi the th N-(ω)-dicarboxylic acid and phosphatidylethanolamine (for example, carbodiimides (e.g., DCC, EDC etc), acylated compounds of urea and so on). Optionally, aqueous solution, as described here, can be mixed with the lipid components with the formation containing liposomes of the composition. In some embodiments the lipid mixture does not include drug or labeled compound. The lipid mixture can be treated with the formation of lipid containing compositions or liposomal composition.

In some embodiments the intermediate lipid mixtures include one or more phospholipids or at least two different neutral lipid-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and drug or labeled compound, where lipid components and the medicinal substance/labeled compound described herein in more detail and where the composition is free of non-modified phosphatidylethanolamine and hydrophilic polymers, such as polyethylene glycol. The mixture may also be essentially free from non-NHS source materials, by-product and/or product splitting associated with the synthesis operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine (for example, carbodiimides (e.g., DCC, EDC etc), acylated compounds m is chevigny etc). Optionally, aqueous solution, as described here, can be mixed with the lipid components with the formation containing liposomes of the composition.

In certain embodiments the intermediate lipid containing compositions include a liposome containing one or more phospholipids or at least two different neutral lipid-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and encapsulated drug substance, where the lipid and drug components described here in more detail and where the liposome is free from non-modified phosphatidylethanolamine and hydrophilic polymers, such as polyethylene glycol. Liposomes can also be essentially free from non-NHS source materials, by-product and/or product splitting associated with the synthesis operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine (for example, carbodiimides (e.g., DCC, EDC etc), acylated compounds of urea and so on). In some embodiments the lipid mixture does not include drug or labeled compound. The lipid mixture can be treated with the formation of lipid containing compositions or liposomal composition.

In certain embodiments the lipid mixture include the Dean or more phospholipids or at least two different neutral lipid-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and encapsulated drug or labeled compound, where lipid components, the smart factor and drug or labeled compound is described here in more detail and where the liposome is free from non-modified phosphatidylethanolamine and hydrophilic polymers, such as polyethylene glycol. In some embodiments the lipid mixtures, liposomes essentially free from non-NHS source materials, by-product and/or fission products associated with the synthesis operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine (for example, carbodiimides (e.g., DCC, EDC etc), acylated compounds of urea and so on). In specific embodiments the lipid is a mixture containing liposome composition (for example, where a drug or labeled compound is added in aqueous solution). Optionally, aqueous solution, as described here, can be mixed with the lipid components with the formation containing liposomes of the composition.

In certain embodiments, the liposomes directional include liposomes containing the ie one or more phospholipids or at least two different neutral lipid-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and encapsulated drug or labeled compound, where lipid components, the smart factor and the medicinal substance/labeled compound described herein in more detail and where the liposome is free from non-modified phosphatidylethanolamine and hydrophilic polymers, such as polyethylene glycol. In some embodiments of liposomes directional, liposomes essentially free from non-NHS source materials, by-product or products of the splitting associated with the synthesis operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine (for example, carbodiimides (e.g., DCC, EDC etc), acylated compounds of urea and so on). However, in some embodiments operations ether derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine may be present in the initial stages of obtaining liposomes directed action (e.g., NHS-NG-PEs (for example, NHS-NG-DOPE, NHS-NG-cells of the dspe etc), for example, before hydrolysis operations ether, which can provide, for example, the NHS and NG-DOPE in the final composition. In certain embodiments, when SuccNωPE not included NR is try liposomes liposome or containing liposome composition may be free or essentially free from the NHS, as in the case of prior learning TF-NωPE and used as educt. In specific embodiments the liposomes directional essentially free from DCC and EDC. In certain embodiments, the liposomes directional essentially free from DCC.

Additionally, each containing liposome compositions, as described here, can be processed to produce liposomes. Obtaining liposomes are well known in the art, can also be carried out in accordance with the described herein ways, for example as described for ways to obtain A and B, described in more detail below. Methods of obtaining liposomes containing liposome compositions include, but are not limited to, extrusion, sonication, bubbles reverse phase, the procedure of freezing and thawing, excluding the size of chromatography, ultrafiltration, etc. and combinations thereof. Liposomes formed from the described herein containing liposome compositions can include drug or labeled compound or can be free from drug or labeled compound (for example, liposomes, also referred to here as "unloaded liposomes"). In particular OPL is the premises containing liposomes of the composition, liposomes (including non-loaded liposomes and liposomes directed action can be written in the form of pharmaceutical compositions, and optionally can be used in the here described methods of treatment and diagnosis and/or sets.

The term "essentially free" refers to the levels and products that are not determinate or determinable minimal traditional analytical methods used in the art. For example, HPLC (see, for example, the European Pharmacopoeia 5thEd.), TLC, gas chromatography, etc. as well as other analytical methods known to the person skilled in the technical field.

For example, lipid containing compositions may contain less than about 0.1%, less than about 0.5%, less than about 1%, less than about 2%, less than about 3%, less than about 4%, less than about 5%, or less than about 6% by weight of a specific source of substance by-product or product splitting associated with the synthesis operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and total lipid component. In specific embodiments of the composition should contain less than about 10%, less than about 7%, less than about 5%, less than about 3%, less than about 2% or less of CA is approximately 1% of the total impurities (for example, % amount of original substance, by-product and product splitting associated with the synthesis operations ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine).

In some embodiments the liposomes directional include liposomes containing phosphatidylcholine (such as neutral, anionic or cationic), cholesterol or a derivative of cholesterol-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and encapsulated drug or labeled compound, where lipid components, the smart factor and the medicinal substance is described here in more detail and where the liposome is free from non-modified phosphatidylethanolamine and hydrophilic polymers, such as polyethylene glycol. In specific embodiments, the phosphatidylcholine is a neutral phosphatidylcholine.

In specific embodiments the liposomes directional include liposomes containing phosphatidylcholine (such as neutral, anionic or cationic), cholesterol or a derivative of cholesterol-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine modified transferrin-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and incapsulate the data drug or labeled compound, where lipid components and drug or labeled compound is described here in more detail and where the liposome is free from non-modified phosphatidylethanolamine and hydrophilic polymers, such as polyethylene glycol. In specific embodiments, the phosphatidylcholine is a neutral phosphatidylcholine.

In specific embodiments the liposomes directional include liposomes containing phosphatidylcholine (such as neutral, anionic or cationic), cholesterol or a derivative of cholesterol-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine modified transferrin-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and encapsulated oxaliplatin, where lipid components described here in more detail and where the liposome is free from non-modified phosphatidylethanolamine and hydrophilic polymers, such as polyethylene glycol. In specific embodiments, the phosphatidylcholine is a neutral phosphatidylcholine.

In certain embodiments described herein lipid containing compositions (including liposomes targeted action and unloaded liposomes) and their compositions can optionally contain lipids obtained by modifying dicarboxylic acid, phosphatidylglycerol, sphingosine, ceramide-derived cholesterol is ina or the like. These derivatives of dicarboxylic acids may be obtained, as described here for derivatives N-(ω)-dicarboxylic acid and phosphatidylethanolamine and in accordance with the methods of getting known to the person skilled in the technical field.

In some embodiments described herein lipid containing compositions (including liposomes targeted action and unloaded liposomes) and their compositions do not include anionic lipids (e.g., phosphatidylserine, phosphatidylinositol, phosphatidylglycerol etc) or cationic lipids (e.g., sphingosine, DOTAP, DOTMA, DC-CHOL and so on). In specific embodiments, the composition is free from anionic lipids. In other embodiments the composition is free from cationic lipids. In certain embodiments the composition is free from cationic and anionic lipids.

In some embodiments lipid containing compositions composition contains a medicinal substance. In other embodiments lipid containing compositions include labeled connection.

In certain embodiments lipid containing compositions of the drug is oxaliplatin, the directional factor (TF) is a transferrin (Tf) and lipid components include: DMPC or DSPC and cholesterol or a derivative of cholesterol, and its derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified Tran is ferrino derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, where modified transferrin-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine includes transferrin attached to the derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine via amide linkages carboxylic acid.

In some embodiments derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine are NG-DOPE or NG-cells of the dspe.

In specific embodiments the lipid components of the lipid containing compositions represent a DMPC, cholesterol, NG-DOPE and modified TF NG-DOPE. In other embodiments the lipid components are DSPC, cholesterol, NG-DOPE and modified TF NG-DOPE. In some embodiments the lipid components are DMPC, cholesterol, NG-cells of the dspe and the modified TF NG-cells of the dspe. In certain other embodiments the lipid components are DSPC, cholesterol, NG-cells of the dspe and the modified TF NG-cells of the dspe. In certain of these embodiments the directional factor (TF) is a transferrin and drug is oxaliplatin.

In specific embodiments the lipid components are DPPC, cholesterol, NG-DOPE and modified TF NG-DOPE. In other embodiments the lipid components are POPC, cholesterol, NG-DOE and modified TF NG-DOPE. In some embodiments the lipid components are DPPC, cholesterol, NG-cells of the dspe and the modified TF NG-cells of the dspe. In certain other embodiments the lipid components are POPC, cholesterol, NG-cells of the dspe and the modified TF NG-cells of the dspe. In certain of these embodiments the directional factor (TF) is a transferrin and drug is oxaliplatin.

In specific embodiments the lipid components are HSPC, cholesterol, NG-DOPE and modified TF NG-DOPE. In other embodiments the lipid components represent the EPC, cholesterol, NG-DOPE and modified TF NG-DOPE. In some embodiments the lipid components are HSPC, cholesterol, NG-cells of the dspe and the modified TF NG-cells of the dspe. In certain other embodiments the lipid components represent the EPC, cholesterol, NG-cells of the dspe and the modified TF NG-cells of the dspe. In certain of these embodiments the directional factor (TF) is a transferrin and drug is oxaliplatin.

The ratio of lipid components

Typically, the molar percentage of the original substances NG-DOPE should be from about 2.5 mol% to about 4.5 mol% in comparison with the total content of lipids. Additionally, the molar percentage of the original substances NHS-NG-DOPE should be from about 0.5 mol% to over emphasis is about 2.5 mol% compared to total lipids. In some embodiments, the relative molar ratio of NG-DOPE to NHS-NG-DOPE should be about 3.4:1. In certain embodiments, the relative ratio of the mol% of NG-DOPE to NHS-NG-DOPE may be approximately 4:1. In specific embodiments, when there are neutral phospholipid and neutral lipid molar ratio (e.g., DMPC:Chol:NG-DOPE:NHS-NG-DOPE) may be 43,0:38,5:3,42:1, which can also be expressed as 50:45:4:1 in mol%.

In certain embodiments, the additional(different) lipid(s) in mol% to NωPE to SuccNωPE (for example, at least two neutral lipid:NωPE:SuccNωPE or one or more phospholipids:NωPE:SuccNωPE, or (one or more phospholipids + neutral(nye) lipid(s):NωPE:SuccNωPE) may be from about 98 mol% to about 87 mol% of additional lipids:from about 1 mol% to about 12 mol% NωPE:from about 0.5 mol% to about 1% SuccNωPE; where the total mole% of all components is 100 mol%. For example, additional lipids:NωPE:SuccNωPE can be approximately 95:4:1, 90:9:1, 92:7:1, 93:6:1 etc.

In specific embodiments, when additional lipids include phospholipids and other lipid, such as cholesterol, derivatives of cholesterol, etc., the range of mol% for each lipid component is from about 30 mol% to about 64%, where the amount of the additional lipid is from about 98 mol% to about 87 mol%.

In certain embodiments, when additional lipids represent two different neutral lipid, the range of mol% for each neutral lipid is from about 30 mol% to about 64%, where the amount of neutral lipid is from about 98 mol% to about 87 mol%.

In the example embodiment, where one additional lipid is a phosphatidylcholine, and the second additional lipid is a cholesterol or a derivative of cholesterol, mol% phosphatidylcholine is from about 30 to about 70 mol% (for example, from about 50 to about 64 mol%, from about 40 to about 65 mol%, from about 40 to about 60 mol%, from about 50 to about 62 mol%, from about 55 to about 60 mol%, from about 35 to about 55 mol%, about 30 mol%, about 40 mol%, about 45 mol%, about 50 mol%, about 55 mol%, about 60 mol%, about 65 mol%, about 70 mol%) and mol% cholesterol or a derivative of cholesterol is from about 30 to about 60 mol% (for example, from about 32 to about 45 mol%, from about 32 to about 40 mol%, from about 40 to about 60 mole is%, from about 35 to about 55 mol%, from about 35 to about 60 mol%, from about 45 to about 60 mol%, from about 35 to about 45 mol%, about 30 mol%, about 35 mol%, about 40 mol%, about 45 mol%, about 50 mol%, about 55 mol%, or about 60 mol%). In some embodiments, the phosphatidylcholine is approximately 50 mol%, about 52 mol%, about 55 mol%, about 58 mol%, about 60 mol%, about 62 mol%, and cholesterol or a derivative of cholesterol is from about 30 mol%, about 32 mol%, about 34 mol%, about 35 mol%, about 37 mol%, about 38 mol%, about 40 mol%, about 42 mol%, about 43 mol%, about 45 mol%.

In specific embodiments mol% NωPE is from about 1 to about 11 mol%, from about 1 to about 10 mol%, from about 1 to about 8 mol%, from about 1 to about 6 mol%, from about 1 to about 5 mol%, from about 1 to about 4 mol%, from about 1 to about 3 mol%, from about 1 to about 2 mol%, from about 2 to about 10 mol%, from approximately the part 2 to about 5 mol%, about 1 mol%, about 2 mol%, about 3 mol%, about 4 mol%, about 5 mol%, about 7 mol%, about 8 mol%, about 9 mol%, about 10 mol%, about 11 mol%, or about 12 mol%.

In certain embodiments the ratio of the first additional lipid:second additional lipid:NωPE:SuccNωPE is, for example, 50:45:4:1. In some embodiments the first additional lipid is a phosphatidylcholine (e.g., DMPC, DOPC, DPPC, DSPC, and so on) and the second additional lipid is a cholesterol. In some embodiments PE NωPE is a DOPE or cells of the dspe. In specific embodiments NωPE is a NG-DOPE or NG-cells of the dspe. In some embodiments, the lipids are DMPC:Chol:NG-DOPE:NHS-NG-DOPE and attitude mol% is 50:45:4:1. In other embodiments, the lipids are DSPC:Chol:NG-cells of the dspe:NHS-NG-cells of the dspe and the ratio of the mol% is 50:45:4:1. In other embodiments, the lipids are DSPC:Chol:NG-cells of the dspe:NHS-NG-cells of the dspe and the ratio of the mol% is 62:33:4:1.

In some embodiments, the total mol% NωPE and TF-NωPE (NωPE+TF-NωPE) is from about 2 to about 13 mol% of total lipid content. For example, from about 2 to about 12 mol%, from about 2 to about 10 mol%, from about 2 to about 8 mol%, about is about 2 to about 6 mol%, from about 2 to about 4 mol%, about 2 mol%, about 3 mol%, about 4 mol%, about 5 mol%, about 6 mol%, about 7 mol%, about 8 mol%, about 9 mol%, about 10 mol%, about 11 mol%, or about 12 mol%.

Usually the total mol% TF-NωPE is from about 0.002 to about 0.2 mol% relative to the amount of lipid content. For example, in some embodiments, the total mol% TF-NωPE is from about 0.002 to about 0.15 mol%, the total mole% TF-NωPE is from about 0.002 to about 0.1 mol%, from about 0.002 to about 0.05 mol%, from about 0.01 to about 0.03 mol%, from about 0.005-about 0.2 mol%, from about 0,007 up to about 0.2 mol%, from about 0,007 up to about 0.05 mol%, from about 0.01 to about 0,025 mol%, from about 0.015 g to about 0,025 mol%, from about 0.01 to about 0.2 mol%, from about 0.02 to about 0.2 mol%, from about 0.04 to about 0.2 mol%, from about 0.06 to about 0.2 mol%, from about 0.08 to about 0.2 mol%, about 0.002 mol%, about 0,008 mol%, about 0.01 mol%, approximately 0,02 mol is%, about 0.03 mol%, about 0,025 mol%, about 0.015 mol%, about 0.06 mol%, about 0.08 mol%, about 0.1 mol%, about 0.15 mol%, or about 0.2 mol%.

Characterization of liposomes containing liposomes compositions

In addition to describing the lipid containing compositions according to the ratio of the components (for example, the ratio of lipids, the ratio of drug substance/labeled compounds to lipids, etc.) described herein containing liposome composition can also be characterized (for example, by physico-chemical properties and so on) using standard analytical methods that should be clear to experienced specialist. Such approved analytical methods include, but are not limited to, the determination of the average diameter of the negotiated amount, the resulting charge (Zeta-potential), the number of captured (i.e. signed) drug substance particle size, stability under various conditions (e.g. during storage, in the form prepared for introduction in vitro), osmotic properties, the number of conjugated guiding factor, etc. Illustrative analytical methods for such characteristics are described below and in the "Examples"section, and to describe the compositions can also be applied to additional ways of the local experienced.

As should be clear to the skilled technician, the content of the drug in the liposomes can be determined using accepted methods of analysis using HPLC, using appropriate controls, as is customary in the art and are additionally described in the "Examples"section. With the use of suitable controls may also be carried out identification of the encapsulated drug substances using HPLC or, in the case of certain medicinal substances (e.g. drugs containing platinum), using analytical methods such as ICP-MS (inductively coupled plasma mass spectrometry), used by experts in the field of technology.

In some embodiments, the amount of medicinal substance (for example, oxaliplatin etc) or labeled compound enclosed in the liposome or containing liposome composition can be from about 0.1 mg/ml to about 15 mg/ml inside the liposomes. For example,,the concentration of the medicinal substance can be from about 0.5 mg/ml to about 15 mg/ml, from about 0.5 mg/ml to about 10 mg/ml, from about 0.1 mg/ml to about 10 mg/ml, from about 0.5 mg/ml to about 5 mg/ml, from about 0.5 mg/ml to bring the flax 3 mg/ml, from about 0.5 mg/ml to about 2 mg/ml, from about 0.5 mg/ml to about 1.5 mg/ml, from about 0.8 mg/ml to about 3 mg/ml, from about 0.8 mg/ml to about 2 mg/ml, from about 0.8 mg/ml to about 1.5 mg/ml, from about 0.7 mg/ml to about 3 mg/ml, from about 0.7 mg/ml to about 2 mg/ml, from about 0.7 mg/ml to about 1.7 mg/ml, about 0.7 mg/ml, about 1.5 mg/ml, about 0.7 mg/ml, about 1.4 mg/ml, about 0.7 mg/ml, about 1.3 mg/ml, about 0.5 mg/ml, about 0.7 mg/ml, about 0.8 mg/ml, about 0.9 mg/ml, about 1 mg/ml, about 1.1 mg/ml, about 1.2 mg/ml, about 1.3 mg/ml, about 1.4 mg/ml, about 1.5 mg/ml, approximately 1.6 mg/ml, about 2 mg/ml, about 3 mg/ml, about 4 mg/ml, about 5 mg/ml, about 6 mg/ml, about 7 mg/ml, about 8 mg/ml, about 9 mg/ml, about 10 mg/ml or about 15 mg/ml inside the liposomes.

The electric potential in the plane of shear is called the Zeta-potential of liposomes. As you know an experienced, Zeta-potential of liposomes can be experimentally determined by using the appropriate equipment, for example, from what uranium using ELS-600 (Otsuka Electronics, Japan), using the method of laser-Doppler microelectrophoresis or other equipment and protocols available for an experienced specialist. For example, J.Colloid and Interface Sci.,39, 670-675 (1972), nition.com/en/products/zeecom_s.htm etc.

In some embodiments described herein liposomes (including liposomes targeted action and liposomes containing liposomes of the composition) must have a cumulative negative Zeta-potential. In some embodiments of the Zeta-potential is from about -10 mV to about -200 mV. For example, from about -50 mV to about -150 mV, from about -50 mV to about -130 mV, from about -60 mV to about -120 mV, from about -50 mV to about -100 mV, from about -75 mV to approximately -90 mV, from about -80 mV to about -90 mV, from about -80 mV to about -85 mV, from about -85 mV to approximately -90 mV, from about -75 mV to approximately -85 mV, from about -70 mV to approximately -90 mV, approximately 75 mV, approximately -80 mV approximately -85 mV, approximately -83 mV, approximately -90 mV, approximately -100 mV, approximately -120 mV.

I believe that after intravenous small liposomes they pass through window sinusoidal liver and usually quickly come into contact with hepatocytes. L is bosomy intermediate size, believed to be trapped in the compartment and blood can circulate for a considerable period of time. However, large liposomes are slower through the sinusoids of the liver and quickly captured kupferschmid cells. Thus, the size of the liposomes is very important to determine the behavior in vivo. See, for example, Liu et al., Biochim. Biophys. Acta (1992)1104(1):95-101; Harashima et al., J. Drug Target. (1995)3(4):253-261 (which is incorporated herein by reference in its entirety), etc.

The size of the liposomal particles can be obtained on the basis of the correlation function by using different algorithms when using photon correlation spectroscopy (PCS; dynamic light scattering or quasielastic light scattering (QELS)). Obtained by using these methods, the particle size is comparable with the average diameter is defined using the PCS. When PCS to describe the size distribution using standard deviation and χ2. In systems PCS χ2determines whether the system is unimodal (Gaussian distribution) or multimodal (Nicomp distribution). The average particle size can be determined by the intensity-weighted measurements and is described on the basis of a Gaussian distribution with χ2≤5. If χ2>5 then use the average of the main peak of the distribution Nicopm. Such an analysis should be the ü mark of an experienced specialist, as well as hardware, for example, Nicomp QELS Particle Sizer, PSS Model 380ZLS, S/N 0103301; pssnicomp.com/zetaspec.htm.

In some embodiments of liposomes, particularly liposomes directional average diameter described here liposomes can be from about 50 to about 275 nm. For example, the average diameter of the liposomes can be from about 50 to about 200 nm, from about 50 to about 265 nm, from about 50 to about 250 nm, from about 50 to about 225 nm, from about 50 to about 175 nm, from about 50 to about 150 nm, from about 50 to about 120 nm, from about 50 to about 100 nm, from about 75 to about 250 nm, from about 75 to about 200 nm, from about 75 to about 175 nm, from about 75 to about 150 nm, from about 75 to about 120 nm, from about 75 to about 100 nm, from about 90 to about 100 nm, from about 90 to about 120 nm, from about 90 to about 150 nm, from about 90 to about 200 nm, from about 95 to about 100 nm, from about 95 to about 120 nm, from about 95 to about 125 nm, from about 95 to about 130 nm, from the roughly 95 to about 150 nm, from about 95 to about 175 nm, about 90 nm, about 95 nm, about 100 nm, about 120 nm, about 130 nm, or about 150 nm. For specific liposomal compositions liposome directed action must have a diameter larger than that from approximately 15 to approximately 25 nm diameter liposomes formed from the same components, but without the inclusion of the guiding factor.

Described here liposomes (for example, liposomes directional unloaded liposomes liposomes in containing liposome compositions can also be characterized by the concentration of the sending of the ligand, which is included in the liposome. Depending on the selected guide ligand experienced specialist should be obvious different means of determining the number of the sending of the ligand. For example, as described in the examples, the content of transferrin (Tf) in the liposomes can be determined by electrophoretic mobility (for example, when measuring using Na-DDS-PAGE) liposomes compared to the respective controls.

Briefly, the confirmation of the content of transferrin in the liposomes can be estimated using two analyses of the content and/or identity of transferrin conjugated to lipsome. First electrophoretic mobility transferrin liposomes, Ana is sireau using Na-DDS-PAGE, you can compare with the nature of the mobility of the purified conjugated transferrin, for example, TF-NωPE. In addition, electrophoretic mobility conjugated transferrin can also be compared with the free transferrin as a reference standard. Additional confirmation of the identity of transferrin in the liposomes can be obtained using ELISA, composition skills to research that shows specific binding of antibodies against the transferrin with liposomes for targeted delivery. Concentration directed by transferrin liposomes can be measured using colorimetric assays of protein, such as BCA, analysis, well known to experienced specialist. An experienced specialist in the light of the guidelines presented here should also be borne in mind similar methods and other known methods in the art to determine the number of different guiding factors.

In short, the amount of transferrin in the liposome can be analyzed using the analytical reagent bicinchoninic acid (BCA). Copper (II) restored to copper (I) protein in alkaline conditions. The resulting ion copper (I) forms soluble intensively coloured complex with BCA. The total associated with the microparticle protein measured by interaction known is th number of a suspension of microparticles with BCA reagents. After the formation of the coloring particles are removed by filtration, and the color is measured spectrophotometrically.

In some embodiments, the concentration of the guide ligand included in the liposome, should be from about 0.5 mg/ml to about 5.0 mg/ml, from about 0.5 mg/ml to about 2.0 mg/ml, from about 1.0 mg/ml to about 2.0 mg/ml, from about 1.0 mg/ml to about 3.0 mg/ml, from about 1.0 mg/ml to about 2.5 mg/ml, from about 1.0 mg/ml to about 2.0 mg/ml, or from about 1.3 mg/ml to about 2.5 mg/ml

The role of Fe ion is very important for the binding of transferrin to the surface of tumor cells. Therefore, the content of Fe ion in liposomes targeted actions, including Tf, is another important way the characteristics of the liposomes. Specialists in the art should be aware of the number of ways of determining the content of iron ions, one method is ICP-MS.

When liposome containing transferrin, Fe ion content in the liposome may comprise, for example, from about 0.25 microgram/ml to about 3 mg/ml, 0.4 μg/ml to about 3 mg/ml, 0.25 μg/ml to about 2 μg/ml, 0.25 μg/ml to about 1.5 μg/ml, 0.25 μg/ml to about 1 μg/ml, 0.4 μg/ml to arr is siteline 2 μg/ml, from 0.4 μg/ml to about 1.5 μg/ml, 0.5 μg/ml to about 2 μg/ml, from about 0.5 μg/ml to about 1.4 μg/ml, or from about 0.5 μg/ml to about 1.5 μg/ml

Liposomes, including liposomes targeted action and unloaded liposomes can also be characterized by their osmotic pressure at a given temperature. The osmotic pressure at a given temperature depends on the molar concentration of sugar solution (sucrose). And it also depends on the total density of ions and the size of molecules in solution. Typically, the osmotic pressure can be measured using a device known as osmometer, which measures the osmotic pressure in a suitable pressure units, as it should be known to experienced specialist.

In some embodiments the osmotic pressure of liposomes, particularly liposomes directional and non-loaded liposomes at room temperature should be from about 310 to about 410 mOsm/kg, for Example, the osmotic pressure can be from about 310 to about 400 mOsm/kg, from about 310 to about 380 mOsm/kg, from about 320 to about 360 mOsm/kg, from about 315 to about 375 mOsm/kg, from about 320 to about 375 mOsm/kg, from about 315 to p is blithedale 370 mOsm/kg, from about 320 to about 370 mOsm/kg, about 360 mOsm/kg, about 350 mOsm/kg, about 340 mOsm/kg, approximately 370 mOsm/kg, or about 380 mOsm/kg at room temperature.

Under various conditions described here (for example, storage conditions, preparation for the introduction and/or in vitro) osmotic pressure may vary less than about 25%, less than about 20%, less than about 15%, while monitoring for a specific period of time associated with various conditions described here. For example, 360±50 mOsm/kg

Receiving lipid containing compositions

There are three main requirements for drug substances and labeled compounds designed to introduce individuals in the course of therapy, namely efficiency, safety and quality assurance. Despite the proven efficacy and safety, the application of medicinal substances in the treatment of vague, if its quality (e.g., purity, uniformity, reproducibility, dose, stability over time, and so on) cannot be guaranteed during production and distribution. Methods of obtaining drugs or labeled compounds, which do not provide appropriate high quality Lek is stannage means, also increases the likelihood of adverse reactions in individuals who inject drugs. During the lifetime of drug or labeled compound, it is important that the product that is being produced and distributed, had the same standards as the product that originally received the permit sanctions. Thus, the ability to properly receive the drug or labeled compound the high quality necessary to obtain safe products, such as drug or labeled compound, drug or labeled compound, which can be received and cleared routine and easy, stand as commercial prospects and security. When, for example, the effectiveness of the labelled compound refers to its ability to be suitable for the diagnosis of specific diseases or conditions in combination with specific diagnostic methods (for example, activity of the labeled compound (e.g., ability to be visualized in a gamma-counter and so on), does not deteriorate from an unacceptable level from batch to batch or during storage.

Below describes the most common methods of obtaining compositions described herein, which can be applied to obtain the corresponding high quality (n is an example, high purity, homogeneity, etc.) liposomes directed action (and their intermediates) and non-loaded liposomes. These methods are also represented schematically in Fig. 4 (a method of obtaining (A) and 5 (method get (B).

Operations esters derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, as here described, are suitable as components of phospholipid complexes such as liposomes, polymeric micelles, micro - and nanospheres, emulsions and water-soluble polymers. Here's how these PE derivatives and methods for their preparation are also known in the art, as noted above.

In particular, operations ester derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine also suitable as components described herein lipid containing compositions. Lipid containing compositions can be obtained in accordance with the ways described here, although modifications of these methods should also be obvious to a person skilled in the art. For example, various methods known to the person skilled in the art can be used for preparation of liposomes from lipid components (e.g., processing ultras what UCU, stirring, extrusion, dehydration etc). For example, as described in patent. the application of the USA, publ. No. 2004/0142025, the contents of which are incorporated herein by reference in its entirety.

The application described here are common methods that are included in the examples to obtain liposomes directed action also covers methods of obtaining other lipid containing compositions (for example, lipid mixtures containing liposome compositions, unloaded liposomes and intermediate liposomes), as described here.

The method of obtaining A

The method of obtaining And depicted schematically in figure 4.

A: the mixture NωPE:SuccNωPE:additional lipid (intermediate 1)

Operations ester derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine (SuccNωPE) and derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine (NωPE), as described here, is mixed with additional(mi) lipid(DAMI) (for example, at least one phospholipid(AMI) (for example, PC (e.g., DMPC, DSPC, etc.), PI, sphingomyelin, phosphatidic acid, etc.)) and at least one additional lipid (e.g., cholesterol), then dissolved in a suitable solvent (e.g. ethanol, T-BuOH, chloroform, isopropyl ether and so on). The amount of solvent is usually from 1 to 100.mass. (with respect to the total mass of the lipids). In certain embodiments it is from 2 to about 20./mass.

Used here as described in the preceding paragraph, SuccNωPE and NωPE can be obtained and purified as described here means or methods known to experienced specialist. SuccNωPE and NωPE, as well as other components described herein to obtain liposomes directed action must have clarity and uniformity sufficient to obtain at the end of liposomes directional with purity and uniformity necessary to comply with the guidance for the introduction of individuals liposomes directed action and compliance with the guidelines for acceptable laboratory practice (GLP) and acceptable manufacturing practice (GMP).

When (in addition to one or more phospholipids) use additional(e) lipid(s) in combination with phospholipid, phospholipid(s) to another(them) more(YM) lipid(s) is approximately 2:1. Mixing ratio derived SuccNωPE to phospholipid is from about 1 to about 12% (from 1:99 to 12:88), or approximately 3-6% (from 3:97 to 6:94) from the total concentration/ratio (NHS-NGPE + NGPE) to (CHOL + phospholipid). For example, illustrative attitude includes 50:45:4:1 (for example, PC:Chol:NG-PE:NHS-NG-PE), where, for example, in the mixture are of 17.5 mg NHS-NG-DOPE, 63,1 mg NG-DOPE, 312 mg Chol and 607 mg of phospholipids.

B: Receive the mixture of the medicinal substance: NωPE:SuccNωPE:additional lipid (intermediate 2)

The mixture NωPE:SuccNωPE:additional lipid obtained at the previous stage A, then mixed with an aqueous solution (e.g., buffer and so on)that contains a drug or labeled compound, subject to encapsulation, for example, an anti-cancer agent (e.g., oxaliplatin, an inhibitor of topoisomerase I, a Vinca alkaloid, etc.), with a mixture of the medicinal substance:NωPE:SuccNωPE:neutral lipid (intermediate 2).

When the drug is oxaliplatin (l-OHP), the concentration of the solution oxaliplatin is approximately 8 mg/ml in approximately 9% sucrose solution. For example, the concentration oxaliplatin in liposomes directed action is approximately 0.8 mg/ml ± 10%.

C: Obtain liposomes drug substance:NωPE:SuccNωPE:additional lipid (intermediate 3)

A mixture of the medicinal substance:NωPE:SuccNωPE:additional lipid (intermediate 2)obtained in stage B, then voice or mix with subsequent evaporation of the solvent with the formation of liposomes drug substance:NωPE:SuccNωPE:additional lipid (intermediate 3). Methods and conditions for scoring, mixing and evaporation and the means of implementation data the x stages are well understood, experienced, as also described in the Examples section below. See, for example, methods of obtaining, using methods bubbles with reversed phase (REV), U.S. patent No. 4235871 (incorporated herein in its entirety by reference). Can also be applied to General methods of obtaining liposomes, such as how simple hydrogenation and ways of introduction of ethanol, known to experienced specialist.

Formed, as described above, the liposome drug substance:NωPE:SuccNωPE:additional lipid next ekstragiruyut in size and allocate the liposome drug substance:NωPE:SuccNωPE:additional lipid. For the concentration of the solution of the liposomes can then be optionally applied ultrafiltration.

When liposome containing l-OHP (drug substance), DMPS (additional lipid/phospholipid (neutral)), cholesterol (CHOL, additional lipid/neutral lipid), N-glutaryl-DOPE (NG-DOPE) and NHS-NG-DOPE, you can select the liposome with an average diameter of approximately 0.2 micrometer (200 nm). Similar size of the liposomes can be also obtained in the case of liposomes containing l-OHP (drug substance), DSPC (additional lipid/phospholipid (neutral)), cholesterol, N-glutaryl-cells of the dspe (NG-cells of the dspe) and NHS-NG-cells of the dspe. Illustrative guide number of lipid components comprise, for example, about 40 mg/ml DMPC (additional lipid/phosphatidylcholine/FOSFA the lipid/neutral lipid), approximately 20 mg/ml CHOL (additional lipid/neutral lipid) and approximately 5 mg/ml of NG-DOPE (the total number of NG-PE and NHS-NG-PE). An illustrative value for the lipid components is 50:45:5 (additional lipid 1 (e.g., phosphatidylcholine): additional lipid 2 (e.g., CHOL): NG-PE (for example, NG-DOPE + NHS-NG-DOPE).

D: Getting liposomes drug substance:NωPE:TF-NωPE:additional lipid (liposomes directed action)

The liposome drug substance:NωPE:SuccNωPE:additional lipid, formed as described in stage C, can then be functionalitywith using the guiding factor for choosing to obtain liposomes drug substance:NωPE:TF-NωPE:additional lipid (also denoted "liposome directional").

Attach the guide factor (TF) (for example, functionalization of the intermediate liposomes (intermediate 3) guiding factor) exercise covalent joining of the guiding factor to SuccNωPE through interaction operations of the residue with a guiding factor. By means of suitable reaction conditions, operations groups on the exposed surface of the liposomes (on the outer surface of the lipid bilayer, where the drug or labeled compound is encapsulated inside liposomes) can be covalent what about modified with the formation of the modified guiding factor derivatives of N-(ω)-dicarboxylic acid and phosphatidylethanolamine (TF-NωPE). Attaching the guiding factor to the liposome leads to the formation of liposomes drug substance:NωPE:TF-NωPE:additional lipid (liposomes directed action).

Specifically, Succinimidyl carboxyl residue SuccNωPE, as described here, functionalitywith under suitable conditions. If the guide factor contains the amino group(s), amino group(s) of the guiding factor enter into interaction with Succinimidyl carboxyl residue with the formation of amide linkages carboxylic acid. Conditions suitable for this reaction are further outlined here, including in the Examples section below, and should be well understood by an experienced specialist. An experienced specialist should also be able to modify the reaction conditions to optimize them for specific combinations of the guiding factor and liposomes, without undue experimentation, based on the data here for instructions.

Various guides factors, as described herein and known to the experienced professional, can be obtained from commercial sources or obtained by methods known to the expert.

When, for example, as a guiding factor in choosing transferrin, transferrin can be obtained from commercial sources in the form of a purified protein, for example, from Celliance Corp., GA, USA. Transferrin can also be obtained by using rivers is minantly ways well known in the art (for example, by applying the prokaryotic cells (E. coli etc), the use of eukaryotic cells (CHO, BHK, etc. and so on). As is well known, transferrin can be obtained and used in the liposomes directed action either APO-or holo-form. Alternatively, liposomes directed action, including APO-transferrin, can be treated with compounds of iron, such as iron citrate, iron chloride (III) and so on, obtaining liposomes directed action, including liposome, a modified holo-transferrin.

Illustrative amounts of transferrin as a guiding factor is, for example, approximately 2 mg/ml transferrin. This should be sufficient for illustrative liposomes directional, comprising about 40 mg/ml DMPC, approximately 20 mg/ml CHOL and about 5 mg/ml of NG-DOPE (the total number of NG-PE and NHS-NG-PE).

After functionalization of liposomes (intermediate 3) guiding factor, as described herein, to obtain the liposomes directional obtained liposomes may not need to be additionally purified by methods known to experts in the art, including as described here cleaning methods, particularly described above in connection with stage C.

When lipase is and contains l-OHP (drug substance), DMPS, cholesterol (CHOL), N-glutaryl-DOPE (NG-DOPE) and Tf-NG-DOPE, you can select the liposome with an average diameter of from about 0.05 micrometer to about 0.2 micrometer (from approximately 50 nm to approximately 200 nm). Similar size of the liposomes can be also obtained in the case of liposomes containing l-OHP (drug substance), DSPC, cholesterol, N-glutaryl-cells of the dspe (NG-cells of the dspe) and NHS-NG-cells of the dspe.

Obtaining liposomes directed action described above (referred to for convenience as a way of obtaining (A) reproducibly gives liposomes directional high purity and homogeneity. In particular, liposomes directional substantially free from non-NHS starting compounds (described here) and by-products (for example, acylated compounds of urea and so on)associated with the formation of SuccNωPEs. In particular, the production and purification SuccNωPEs before the formation of liposomes gives liposomes (intermediate 3) and liposomes, which are focused largely free from carbodiimide initial substances (e.g., DCC, EDC and so on)used for functionalization NωPEs for education SuccNωPEs. As mentioned earlier, drug and labeled compounds, including liposomes targeted action, comprising a medicinal substance and labeled compounds, which are designed to introduce the individual to the minds in the course of treatment or diagnosis, must be of high quality.

Optional lipid mixture, described in A (intermediate 1)can be processed guiding factor with the formation of lipid mixtures containing TF-NωPE. Alternatively, the lipid mixture can be mixed with a water solution with the formation containing the liposome composition. Finally, containing the liposome composition can be processed by receiving liposomal composition. Optional aqueous solution may include drug or labeled compound.

An alternative way to obtain (method B)

The method of obtaining B schematically depicted in Fig. 5.

A. Obtaining liposomes drug substance:NωPE:additional lipids

Liposomes directed action, as described here, can also be obtained by dissolving additional lipids and NωPE in a suitable solvent (e.g. ethanol, tert-BuOH, chloroform, isopropyl ether, etc.), dispersing the resulting solution in an aqueous solution, optionally containing a drug or labeled compound, and the subsequent processing of the obtained dispersion ultrasound or formation of vesicles with reversed phase with obtaining liposomes (drug substance:NωPE:neutral lipids). A solution of liposomes can be concentrated by ultrafiltration.

In the operation of a non-limiting example of a method of obtaining liposomes formation of vesicles with reversed phase (REV) (U.S. Patent No. 4235871, included as a reference). Of course, can also be applied to conventional methods of obtaining liposomal compositions, such as how simple hydration and injection of ethanol.

For stable retention NωPE(s) in the lipid bilayer NωPE(s) can be accessed and cleaned, and then NωPE(s) together with additional lipids (e.g., phospholipid(s), cholesterol etc) is used to produce liposomes in accordance with methods known to experienced specialist.

As a non-limiting example is the mixing and dissolving in a suitable solvent additional lipids, for example, one or more phospholipids (e.g., DSPC, DMPC, and so on), another optional additional lipid (e.g., cholesterol) and at least one NωPE.

When additional lipid is a phospholipid and cholesterol, the ratio of mixed phospholipid and cholesterol may be, for example, approximately 1:1, for example, about 1.1:1, about 1.2:1, ~ 0.9:1 (e.g., DMPC and cholesterol 50:45 (mol%). Content NωPE(s) as a share of total lipid content is, for example, 6% with respect to the phospholipid. The resulting solution is mixed with a solution oxaliplatin in aqueous buffer. Content NωPE can be from about 0.8 mol% to about 12 mol% of BEGO lipid content. For example, from about 1 mol% to about 10 mol%, from about 1 mol% to about 8 mol%, from about 1 mol% to about 6 mol%, from about 1 mol% to about 5 mol%, from about 1 mol% to about 4 mol%, from about 1 mol% to about 3 mol%, from about 1 mol% to about 2 mol%, from about 2 mol% to about 12 mol%, from about 2 mol% to about 10 mol%from about 3 mol% to about 8 mol%, about 1 mol%, about 2 mol%, about 3 mol%, about 4 mol%, about 5 mol%, about 6 mol%, about 8 mol%, about 10 mol%, or about 12 mol%.

The concentration of drug or labeled compound in the solution may be as described above and, in particular, as described above in the method of obtaining A. similarly, a solution containing a drug or labeled compound, described here includes components of the solution.

Liposomes containing l-OHP (drug substance), DSPC, cholesterol and N-glutaryl-cells of the dspe (NG-DOPE)obtained by this method, can be selected with the receipt containing oxaliplatin liposomes (e.g., using gel filtration, chromatography with the exception of size, ultrafin the ation, of ultracentrifugation and so on), having an average diameter of approximately 0.2 μm.

B. Obtaining liposomes drug substance:SuccNωPE:NωPE:additional lipids

After stage A part NωPE in the liposome obtained in stage A (the liposome drug substance:NωPE:additional lipids), functionalitywith obtaining liposomes, including SuccNωPE (i.e. liposomes drug substance:SuccNωPE:NωPE:additional lipids), which can then be modified with the formation of the TF-NωPE.

For education SuccNωPE carboxyl group at the end of NωPE modify of obtaining operations of the group. Such functionalization may be carried out by the methods described for obtaining SuccNωPE(s).

For example, in the presence of liposomes injected into the interaction of carbodiimide (e.g., EDC, DCC, etc.) and N-hydroxysultaine (NHS) to obtain the liposomes drug substance:SuccNωPE:NωPE:additional lipids.

C. Obtaining liposomes drug substance:TF-NωPE:NωPE:additional lipids

After stage B obtained in stage B liposome drug substance:SuccNωPE:NωPE:additional lipids enter into interaction with a guiding factor with the formation of liposomes drug substance:TF-NωPE:NωPE:additional lipids. The methods and conditions of the reaction are the same as described in the s for a method of obtaining A stage D.

Obtained according to the method of obtaining B liposome drug substance:TF-NωPE:NωPE:additional lipids can be purified and concentrated described herein and known experienced ways.

Comparison of methods of obtaining

The method of obtaining A has a number of advantages compared to the method of obtaining B, although to obtain liposomes drug substance:TF-NωPE:NωPE:additional lipids (liposomes directed action, not necessarily containing either drug or labeled compound) can be used both ways. The most important thing is that liposomes obtained by method A, should be free or substantially free from contamination (e.g., non-NHS starting compounds and/or by-products)associated with obtaining SuccNωPEs. In particular, as noted earlier, liposomes directional obtained by the method of obtaining A should be free or substantially free from, for example, carbodiimides (e.g., EDC, DCC etc) and acylated derivatives of urea. In some embodiments, when SuccNωPE not include the interior of the liposome, the liposome or containing the liposome composition can be free or substantially free from the NHS. In addition, the larger the scale of the reaction, the longer time is of Holocene. Time for a method of obtaining A considerably less time for a method of obtaining a B.

Although the clearance of liposomes drug substance:TF-NωPE:NωPE:additional lipids obtained using the method of obtaining B should reduce the amount of pollutants, such liposomes (for example, liposomes drug substance:SuccNωPE:NωPE:additional lipids obtained in stage B of the method of obtaining B) harder to clean than lipids (for example, SuccNωPEs obtained and purified to the stage of A method of obtaining (A). As part SuccNωPE, apparently, is focused inside liposomes (e.g., functional group operations ether complex is located on the inner side of the lipid bilayer and is not available for interaction with the guiding factor), it seems probable that the liposomes directional obtained by the method of obtaining A, may contain some residual amount stored in SuccNωPE.

A further advantage of the method of obtaining A is that the content of TF-NωPE relatively NωPE end liposomes drug substance:TF-NωPE:NωPE:additional lipids can be controlled more precisely in the application of the method of obtaining A. the Relative amounts of these lipids directly correlated with the relative amounts SuccNωPE and NωPE used as starting substances at the stage of A method of producing A. That is they way the number of modified TF SuccNωPE also can be controlled more precisely.

When applying the method of obtaining a B number NωPE on the number SuccNωPE depends on the efficiency of the reaction stage B of method B. This reaction is considered to end when the content NωPE in the liposome of approximately 10%, but can be expected experimental variation between batches. The low efficiency of the reaction, probably partly due to steric hindrance pre-formed liposomes. When SuccNωPE is formed from the selected NωPE (lipid), steric hindrance is much smaller, and the reaction goes to completion. In addition, after the formation of the SuccNωPE (only in lipid form) the product obtained can be purified from the reaction mixture, which thereby provides the removal of unreacted NωPE, carbodiimide and NHS, as well as other by-products that may form during the reaction.

Liposomes formed using any method, apparently, are more homogeneous (and therefore can be used to obtain a more reproducible drug/diagnostic product) in comparison with liposomes that include PEG or other hydrophilic polymers, such as described in the prior art of the present description the Oia. Usually, when PEG or other hydrophilic polymers are used to increase the circulation time of liposomes (e.g., to protect liposomes from capture RES), their use leads to the formation of liposomes with an extended range of molecular weight due to the wide distribution of PEG or hydrophilic polymers. This distribution increases the complexity associated with manufacturing (e.g., reproducibility, and/or cleaning), and may also increase the variability of clinical effectiveness. Liposomes directional obtained by methods A or B, should have advantages in these respects.

Additional methods of obtaining

The lipid mixture and containing the liposome composition (which can be used to produce liposomes) can also be obtained by modification of the methods of obtaining A and B. for Example, in some embodiments the lipid mixture and containing the liposome composition, which includes an additional(e) lipid(s) component(s):NωPE:TF-NωPE or more(e) lipid(s) component(s):NωPE:TF-NωPE:medicinal substance/labeled compound (where "additional(e) lipid(s) component(s)" refers to one or more phospholipids (e.g., one or more neutral, one or more anionic, one or more cationic phospholipids or combinations of two or more of these you who e), optionally additionally comprising one or more additional lipids as described herein (e.g., cholesterol or its derivative); or at least two different neutral lipid as described herein (e.g., at least one phospholipid (for example, PC (e.g., DMPC, DSPC, etc.), PI, sphingomyelin, phosphatidic acid, etc. and at least one other neutral lipid (e.g., cholesterol), can be obtained using the methods of preparation, described below, as well as other modifications ways supposed to be an expert in the light of the guidance provided in this description. Components additional lipids, NωPE, TF-NωPE and, if present, drug or labeled compound may be such as described in the present description. Similarly, the relative amounts of the components are such as described in the present description.

In some embodiments the lipid mixture obtained in the first stage of the method of obtaining B (lipid mixture obtained by dissolving additional lipids and NωPE in a suitable organic solvent)may be modified by the inclusion of the NHS and then modified TF obtaining lipid mixtures extra(s) of the lipid(s) component(s):NωPE:TF-NωPE. This lipid mixture can then be mixed with an aqueous solution (containing optional Leka is the only substance or labeled compound) with the formation containing the liposome composition. Alternatively, the drug or labeled compound may be included after receipt containing the liposome composition. In some embodiments of the drug or labeled compound, free from water solution, may be included in the lipid mixture formed after the modification using the NHS and TF, with the formation of lipid mixtures extra(s) of the lipid(s) component(s):NωPE:TF-NωPE:drug or labeled compound. This lipid mixture can then be mixed with an aqueous solution with the formation containing the liposome composition.

In some embodiments the lipid mixture obtained in the first stage of the method of obtaining B (lipid mixture obtained by dissolving additional(s) of the lipid(s) component(s) and NωPE in a suitable solvent, may be mixed with the aqueous solution with the formation containing the liposome composition (extra(s) of the lipid(s) component(s):NωPE). This contains the liposome composition can then be processed NHS and TF and then mix with the drug or labeled compound with the formation containing the liposome composition of the additional(s) of the lipid(s) component(s):NωPE:TF-NωPE:medicinal substance/labeled compound. Alternative containing the liposome composition of the additional(s) of the lipid(s) component(s):NωPE you can handle Lek is rstendamm substance or labeled compound and then modify the NHS and then TF.

In some embodiments the lipid mixture obtained in the first stage of the method of obtaining B (lipid mixture obtained by dissolving additional(s) of the lipid(s) component(s) and NωPE in a suitable solvent)may be mixed with the drug or labeled compound (optional comprising an aqueous solution) with the formation of the lipid mixture (where the drug or labeled compound does not include an aqueous solution or containing the liposome composition. When a lipid mixture, the lipid mixture can then be processed NHS and TF with the formation of lipid mixtures extra(s) of the lipid(s) component(s):NωPE:TF-NωPE:medicinal substance/labeled compound, which can then be mixed with an aqueous solution with the formation containing the liposome composition. Alternatively, when formed containing the liposome composition (for example, when a drug or labeled compound is included in an aqueous solution), this contains the liposome composition can then be processed NHS and TF also with the receipt containing the liposome composition of the additional(s) of the lipid(s) component(s):NωPE:TF-NωPE:medicinal substance/labeled connection.

In an additional alternative method of obtaining, method C, the individual components are simultaneously mixed in an organic solvent with the formation of the lipid is a mixture of (C-1)(for example, ingredients: extra(s) of the lipid(s) component(s); NωPE; TF-NωPE or components: extra(s) of the lipid(s) component(s):NωPE:TF-NωPE:medicinal substance/labeled compound), where TF-NωPE get and need not be cleaned prior to mixing. The lipid mixture of C-1 can then be mixed with an aqueous solution with the formation containing the liposome composition C-2 (extra(s) of the lipid(s) component(s):NωPE:TF-NωPE, optionally containing a drug or labeled compound). When formed in this way contains the liposome composition contains no drug or labeled compound, drug or labeled compound can be added after the formation containing the liposome composition (C2-A). Alternatively, when the source component is used comprising an aqueous solution of a medicinal substance or labeled compound containing the liposome composition can be formed simultaneously with the mixing of all of the original components.

C-2 (optional containing the drug or labeled compound) or C2-A can be treated with the formation of liposomes (C-3). When C-3 does not include drug or labeled compound, the liposome will be unloaded by the liposome, as described previously (e.g., liposome additional(s) of the lipid(s) component(s):NωP:TF-NωPE). When C-3 includes a drug or labeled compound, the liposome will liposome directed action, as described here. When C-3 is unloaded by the liposome, drug or labeled compound may be as described previously added to the non-loaded liposome at the next stage, with the formation of liposomes directed action that can be performed immediately after receiving the C-3 or some time later, which may include the retention period unloaded liposomes C-3.

In an additional alternative method of obtaining, method D, the individual components are simultaneously mixed in an organic solvent to form a lipid mixture (D-1) (e.g., ingredients: extra(s) of the lipid(s) component(s); NωPE; or components: extra(s) of the lipid(s) component(s):NωPE:drug or labeled compound). The lipid mixture D-1 can then be mixed with an aqueous solution with the formation containing the liposome composition D-2 (extra(s) of the lipid(s) component(s):NωPE:(optional containing the drug or labeled compound). Containing the liposome composition D-2 can then be mixed with TF-NωPE with the formation containing the liposome composition D-3 (extra(s) of the lipid(s) component(s):NωPE:TF-NωPE(optional containing medicinal substances which STV or labeled compound)), where TF-NωPE get and need not be cleaned prior to mixing. When formed in this way contains the liposome composition contains no drug or labeled compound, drug or labeled compound can be added after the formation containing the liposome composition (D3-A). Alternatively, when the source component using drug or labeled compound, comprising an aqueous solution containing the liposome composition can be formed simultaneously with the mixing of all components.

D-3 (optional containing the drug or labeled compound) or a D3-A can then be processed to obtain liposomes (D-4). When D-4 does not include drug or labeled compound, the liposome will be unloaded by the liposome, as described previously (e.g., liposome additional(s) of the lipid(s) component(s):NωPE:TF-NωPE). When D-4 includes drug or labeled compound, the liposome will liposome directed action, as described here. When D-4 is unloaded by the liposome, drug or labeled compound may be as described previously added to the non-loaded liposome at the next stage, with the formation of liposomes directed action that can be performed immediately after receiving D-4, or SP is the degree for some time, which may include the retention period unloaded liposomes D-4.

As in the case of lipid mixtures containing liposome compositions and liposomes (including liposomes targeted action, unloaded liposomes, etc.)formed by the method of obtaining A lipid containing composition obtained by the method of obtaining a C or D must be substantially free from non-NHS source materials, by-product and/or product decay associated with the synthesis operations of ester derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine (for example, carbodiimides (e.g., DCC, EDC etc), acylated compounds of urea and so on), as far as the original substance (e.g., TF-NG-PE) largely free from these substances to the initial stage of the method of obtaining a C or method D. When SuccNωPE not include as a source of the substance (and, therefore, do not include inside liposomes, liposome or containing a liposome composition can also be free or substantially free from the NHS, as in the case of prior learning and use as the initial substance TF-NωPE. In some embodiments lipid containing composition obtained by the method of obtaining a C or method D, substantially free from DCC and EDC. In some embodiments lipid containing compositions, recip is by way of obtaining a C or method D, largely free from DCC.

In some additional embodiments(s) of the lipid(s) component(s) include(s) one or more phospholipids (e.g. phosphatidylcholine, etc.) and cholesterol or a derivative of cholesterol. In some embodiments, the phosphatidylcholine is a DMPC, POPC, DSPC, etc. as described herein. In some embodiments, the phosphatidylcholine is a DMPC or DSPC. In some additional embodiments, lipid(s) are phospholipid and cholesterol. In some embodiments, the phospholipid is a neutral phospholipid.

In some additional embodiments(s) of the lipid(s) component(s) include(s)at least two different neutral lipid, which include phospholipids (e.g. phosphatidylcholine, etc.) and cholesterol or a derivative of cholesterol. In some embodiments, the phosphatidylcholine is a DMPC, POPC, DSPC, etc. as described herein. In some embodiments, the phosphatidylcholine is a DMPC or DSPC. In some embodiments, at least two different neutral lipid represent a phospholipid and cholesterol.

In some embodiments NωPE is a NG-PE. In some embodiments NωPE is an Nω-DOPE or Nω-cells of the dspe. In some embodiments NωPE is a NG-DOPE or NG-cells of the dspe.

In some embodiments TF represents,for example, asialoglycoprotein, folate, transferrin, etc. In some embodiments TF is a transferrin (Tf). In some embodiments TF-NωPE is a Tf-NωPE (for example, Tf-NG-DOPE or Tf-NG-cells of the dspe).

In some embodiments the medicinal substance is, for example, an anti-cancer agent (e.g., oxaliplatin, an inhibitor of topoisomerase I, a Vinca alkaloid, and so on). In other embodiments the lipid mixture, or containing the liposome composition comprises labeled compound. In some embodiments the lipid mixture, or containing the liposome composition does not include a labeled compound or drug.

As mentioned earlier, each of the lipid mixtures can be mixed with a water solution with the formation containing the liposome compositions each containing the liposome compositions can be processed with the formation of the corresponding liposomes (for example, liposomes directed action (e.g., drug or labeled compound), intermediate liposomes, unloaded liposomes and so on), as described here.

As for the variations described are methods of obtaining, it is understood that modification NωPE with the NHS, modification NHS-NωPE using TF, receipt containing the liposome compositions of the lipid mixtures and obtain liposomes containing liposome compositions can the t to be carried out by an experienced technician, as described herein, without undue experimentation, taking into account the guidance offered in the present description, including, in particular, a detailed description of how to obtain A and B, and as shown in the examples.

Pharmaceutical compositions

In another aspect of the present invention offers pharmaceutical compositions for the treatment or diagnosis of vulnerable individuals, including lipid containing compositions as described herein and one or more pharmaceutically acceptable carriers, excipients, diluents, stabilizers, preservatives, or other inactive ingredients, including combinations of the above-mentioned, known, experienced professionals and additionally described here.

In some embodiments pharmaceutical compositions containing the lipid composition is a liposome directed action, as described here. In other embodiments containing the lipid composition is a containing liposome composition. In some embodiments containing the lipid composition is an unloaded the liposome. In some embodiments, the composition includes the medicinal substance. In other embodiments, the composition includes labeled connection.

In some embodiments, the carrier may include one or more of sterile water, buffer solution or fiziologicheskogo the solution, diluent or combinations thereof.

The pharmaceutical compositions can further include one or more of the various salts, sugars, proteins, starch, gelatin, vegetable oils, polyethylene glycol, and the like, including combinations of two or more of the above.

An additional aspect of the invention includes the use of compositions and formulations as described herein for the manufacture of a medicinal product. In particular, the manufacture of a medicinal product for use in treating or diagnosing conditions, as described herein. In addition, active compositions and formulations described herein with the various parties that are also designed for use in the manufacture of a medicine for use for the treatment or diagnosis of the conditions and in accordance with the described herein means, unless specified otherwise.

The use of formulations

Introduction

As previously noted, in one aspect provides methods of treating or diagnosing conditions, as described here, with the application containing the drug or labeled compound containing lipid compositions (e.g., liposomes directional containing drug or labeled compound containing the liposome compositions) and pharmaceutical compositions, as described herein.

In one embodiment, the methods can mytreasury in the form of a therapeutic approach to the treatment described here States. Thus, in a particular embodiment containing the medicinal substance containing lipid compositions or pharmaceutical compositions can be used to treat these conditions in vulnerable individuals, including humans. The methods typically include the introduction of individual compositions or composition described herein in an amount effective to treat the condition.

In another embodiment, the methods can be implemented in the form of a diagnostic approach to diagnosis described here States. Thus, in a particular embodiment containing the labeled compound containing lipid compositions or pharmaceutical compositions can be used to diagnose these conditions in vulnerable individuals, including humans. The methods typically include the introduction of individual compositions or composition described herein in an amount effective to diagnose the condition. This introduction is usually produced in conjunction with the methods to detect the condition.

In some embodiments the individual is a mammal, including, but not limited to, human, cow, horse, cat, dog, rodent, or Primate. In other embodiments the individual is the man.

The terms "pharmaceutically effective amount" or "therapeutically effective amount" refers to the number of the TSS of the composition, sufficient for the treatment of certain disorders, conditions or diseases, or one or more symptoms and/or prevent diseases or disorders. In relation to cancer pharmaceutically or therapeutically effective amount comprises an amount sufficient to, among other things, for the induction of growth reduction or to reduce the rate of tumor growth.

The terms "an amount effective to diagnose"or "diagnostically effective amount"or "amount effective for diagnosis" are close to each other and refer to the amount of the composition sufficient to establish the diagnosis of certain disorders, conditions or diseases and/or one or more of its manifestations, where the diagnosis includes the establishment of the disease and/or determining the degree or severity of the disease. For example, in the treatment of cancer "diagnostically effective amount" includes an amount sufficient for detection, for example, the presence and/or concentration of one or more malignant cells, tumor(s) or other manifestations of cancer. Often diagnosis should be performed with respect to the basal level or background level determination observed in individuals without the condition. The levels of determination in excess of background or basal levels (elevated levels of definition), serving as the indication, and in some cases the severity of the condition.

When it is used in relation to methods of treatment and use containing the medicinal substance containing lipid compositions, "in need" an individual may be an individual, who has already been diagnosed with the condition or who has previously been treated in the condition to be treated. In relation to methods of diagnosis and application containing the labeled compound compositions "in need" an individual may be the individual who suspect status, has the risk condition (for example, a family history of the condition, lifestyle factors indicating risk status (e.g., Smoking as a risk factor for lung cancer and so on)or who was previously diagnosed condition (for example, the diagnosis may include the monitoring of gravity (for example, the progression/regression of disease in time and/or therapy).

In some embodiments the state of the subject to treatment or diagnosis is cancer. In some embodiments the cancer can be cancer of the stomach, colon, colon, rectum, or breast cancer. In some embodiments the cancer is a breast cancer. In other embodiments the cancer is a cancer of the stomach. In some embodiments the cancer is a pancreatic cancer, non-small cell lung cancer, melcochita the hydrated lung cancer, brain cancer, liver cancer, kidney cancer, prostate cancer, bladder cancer, ovarian cancer or hematological malignancy (e.g., leukemia, lymphoma, multiple myeloma etc).

Containing the medicinal substance compositions, including compositions described herein can be used separately or in combination (e.g., before, simultaneously with or after) with other treatment modalities (for example, the additional cancer therapy, treatment of combined methods). For example, in combination with other therapeutic agents (e.g., anticancer chemotherapeutic agents, as described herein and known to experts in the art (e.g., alkylating agents, takanami, metabolic antagonist, anticancer antibiotics, plant alkaloids, drug for hormone therapy, the drug with the target molecule, etc.)), surgery and/or radiotherapy. When subjected to a treatment condition is cancer, compositions described herein can be introduced in combination with one or more anti-cancer agents or cytotoxic compounds, as described herein and as known in the art, one or more additional agents to reduce the occurrence and/or severity of adverse reactions and/or their manifestations, x is surgical operation (for example, removal of the tumor or lymph nodes etc) or irradiation. When one or more surgical procedures or radiation to form part of a treatment regimen, the composition can be entered before, simultaneously or after radiotherapy or surgery. Similarly, the compositions and formulations as described herein can be entered before, concurrently or after administration of one or more anti-cancer agents. Liposomes directed actions and their compositions described herein can also be entered in combination (e.g., before, simultaneously with or after) with medicinal substances to relieve symptoms associated with the condition or treatment regimen (e.g., medicinal substances to suppress vomiting, hair loss, immunosuppression, diarrhea, rashes, sensitivity disorders, anemia, fatigue, stomatitis syndrome hand-foot and so on). Liposomes directed action can also enter more than one stage (including over) regimens (for example, after surgery with or after radiotherapy and so on).

Containing the labeled compound compositions, including compositions described herein can be used separately or in combination (e.g., before, simultaneously with or after) methods of treatment (for example, the additional cancer therapy, treatment of combined methods). For example, composition and can be used to monitor the success of treatment. For example, to determine whether detected whether subject to treatment before, after or simultaneously with the treatment regimen (as described above with respect to methods of treatment).

In some embodiments the composition is administered before or after surgery (such as removal of the tumor or lymph nodes etc). In other embodiments the composition is administered after surgery and prior to, simultaneously with or after radiotherapy. The optimal combination of one or more of surgery and/or radiotherapy in combination with the introduction of the compositions described herein, and optionally one or more chemotherapeutic agents can be determined by the attending physician, depending on the specific case and taking into consideration various factors that affect a specific individual, including those described here.

In some embodiments containing the medicinal substance compositions or pharmaceutical compositions can be introduced in combination with one or both of the 5-toralla and/or leucovorin. In other embodiments containing the medicinal substance composition or pharmaceutical compositions can be introduced in combination with one or more other anti-cancer drug, such as capecitabine, UFT/LV, tegafur-uracil and leucovorin, irinotecan, antibody against EGFR (e.g., zetoc IMAP etc), the anti-VEGF antibody (e.g., Avastin and so on), a tyrosine kinase inhibitor (e.g., erlotinib), etc. This introduction can also be combined with treatment, including radiotherapy and/or surgery. In some embodiments enclosed in the liposome directional drug is oxaliplatin.

In connection with the described here by way of application containing lipid compositions or pharmaceutical compositions of the present invention can impose dropped. Parenteral administration can be produced by bolus injection (in/in), infusion (in/in), intraperitoneal injection or by local injection (such as intracranial injection). In some embodiments of the introduction is by bolus injection or continuous infusion.

Continuous intravenous infusion may last for minutes or hours. For example, but not limited to, from about 10 minutes to about 5 hours, from about 15 minutes to about 4 hours, from about 30 minutes to about 4 hours, from about 45 minutes to about 4 hours, from about 60 minutes to about 4 hours, from about 45 minutes to about 3 hours, from about 60 minutes to about 2 hours, from about 90 minuto approximately 3 hours from about 90 minutes to about 2 hours, about 10 minutes, about 15 minutes, about 20 minutes, about 30 minutes, about 45 minutes, about 50 minutes, about 60 minutes, 80 minutes, about 1.5 hours, about 2 hours, about 2.5 hours, about 3 hours, about 3.5 hours, about 4 hours, about 5 hours, about 12 hours, about 24 hours, about 36 hours, or about 48 hours.

Composition and dosage

As mentioned above, containing the lipid compositions and pharmaceutical compositions, as described herein, can be entered in need individuals to treatment or diagnosis of these conditions in relation to the here described methods of use.

Described herein containing lipid composition and, in particular, described herein liposomes directed action usually must be used in amounts effective to achieve the desired result, for example, in an amount effective for treatment or prevention of a subject to treatment of a particular condition. Song(s) you can enter a therapeutically to achieve therapeutic action. Therapeutic effect means suppressing or alleviating subjected to treatment of the underlying violation and/or suppression or alleviation of one or more proc is Ohm, associated with the primary disease, resulting in the patient reported improved health or condition, notwithstanding that the patient may suffer from underlying disease. Therapeutic action also includes a stop or slowing of disease progression, regardless of whether there is an improvement.

In some embodiments, when subjected to a treatment condition is a cancer, an effective amount is an amount sufficient to reduce tumor growth (e.g., as measured by the rate of increase in the average volume of tumors before and/or after treatment). In some embodiments an effective amount is the amount sufficient to decrease the average tumor volume (for example, when the average tumor volume after treatment is reduced in comparison with the average tumor volume before treatment).

The number of tracks to be entered for the introduction of an effective amount of the encapsulated drug substance (for example, oxaliplatin), must depend on many factors, including, for example, want to treat a specific condition, the route of administration, the severity of the subject to treatment status, age, and weight of the patient, the bioavailability of the composition, the expected adverse effects in the subject of the treatment of the individual and so the effective dose also find the camping in the competence of specialists in the art in light of the proposed instructions here.

In some embodiments, the dose of encapsulated oxaliplatin at a specific point in time must be in the range of from about 1 to about 400 mg/m2/day. For example, in the range of from about 1 to about 350 mg/m2/day, from 1 to about 300 mg/m2/day, from 1 to about 250 mg/m2/day, from 1 to about 200 mg/m2/day, from 1 to about 150 mg/m2/day, from 1 to about 100 mg/m2/day, from about 5 to about 80 mg/m2/day, from about 5 to about 70 mg/m2/day, from about 5 to about 60 mg/m2/day, from about 5 to about 50 mg/m2/day, from about 5 to about 40 mg/m2/day, from about 5 to about 20 mg/m2/day, from about 10 to about 80 mg/m2/day, from about 10 to about 70 mg/m2/day, from about 10 to about 60 mg/m2/day, from about 10 to about 50 mg/m2/day, from about 10 to about 40 mg/m2/day, from about 10 to about 20 mg/m2/day, from about 20 to about 40 mg/m2/day, from about 20 to about 50 mg/m2/day, from about 20 to AP is sustained fashion 90 mg/m 2/day, from about 30 to about 80 mg/m2/day, from about 40 to about 90 mg/m2/day, from about 40 to about 100 mg/m2/day, from about 80 to about 150 mg/m2/day, from about 80 to about 140 mg/m2/day, from about 80 to about 135 mg/m2/day, from about 80 to about 130 mg/m2/day, from about 80 to about 120 mg/m2/day, from about 85 to about 140 mg/m2/day, from about 85 to about 135 mg/m2/day, from about 85 to about 135 mg/m2/day, from about 85 to about 130 mg/m2/day, from about 85 to about 120 mg/m2/day. Dose, administered at a specific point in time, can also be approximately 130 mg/m2/day, about 120 mg/m2/day, about 100 mg/m2/day, about 90 mg/m2/day, approximately 85 mg/m2/day, about 80 mg/m2/day, about 70 mg/m2/day, about 60 mg/m2/day, about 50 mg/m2/day, about 40 mg/m2/day, about 30 mg/m2/day to about 20 mg/m2/day, about 15 mg/m2/day or priblisitelno mg/m 2/day.

Enter the dose may be higher or lower than described here ranges of doses, depending, among other factors, the bioavailability of the composition, the stability of the individual to adverse side effects, route of administration and various factors discussed above. The magnitude of the dose and interval may be adjusted individually to provide levels of composition in plasma, sufficient to maintain therapeutic effect, in accordance with the decision of the attending physician. Experienced professionals should be able to optimize effective local dosages without undue experimentation in light of the proposed instructions here.

Dose can also be determined using animal models in vivo, which should be clear to experts in the field of technology.

Multiple doses (e.g., continuous or bolus) of the compositions as described herein can also be entered in need individuals for hours, days, weeks, or months. For example, but not limited to, daily, every other day, every 10 days, weekly, monthly, twice a week, three times per week, twice per month, three times a month four times a month, five times per month, after month, every third month, every fourth month and so on

Sets

There are also kits for the introduction described the here and compositions including pharmaceutical compositions, including compositions.

In some embodiments the kits may include unit dose (for example, used for therapy or diagnosis), at least, one containing the lipid composition or pharmaceutical composition as disclosed here. The kits may additionally include suitable packaging and/or instructions for use of the composition. The kits can also include means of delivery of a composition or pharmaceutical composition, such as a syringe for injection or other device as described herein and known to specialists in this field of technology.

In some embodiments the kits may include unit dose (for example, used for therapy or diagnosis) unloaded liposomes or its pharmaceutical composition as disclosed here. The kits may additionally include suitable packaging and/or instructions for use of the composition. The kits can also include means of delivery of a composition or pharmaceutical composition, such as a syringe for injection or other device as described herein and known to specialists in this field of technology. In addition, in some embodiments, the kit may contain a unit dose of drug or labeled compound to be included in an unloaded the liposome.

In addition, containing the ipid composition or pharmaceutical composition can be assembled in kit form. In the set of features containing the lipid composition or pharmaceutical composition and reagents for producing compositions for injection. The composition may be in dry lyophilized form or in solution, in particular a sterile solution. When the composition is in dry form, the reagent may include a pharmaceutically acceptable diluent, to obtain a liquid composition. Such diluents include those known to experts in the art, for example, solutions of sugar, such as dextrose, sucrose, etc. In some embodiments, the solution may include a solution of sugar with a sugar concentration from about 1% to about 20%, from about 1% to about 18%, from about 1% to about 15%, from about 1% to about 10%, from about 3% to about 10%, from about 3% to about 6%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 12%, about 15%, about 18%, or approximately 20%. In some embodiments, the solution may be a solution of dextrose (with a concentration of dextrose, for example, approximately 1%, approximately 2%, approximately 5% etc). In some of the s incarnations containing the lipid composition can be represented as, for example, the liposome directional, unloaded the liposome, the lipid mixture, or containing the liposome composition (optionally containing a drug or labeled compound).

The kit may also contain a device for insertion or for the separation of compositions, including, but not limited to, a syringe, pipette or other device known in the art. In liquid form, the composition can be stored in vials or other sterile closed container, including those known to specialists in this field of technology.

Kits can include other therapeutic compounds for use in combination with the compounds described herein. In one embodiment, therapeutic agents are other anticancer agents. These agents may be proposed in a separate form or in a mixture with compounds of the present invention, proposed so that such mixing does not reduce the effectiveness of any of the additional therapeutic agents described herein compositions and formulations. Similarly, the kits can include additional agents for adjunctive therapy. For example, agents to reduce the adverse effects of drug substance (for example, protivotoshnotnoe agents, agents against baldness, immunostimulatory agents, and so on).

Kits which contain instructions for the preparation and introduction of the composition, information about the side effects of compositions and other relevant information. The instructions may be in any suitable format, including, but not limited to, printed material, videotape, machine-readable disk, or optical disk.

In another aspect of the invention provides kits for the treatment of an individual suffering from these conditions or susceptible thereto, comprising a first container that includes a metered quantity containing the lipid composition or compositions as described herein and instructions for use. The container may be any known in the art and suitable for storage and delivery of intravenous formulations. In some embodiments, the kit further includes a second container comprising a pharmaceutically acceptable carrier, diluent, adjuvant, etc. to obtain a composition to be administered to the individual.

May also offer kits that contain doses disclosed here compositions or formulations, sufficient to ensure effective treatment of the individual over an extended period of time such as a week, 2 weeks, 3 weeks, 4 weeks, 6 weeks, 8 weeks, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months or more.

The kits may also include multiple doses containing whether the ID of the composition or compositions and instructions for use, Packed in quantities sufficient for storage and use in pharmacies, for example pharmacies with clinics and prescription pharmacy.

All cited here the patents, patent applications and publications included in the present description by reference in its entirety.

Examples

The present invention is additionally described with reference to the following examples; these examples, however, do not limit the scope of the present invention.

Example 1: Test for cytotoxicity oxaliplatin

The solution oxaliplatin (l-OHP) was obtained by dissolving oxaliplatin in a 9% sucrose solution (sucrose/distilled water) at a concentration of 8 mg/ml cell Viability was determined using a commercially available kit for analysis of cytotoxicity (WST-I kit, Wako Pure Chemical Industries, Ltd., Japan).

Cells AsPC-1 (provided by Dr. Hironobu Yanagie of the Research Center for Advanced Science and Technology, the University of Tokyo, Japan)and cultured in medium RPMI 1640 with the addition of 10% FCS (serum fruits calves; SIGMA, USA), treated by solutions of l-OHP concentrations [200 × (l/2)(0-10)nm] at 37 ° C in 5% CO2within 48 hours. After that, the medium was removed, and added the substrate (WST-I, Cell Counting Kit, Dojindo Laboratories, Japan) to the cells, which were incubated at 37 ° C in 5% CO2within 2 hours for the appearance of a colored product. The developed color was measured by absorption at 450 nm (the end of the control wavelength: 620 nm) Measurement Mini NJ-2300 (Cosmo Bio Co., Ltd., Japan).

The results are shown in Fig. 6. It was found that the cytotoxicity of l-OHP is LD50> 8 mcg/ml

Example 2: Determine the number of transferrin receptors on the cell surface

The experiment used a normal human leukocytes and originating from a malignant tumor human cell lines (K562, MKN45P and HL60), obtained from the following sources: K562:TKG0210 (Cell Resource Center for Biomedical Research, Institute of Department, Aging and Cancer, Tohoku University, Japan); MKN45P: Dr. Hisae Iinuma, Teikyo University School of Medicine, Japan; HL60:TKG0345 (Cell Resource Center for Biomedical Research, Institute of Department, Aging and Cancer, Tohoku University, Japan).

The number of receptors Tf (transferrin) on the surface of each cell type was determined using katchanovski analysis (Comp. Biochem. Physiol., 116B, 137-160 (1949), using Microsoft Excel). Solution125I-labeled Tf (Na125I (PerkinElmer Japan Co., Ltd., Japan) and h-Tf (T-4132, SIGMA, USA) was connected iodophenyl method (Biochem. Biophys. Res. Commun., 122, 319-325 (1984)), was added to each cell culture at various concentrations in the range [300 × (1/2)(0-9)nm] at 4C, and incubated for 1 hour.

Concentration125I-labeled Tf was determined by quantitative measurement of protein by Lowry method (J. Biol. Chem., 193, 265-270 (1951)), and radioactivity was measured using a gamma counter (Auto Well Gamma System ARC-300, Aloka Co., Ltd., Japan). Briefly, the solution was centrifuged to precipitate the cells, the cell fraction was washed chilled on ice SFR (180 × g (gravity) for 3 min, that was repeated 3 times, followed by measurement of radioactivity using a gamma counter to determine the concentration of Tf associated with the cell surface). The number of cells was determined by quantitative measurement of protein using the Lowry method (J. Biol. Chem., 193, 265-270 (1951)).

For each experimental point, the concentration of unbound Tf was determined by subtracting the concentration of bound Tf of known concentration added to Tf. Katchanovski graph was obtained by applying the concentration of bound Tf on the horizontal axis and the relationship of the concentration of bound Tf to the concentration of unbound Tf on the vertical axis. The amount of bound Tf (i.e. the number of receptors) were determined from the intersection with the x axis of the graph, as described in Proc. Natl. Acad. Sci USA, 80 2263-2266 (1983); J. Cell Physiol, 132, 492-500 (1987); Proc. Natl. Acad. Sci USA, 92 3318-3322 (1995); J. Pharm. Sci, 84, 216-220 (1995); Eur. J. Biochem., 186, 367-373 (1989); J. Biol, Chem., 258, 4715-4724 (1983), which here by reference in its entirety.

The number of125I-Tf associated with the cell surface of various cell types shown in Fig. 7. It was found that the number of receptors for transferrin (Tf) on the cell surface of the cell lines derived from human malignant tumors, significantly higher than in normal leukocytes.

Example 3: Receiving NHS-NG-DOPE

200 mg NG-DOPE (Avanti Polar Lipids, Inc., USA) (Cat. No.. 870242,M.M. 880,13) was weighed in a conical vessel with 2 conclusions. To the vessel was added 39,2 mg NHS (Sigma, USA, mm = 115,09). Then added 5 ml of a mixture of chloroform/ethyl acetate (1:1(vol./vol.), Wako Pure Chemical Industries, Ltd., Japan) and mixed by rotation for the early dissolution of NG-DOPE and the NHS. Was observed a slight haze.

After the initial mixing was added rod stirrer, the vessel was gently purged with gaseous nitrogen (filled with gaseous nitrogen cylinder) through one of the conclusions of the vessel was closed with a rubber stopper. Stirring in nitrogen atmosphere produced using rod stirrer and magnetic stir bar. The second conclusion was closed by the tube. The reaction was conducted at room temperature (20-23ºC). The mixture was stirred for 5-10 minutes. Took 20 μl sample of the reaction mixture lipid + NHS for use in quality control at TLC.

In a separate vessel received a solution of DCC (99%, Aldrich, USA, M.M.: 206,33 g/mol) by dissolving 70 mg of DCC in 5 ml of ethyl acetate. DCC was dissolved in the solvent rapidly with the formation of a transparent solution. Thus obtained solution of DCC (approximately 5 ml) then was added dropwise to the reaction mixture, the lipid/NHS for 10-15 minutes. After addition of DCC, the reaction mixture became more and more muddy.

TLC was performed with control (lipid/NHS) and an aliquot of the lipid/NHS/DCC at time 0 as reference points the following about the time. A sample of 50 µg (2,5 μl of a solution of 20 mg/ml) was applied on the plate for TLC (aluminium plate silica gel 60F254from EM Science (Gibbstown, NJ, USA) cat. No.. SP05554M), dried and then placed in a processing chamber in which the solvent (a mixture of 70% chloroform, 28% methanol, 2% water) could move. Pointed to the front of the solvent, and then the TLC plate was dipped in ammonium molybdate (5% ammonium molybdate in 10% H2SO4and dehydrational using the drying apparatus.

The reaction mixture lipid/NHS/DCC was stirred under a stream of nitrogen and the formation of product (Rf 0.3 to 0.4) was monitored in time.

After 18 hours the transformation in the NHS-NG-DOPE was not full, and added additional number of NHS (26 mg in 2 ml ethyl acetate) and DCC (47 mg in 1 ml ethyl acetate). The course of the reaction was again tested by TLC in time 20 hours.

The reaction was let go over the weekend at ambient temperature while blowing with nitrogen and stirring (with protection from light). Before cleaning was still some amount of substance.

Clearance: The reaction mixture was cooled on ice for ~30 minutes. The cooled reaction mixture is then filtered through BONEROWSKI funnel and then washed 3 times with 2×5 ml of chloroform. All the obtained liquid was collected and dried on a rotary evaporator. After evaporation there was obtained semi-solid paste. Pas who have then resuspendable in 2-3 ml of chloroform.

Cleaning suspended paste on silica gel was performed using a 4 g silica (400 mesh), wetted with chloroform. The silica gel was Packed in a column 1 cm × 28 cm with shutoff valve. The approximate size of the layer was 1 cm × 14 cm Column was balanced with chloroform (packing under gravity).

The sample was applied to a balanced (but not dried) column of silica gel. In a column was added 10 ml of chloroform (5×2 ml). Collected fractions 5×2 ml of the current Velocity was determined by gravity, but the fraction of 5×10 ml were collected within 10-20 min and labeled fractions 1-5.

Then the column was applied to 50 ml (5×10 ml) mixture of chloroform/methanol (90/10, about./vol.). Collected fractions 5×10 ml, which were designated as fractions 6-10.

After collecting fractions 6-10 column was applied a mixture of chloroform/methanol (5/1, (about./about.)) a volume of 100 ml (10×10 ml). Collected additional fractions of 10×10 ml, which were designated as fractions 11-15.

Fractions 6-15 (5 µl aliquots) were analyzed by TLC as described above.

After TLC fractions 6-15 fractions 7-11 were combined and dried to a thin film using a rotary evaporator. The final product obtained after evaporation, had a weight of 130 mg (yield 65%)as it was determined by TLC when compared with the crude product of the reaction and comparison with the standard product (NHS-NG-DOPE), obtained from NOF (Japan is).

Example 4: Receiving NHS-NG-DOPE

Previously received and cleared NG-DOPE (200 mg) (NOF Corporation, Japan) and NHS (N-hydroxysultaine; 34 mg) was weighed and placed in a 5 ml conical vessel with 2 holes. One hole was closed with a rubber stopper, and through the remaining hole was added rod stirrer.

The vessel was then placed in a vacuum and carefully filled the stream of gaseous nitrogen (repeated three times). The vessel was then left in an atmosphere of nitrogen when using a cylinder with nitrogen.

After placing in a nitrogen atmosphere in the vessel was added 2.5 ml of anhydrous chloroform and stirred using a rod stirrer and magnetic stir bar. The reaction was carried out at ambient temperature for about 30 minutes, and to dissolve the starting compounds used rotation. Took 20 μl of the sample mixture lipid + NHS for use in controlling/monitoring the reaction by TLC.

Then got the solution by dissolving 61 mg of DCC (1,3-dicyclohexylcarbodiimide) in 2.5 ml anhydrous chloroform (quickly formed a clear solution). The DCC solution was added dropwise to a mixture of lipid/NHS for 15 minutes. After addition of DCC solution was Motel.

At time 0 was performed TLC (70% chloroform, 30% methanol, 5% water) mixture of lipid/NHS and lipid/NHS/DCC to monitor the reaction by application of 50 mg of chloroform on the plate for TLC, prussiani and space in the reaction chamber (70% chloroform, 30% methanol, 5% water) to move.

The reaction mixture was continued to stir under a stream of nitrogen and the formation of product (Rf 0.3 to 0.4) was monitored in time.

The reaction was allowed to take place for 2-3 days at ambient temperature while blowing with nitrogen and stirring.

The reaction mixture was then filtered through BONEROWSKI funnel and washed twice with 2×5 ml of chloroform. The entire solution was collected and dried on a rotary evaporator. Was obtained semi-solid paste.

Semi-solid paste resuspendable 2×3 ml of chloroform and then filtered and dried. This procedure was repeated three times. Finally, after three times the product resuspendable 2×3 ml of chloroform.

Column of silica gel was obtained by mixing silica with chloroform, followed by packing in a column 1 cm × 28 cm with shutoff valve. The approximate size of the layer column was 1 cm × 14 cm Column was balanced with chloroform (packing under gravity).

Samples were applied to a balanced (but not dried) column of silica gel. Then the column was applied 100 ml of chloroform, and collected in aliquots of 100 ml fractions. The flow rate was determined by gravity (fraction 1).

Fraction 1. The column was applied to 100 ml of a mixture of chloroform/methanol (90/10, about./vol.). Collected in 100 ml fractions (fraction 2).

The column was applied to 200 ml (20×10 ml) the mixture is chloroform/methanol (50/10, about./vol.). Collected 20×10 ml fractions (fractions 3-23).

Fractions 1 through 23 were analyzed using an aliquot of 5 ml by TLC.

Fractions 9 through 22 were combined and dried to a thin film using a rotary evaporator and liofilizirovanny. The final weight of the NHS-NG-DOPE of these fractions accounted for 61.9 mg (yield of 27.9%).

Example 5: Receiving lipid mixture (NG-DOPE:Tf-NG-DOPE:DMPC:CH)

583 mg DMPC (NOF corporation, Japan), 299 mg of cholesterol (Wako Pure Chemical Industries, Ltd., Japan) and to 75.7 mg NG-DOPE (NOF corporation, Japan) were mixed and dissolved in t-BuOH (10 wt./about. with respect to lipids (10 ml) at 45-50°C.

The resulting solution was poured into a vessel and froze about 8 hours on the shelf at -40°C. the Pressure was reduced to approximately 0.1 mm, RT. Art. and the vessel was left under reduced pressure for 2 days in a stepwise increase in temperature from -40°C to 25°C, and the result of this process was obtained liofilizovannye lipid mixture.

Obtained as described above, the powder liofilizovannyh lipid mixture was mixed with 20 mg of powdered Tf-NG-DOPE (obtained as in example 29) and milled. Thus was obtained a homogeneous powder lipid mixture with a lipid ratio of 50:45:5 (DMPC:Chol:NG-DOPE+Tf-NG-DOPE).

Example 6: a Receipt containing the liposome compositions

The lipid mixture was obtained according to the previous examples with the components shown below:

Part 1: DMPC/Chol/NG-DOPE (155 mg/79,4 mg/16,1 mg)

Part 2: DMPC/Chol/NG-DOPE (155 mg/79,4 mg/l6,l mg)

Part 3: DMPC/Chol/NG-DOPE/NHS-NG-DOPE (l52 mg/77.9 mg/15.8 mg/of 4.38 mg)

Part 4: DMPC/Chol/NG-DOPE/NHS-NG-DOPE (152 mg/77.9 mg/15.8 mg/of 4.38 mg)

Part 5: DMPC/Chol/NG-DOPE/Tf-NG-DOPE (148 mg/76,0 mg/15,4 mg/4,8 mg)

Party 6: DMPC/Chol/NG-DOPE/Tf-NG-DOPE (148 mg/76,0 mg/15,4 mg/4,8 mg)

Each of the parts 1, 3 and 5 were hydrational and stirred with 300 mm aqueous solution of sucrose (about 20./mass. with respect to lipids (5 ml)(5 ml of sucrose solution (20 vol./mass.) added to the dry lipid mixture and stirred for 30 min at 40-45°C. Each of the parties 2, 4 and 6 were hydrational and mixed with an aqueous solution of l-OHP (8 mg l-OHP/ml (about 20./mass. with respect to lipids (5 ml) in 300 mm sucrose solution for 30 min at 40-45°C. Thus was obtained containing the liposome mixture.

The diameter of the liposomes was determined using QELS, and the results are presented on Fig. Liposomes in containing the liposome mixture have an average diameter of 500-2000 nm and have a wide size distribution in the field of 100-10000 nm.

Example 7: a Receipt containing oxaliplatin liposomes (NG-DOPE:Tf-NG-DOPE:DMPC:CH)

The composition of the liposomes was as follows:

Dimyristoylphosphatidylcholine (1,2-dimyristoyl-sn-glycero-3-phosphocholine: DMPC) (NOF Corporation, Japan)

Cholesterol (CH) (Solvay Pharmaceuticals B.V., the Netherlands)

N-hultarilldricteltnoh (N-glutaryl-1,2-dioleoyl-sn-glycero-3-phospho shall canoemen, sodium salt: DOPE-CO- (CH2)3-COOH, denoted hereafter as NG-DOPE) (NOF Corporation, Japan)

Succ-N-hultarilldricteltnoh (N-(succinimidylester)-1,2-dioleoyl-sn-glycero-3-phosphoethanolamine, sodium salt: DOPE-CO-(CH2)3-CO-OSu; referred to hereafter as the NHS-NG-DOPE) (NOF Corporation, Japan)

DMPC:CH:NG-DOPE:NHS-NG-DOPE = 50:45:4:1 (m/m).

As the aqueous phase used an aqueous solution of l-OHP (8 mg/ml in 300 mm sucrose solution).

A mixture of DMPC, CH, NG-DOPE and NHS-NG-DOPE (molar ratio of 50:45:4:1) was dissolved in 4 vol./mass. (with respect to the total mass of lipids) hot solvent which is a mixture of ethanol/tert-butanol/water. The lipid solution was injected into 300 mm sucrose solution containing about 8 mg/ml of l-OHP at around 45C, so that the concentration of the solvent was approximately 14% vol./about.

The suspension was passed through an extruder, an indoor five pieces of filter 100 nm (Cat. No. 112105, Whatman plc, UK) at a pressure of approximately 200-800 pounds per square inch at about 45°C. Thus was obtained liposome having an average diameter of about 100 nm. The diameter of the liposomes was determined by QELS.

6 l phosphate buffered saline (pH 7,9), 6 l of a solution of transferrin (Cat. No.. 4455, Selorogicals, GA, USA) and 18 l of a suspension of liposomes were mixed and stirred at 30 ° C for 15-60 minutes uh what about gave the reaction mixture, containing 4 mg/ml transferrin and 20 mg/ml lipid.

Quantitative analysis of transferrin was performed using tests with bicinchoninic acid (ICA) in accordance with the instructions provided by seller.

The increase in molecular weight after switching transferrin was investigated using DDS-Na-PAGE (polyacrylamide gel electrophoresis with sodium dodecyl sulfate). Analysis of NG-DOPE was performed using high performance liquid chromatography (HPLC) with evaporative light scattering detector (ELSD2000, Alltech, MD, USA) using a column of silica gel (YMC PVA Silica Column of 4.6×250 mm, 5 μm).

Example 8: a Receipt containing oxaliplatin liposomes (NG-Cells of the dspe:Tf-NG-cells of the dspe:DSPC:CH)

The composition of the liposomes was as follows:

Distearoylphosphatidylcholine (1,2-distearoyl-sn-glycero-3-phosphocholine: DSPC)

Cholesterol (CH)

N-glutamyltranspeptidase (N-glutaryl-1,2-distearoyl-sn-glycero-3-phosphoethanolamine, sodium salt: cells of the dspe-(CH2)3-COOH, denoted hereafter as NG-cells of the dspe)

DSPC:CH:NG-cells of the dspe=2:1:0.2 (mol/mol).

As the aqueous phase used an aqueous solution of l-NRO (8 mg/ml in a 9% sucrose solution), as described in example 1.

A mixture of DSPC (M, NOF, Japan), cholesterol (038-03005, Wako Pure Chemical Industries, Ltd., Japan) and NG-cells of the dspe (Dr. Kazuo Maruyama, Teikyo University, Faculty of Pharmaceutical Sciences, Japan) in the ratio 2:1:0,2 (m/m) was dissolved in chloroform and isopr Pilbeam alcohol.

To the resulting solution was added a solution of l-OHP (9% sucrose solution), and then the resulting mixture was treated with ultrasound for about 15-30 minutes. The solution is then evaporated using a rotary evaporator at 60°C to remove solvent and frozen/thawed five times. The suspension was frozen (by immersion in a bath of dry ice/acetone) and thawed (immersion in warm water). This was repeated five times.

Then the obtained product was sorted by size at 60°C using filters for extruder (twice at 400 nm and then five times with 100 nm) (LipexTM Extruder, Model No. T-001, Northern Lipids Inc., Canada) and was ultracentrifugal (200000×g, 60 min, approximately 4°C). Sediment resuspendable in a 9% sucrose solution or MES buffer (pH 5.5) (MES buffer. Cat. No. 345-01625, Dojindo Laboratories, Japan) with a receipt containing l-OHP liposomes NG-cells of the dspe:DSPC:CH.

Then implicating l-OHP the liposome NG-cells of the dspe:DSPC:CH modified with transferrin (Tf). To the thus obtained containing l-OHP the liposome NG-cells of the dspe) was added l-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC; Cat. No. #22980, Pierce Biotechnology, Inc., USA) (average of 2.7% by weight of lipid) and N-hydroxysultaine (S-NHS; 038-0432, Wako Pure Chemical Industries, Ltd., Japan) (average of 7.3% by weight of lipid), and the mixture was left at room temperature for 10 minutes.

Then to the resulting process is the added transferrin (Tf), approximately 20% by weight of the lipid (Cat. No. T4132, SIGMA, USA), and stirred at room temperature for 3 hours. Was added a solution of transferrin (Tf) (in an amount of about 20% of the total volume of the reaction mixture) in 1 mm SFR (phosphate buffered saline) and 1 mm SFR (in the amount of approximately 20% of the total volume of the reaction mixture, and the resulting solution was stirred at room temperature for 1 hour.

To the thus obtained suspension APO-form liposomes Tf-NG-cells of the dspe was added 10-40 EQ. (with respect to transferrin) citrate, iron sodium citrate (Wako Pure Chemical Industries, Ltd., Japan) and was stirred at room temperature for 15 minutes. The resulting solution was subjected to ultrafiltration as described above. The precipitate is then resuspendable in a 9% sucrose solution with obtaining thereby holo-form liposomes Tf-NG-cells of the dspe. The solution was subjected to ultrafiltration (200000 × g, 60 min, approximately 4°C), and then the precipitate resuspendable in a 9% sucrose solution.

Quantitative analysis of transferrin was performed using tests with bicinchoninic acid (BCA)made in accordance with the instructions of the seller (Cat. No. 23227, BCATMProtein Assay Kit, Pierce Biotechnology, Inc., USA).

The increase in molecular weight after modification was investigated using DDS-Na-PAGE (electrophor is in polyacrylamide gel with sodium dodecyl sulfate). Analysis of NG-cells of the dspe was performed using high performance liquid chromatography (HPLC) with evaporative light scattering detector (ELSD2000, Alltech, MD, USA) using a column of silica gel (YMC PVA Silica Column of 4.6 × 250 mm, 5 μm).

Example 9: Getting PEG containing oxaliplatin liposomes

In accordance with the Protocol of the experiment in example 8 were obtained liposomes of DSPC:cholesterol:cells of the dspe-PEG(2K)-OMe:cells of the dspe-PEG(3.4K)-COOH (Tf-PEG-liposomes). In these liposomes the ratio of the components was as follows: DSPC:cholesterol:cells of the dspe-PEG(2K)-OMe:cells of the dspe-PEG(3.4K)-COOH=2:1:0,16:0,03.

This liposome contained 6 mol% PEG-lipid and 1 mol% PEG-COOH-lipid, and Tf is attached to the liposome through PEG-COOH.

By way of example 8 were obtained liposomes Tf/PEG-cells of the dspe (Tf/PEG-NG-cells of the dspe liposomes).

In these liposomes the ratio of the components was as follows: DSPC:cholesterol:cells of the dspe-PEG(2K)-OMe:NG-cells of the dspe=2:1:0,16:0,03.

This liposome modified with PEG, and Tf is attached to the liposome through NG-cells of the dspe. Derivatives of PEG-liposomes can also be obtained by the methods described in patent applications U.S. publication Nos. 2003/0224037 and 2004/0022842, the disclosure of which is incorporated here in their entirety.

Example 10: Getting unloaded liposomes

A mixture of DMPC, Chol (Wako Pure Chemical Industries, Ltd., Japan), NG-DOPE (NOF Corporation, Japan) and NHS-NG-DOPE (NOF Corporation, Japan) (in a molar ratio of 50:45:4:1; 410 g of DMPC, 211 g Chol, 43 g of NG-DOPE and 12 g of NHS-NG-DOPE, respectively) was dissolved in 4 vol./mass. (what about the relation to the total weight of lipid) warm solvent ethanol/tert-butanol/water. The resulting suspension with a volume of 20 l were incubated at 45°C under stirring and was passed through an extruder (Stevested Machinery &Engineering Ltd., Canada), which was covered with five layers of 100 nm polycarbonate filters (Cat. No. 112105, Whatman plc, UK) at a pressure of approximately 200-800 pounds per square inch at about 45°C. Thus was obtained liposomes having an average diameter of about 100 nm. The diameter of the liposomes was determined by QELS.

A suspension of liposomes, SFR buffer (pH 7,9) and the solution in TFR transferrin (Cat. No. 4455, Selorogicals, GA, USA) (pH 7.0) were mixed in a ratio of 3:1:1 (vol./vol.), then was stirred for 15-60 min at 30°C. Thus was obtained approximately 6 l unloaded liposomes.

20 grams (approximately 19 ml) solution of liposomes were made in a container and freeze for approximately 8 hours on the shelf at -40°C. the Pressure was reduced to approximately 0.1 mm, RT. Art. and the vessel was left under reduced pressure for 2 days in a stepwise increase in temperature from -40°C to 25°C over a 2-day period. At the end of this process was obtained approximately 3.5 g of liofilizovannyh unloaded liposomes. Liposomes were then stored at 4°C.

Example 11: the Inclusion oxaliplatin in prior unloaded the liposome

An aqueous solution of l-OHP (8 mg/ml in 300 mm sucrose solution) was added to approximately 3.5 g lyophilisate the Noi non-loaded liposome and re-hydrational by stirring for 2 hours at 40°C. After stirring liposomal l-OHP was separated from free l-OHP fractionation using Sephadex G-25 (Ø1×45 cm). Liposomal l-OHP and free l-OHP was followed up by VIS 600 nm and UV 210 nm, respectively.

Measured the amount of l-OHP and cholesterol. The concentration of l-OHP was calculated for the case of condensation of liposomal fraction relative to the initial concentration of cholesterol, and, finally, the output of the l-OHP was determined by comparing the concentration of l-OHP liposomes and the initial concentration of l-OHP.

The total concentration of l-OHP for the case of condensation of liposomal fraction relative to the initial concentration of cholesterol was 210 ág/ml, And the yield of l-OHP was equal to 2.6%.

This means that 210 µg/ml l-OHP was encapsulated in liofilizovannye unloaded the liposome.

Example 12: Comparison of levels of liposomes in the blood and organs

Comparative study was carried out to assess delays in the blood and accumulation in organs containing l-OHP Tf-modified liposomal compositions in mice-carriers of the tumor. As animal models used male mice of BALB/c at 5 weeks of age, as well as tumor cells used cells Colon 26 (originating from colon cancer mouse). Cells were obtained from the Laboratory of Biopharmaceutics, Teikyo University School of Pharmaceutical Sciences, Japan.

Cells Colon 26 (2×106cells), in addition Kul is iferouane in vitro, implanted subcutaneously in the back of the mouse. As mouse-media tumors of the colon used the mouse with a tumor diameter of approximately 8 to 10 mm (on average after growth for 8 to 10 days). The tail vein is injected with a solution of each of the liposomes obtained in examples 5 and 6, or l-OHP (8 mg/ml in a 9% sucrose solution). In each case the concentration oxaliplatin brought up to 5 mg l-OHP/kg of body weight. As liposomes used the liposome Tf-NG-cells of the dspe ((<); example 5), the liposome Tf/PEG-NG-cells of the dspe ((▲); example 6) and the liposome Tf-PEG-cells of the dspe ((♦); example 6).

Blood, plasma, liver, spleen, kidney, heart, lung and tumor tissue were taken from 3 mice of each group for each time point after 1, 3, 6, 24, 48 and 72 hours after injection. The Pt concentration in the blood, every organ and tumor tissues were determined using atomic absorption (AA), and the concentration of l-OHP was calculated and represented as a ratio (%) to the dose. Concentration in blood is shown in Fig. 9.

Liposome Tf-NG-cells of the dspe showed essentially the same delay in the blood 3 hours after administration, and liposome Tf-PEG-cells of the dspe, and liposome Tf/PEG-NG-cells of the dspe. However, after 6 hours liposome Tf-NG-cells of the dspe showed a slight delay in the blood, but disappeared from the blood faster than PEG liposomes. Concentration in tumor tissues is shown in Fig. 10. Liposome Tf-NG-cells of the dspe showed essentially the same accumulation in tumor tissues, as lipase is and Tf-PEG-cells of the dspe, and liposome Tf/PEG-cells of the dspe, despite the delay in the blood in lower concentrations over time.

On the basis of the above results, it was found that to deliver a sufficient concentration of drug in tumor tissue from mice on a significant or a higher level is necessary and sufficient delay in the blood for approximately 6 hours after administration. Suppose that the delay time in the blood for a longer period of time may increase the likelihood of providing adverse effects on normal tissue.

Example 13: Retrieving diagnostic liposomes and accumulation125I in tumor tissue

Liposomes were obtained by the same method as in example 7, except that l-OHP was replaced with [125I]-termineonline (in solution SFR), and were obtained liposomes of DMPC/CH/NG-DOPE/Tf-NG-DOPE/[125I]-termineonline. Lipid components were obtained as described in example 7. Received two liposomal composition with the components shown below. Liposome containing no Tf-NG-DOPE, served as a control for non-directional distribution of the liposomes.

Liposome directional: DMPC/CH/NG-DOPE/Tf-NG-DOPE (63,3/31,7/4/1 (m/m)

Omnidirectional liposome (control): DMPC/CH/NG-DOPE (63,3/31,7/5 (m/m)

125I was attached to termineonline connection Na125I (PerkinElmer Japa Co., Ltd., Japan) and termineonline (Dr. Kazuo Maruyama, Teikyo University, Faculty of Pharmaceutical Sciences, Japan) using iodophenol method (Biochem. Biophys. Res. Commun., 122, 319-325 (1984), incorporated by reference in its entirety). Thus was obtained125I termineonline. Solution125I termineonline/SFR (') with a concentration of approximately 1 mg/ml was then encapsulated in a liposome as described in example 7.

100 μl each of solutions liposomes were injected with the tail vein of the mouse-media murine colon cancer described in example 12. Tumor tissue and tail had been confiscated from 5 mice of each group for each time point after 1, 6, 24, and 48 hours after injection. Measured the mass of the tumor tissue, and radioactivity (unit: pulse/min) in tumor tissue and the tail was measured using a gamma counter (Aloka Auto Gamma System ARC-300, Japan). The results were expressed as distributed number in the tumor tissue (% of dose/g of tumor) = [(value account in tumor tissue) - (background)] × 100 / [(amount of invoice Std.) - (the value of the account at the tail)] / (mass of tumor tissue (g)). The half-life of radioactivity125I is approximately 60 days.

Radioactivity 100 µl injected solution (standard: Std.) took over 100%, and the value of the account is empty tubes for testing was assumed to be background noise (b.g.). The results are shown figure 11. It is to be seen from 11, modified Tf liposome showed high accumulation in tumor tissue, whereas undirected liposome showed high accumulation. These results indicate that liposome comprising a radioactive compound, a method to identify tumor tissue.

Example 14: Comparison of antitumor effects of liposomes

Comparative study was carried out to evaluate the antitumor effects in mice carrier for colon cancer Colon 26, comprising a l-OHP modified Tf liposomal compositions (liposomes Tf-PEG, obtained in example 9, liposomes Tf-NG-cells of the dspe:NG-cells of the dspe:DSPC:CH, obtained in example 8, liposomes Tf/PEG-NG-cells of the dspe obtained in example 9; 9 mice in each group) and each of the liposomal compositions to which transferrin was not attached ((-)TF; 6 mice in each group).

Mice-carriers of the tumor was obtained in the same manner as in example 12. As control was used a solution of l-OHP (8 mg/ml in a 9% sucrose solution). The date when the l-OHP was administered at a dose of 5 mg/kg was determined as the start date, and on the 4th day of l-OHP was again administered at a dose of 5 mg/kg tumor Size at day 0 was taken as a 1, and a size expressed as the ratio, based on the original size. Tumor size was measured on 0, 2, 5, 7, 10, 13, 15, 18 and 21 days, and recorded the days of survival.

The results are shown in Fig.

As you can see in Fig, composers is AI liposomes, to which was attached transferrin, showed an inhibitory effect on tumor growth. On the other hand, the composition of liposomes to which transferrin was not attached, had a weaker inhibitory effect on tumor growth compared with that of the compositions of liposomes to which transferrin was added. Based on the results presented on figures 9 and 10, it was found that to provide inhibitory effect on tumor growth and ensure sufficient concentration of accumulated drug in tumor tissue to a significant and essentially the same level necessary and sufficient delay in the blood for approximately 6 hours after administration of liposomes is attached to transferrin. Suppose that the delay time in the blood for a longer period of time may increase the likelihood of providing adverse effects on normal tissue.

Example 15: Optimizing content NG-cells of the dspe

To determine the optimal relationship NG-in the cells of the dspe liposome mixing explored the delay in the blood of normal mice NG-cells of the dspe, in which the anticancer agent is not concluded. Liposomal compositions, in which the anticancer agent is not concluded, was obtained in the same manner as in example 8, but using water instead of the solution of l-OHP as the aqueous phase, when choosing the x quantities NG-cells of the dspe.

The total molar quantity of all lipid components constituting the liposome, taken as 100%, and the content of NG-cells of the dspe was expressed as the ratio (mol%) NG-cells of the dspe to the total lipid components. In addition, received a liposome containing 6 mol% of lipid MPB (MPB-cells of the dspe) or lipid PDP cells of the dspe) as a constituent lipid. The liposome MBP receive education liposomes by the addition of maleimidomethyl (MPB) to the amino group of ethanolamine lipid and attach Tf to the liposome through MPB (870013(16:0), Avanti Polar Lipids, Inc., USA). The liposome PDP (870205(16:0, Avanti Polar Lipids, Inc., USA) is obtained by formation of liposomes by attaching 2-pyridylmethylamine (PDP) to the amino group of ethanolamine lipid and attach Tf to the liposome through a PDP.

In the experiment used 105 mice (males line ICR at the age of 6 weeks) (Tokyo Laboratory Animal Science Co., Ltd., Japan). As a label to termineonline (obtained as described in example 13) was added125I, and the solution of inulin with a concentration of approximately 1 mg/ml was made in the liposome. Measured the mass of the collected blood and organs for each case, and radioactivity (unit: pulse/min) liposomal marker was measured using a gamma counter (Aloka Auto Gamma System ARC-300, Japan). In addition, we measured the radioactivity of each injected into the tail vein of a solution (100 μl). Radioactivity 100 ál of injectate (standard: Std) took 10%, and the amount (% of dose) for each organ was expressed as percentage. The total number of blood was taken equal to 7.3% of body weight, and the amount of liposomes in the blood expressed in the form of the number in the whole blood. The value of the account is empty tubes for testing was assumed to be background noise (b.g.), which is subtracted from the value of the account each sample.

Distributed quantity in the blood (%) = [(value of the accounts of the blood) - (value b.g.)] × (body weight of mouse (g)) × 0,073 x 100/[(amount of invoice Std.) - (the value of the accounts of the tail) × (mass of blood (g))].

The results are shown in Fig. With regard to the concentration in the blood after 6 hours, the liposome NG-cells of the dspe showed high latency in the blood, when the content of the lipid is 3 mol% or more. In the case of maleimide-liposomes (MPB 6%) delay in blood was low.

Example 16: Effect of Tf and dicarboxylic acid in a delay in the blood

To study the effect of presence or absence of attached to the liposome transferrin and type of dicarboxylic acid (for example, glutaryl, succinyl etc.) was determined delay in blood liposomes with attached transferrin, which has not entered into an anti-cancer agent. The experimental method was the same as described in example 15.

Was obtained liposome containing a phospholipid to which instead of glutaric acid was added succinic acid.

NG-cells of the dspe (glutaric Ki is the lot) was prepared as follows. In the darkness under a stream of nitrogen gas cells of the dspe (ME-8080, NOF Corporation, Japan) suspended in 10-fold volume of anhydrous chloroform in relation to the volume of the cells of the dspe. Then dropwise at room temperature was added triethylamine (208-02643, Wako Pure Chemical Industries, Ltd., Japan) and a solution of anhydrous glutaric acid (G0071, Tokyo Chemical Industry, Japan) in anhydrous chloroform (dissolved in anhydrous chloroform same size as the cells of the dspe) in amounts equal to 1.3 equivalents. After the solution was allowed to interact at 30°C for 2 hours under stirring.

After that, the reaction solution was washed 3 times with acetate buffer (pH 4.5), and the organic layer dehydrational magnesium sulfate and filtered by filtering with suction using a water-jet pump. Thereafter, the filtrate was concentrated under reduced pressure at 30°C. When it was oily (when the volume is equal to approximately 2 volumes of cells of the dspe)was added to methanol to form crystals and then filtered. This was again dissolved in chloroform, and this procedure was repeated twice. Then the crystals were dried under reduced pressure at room temperature, the result of which was obtained the desired product as white crystals. The liposome NG-cells of the dspe was obtained in the same manner as in example 8.

The results are shown in Fig. Liposome to which transferrin is connected through D. the carboxylic acid (NG-cells of the dspe:N-glutamyltranspeptidase, NG-cells of the dspe:N-succinylsulfathiazole), exhibits high latency in the blood. However, in the case of liposomes to which transferrin connected via S-S connection using maleimide (MPB), a delay in blood was low, even though it was annexed to the same ligand, transferrin.

Example 17: Electrophoretic analysis of liposomes

As an example, analytical methods, characterization of liposomes presents an example of the electrophoresis. The liposome was dissolved and was denaturiruet at 95°C for 5 min in buffer for sample containing 2.5% VAT-Na and 5% 2-mercaptoethanol. Using polyacrylamide gel from about 7.5% to 10% (Funakoshi, Easy gel (II)precast gel, Japan), 5 µl of each sample were applied to the gel, and electrophoresis was performed at a constant current of 20 mA for 1 to 2 hours.

After electrophoresis the gel was stained with silver using the kit silver staining (Wako Pure Chemical Industries, Silver Staining II Kit Wako, Japan). In Fig. 15 shows the results for the following liposomes: track 6 (liposome transferrin-N-glutamyltranspeptidase (liposome Tf-NG-cells of the dspe)); track 5 (liposome transferrin-polyethylene glycol-distearoylphosphatidylcholine (liposome Tf-PEG-cells of the dspe)). Lanes 1-4 contain h-apo-Tf (240 ng), h-apo-Tf (120 ng), h-apo-Tf (60 ng) and h-apo-Tf (30 ng), respectively.

In the case of the example for comparison, liposomes Tf-PEG-cells of the dspe, due to some not is narodnosti molecular weight of polyethylene glycol arose complicated picture electrophoresis with multiple bands. In the case of liposomes Tf-NG-cells of the dspe there was one band, which is much easier to analyze, and which increases ability to clean liposomes. These results show that liposomal compositions of the present invention, the analytical method of testing is simpler than for the derivative of PEG liposomal composition.

Example 18: Effect of free PE liposomal composition

To study the influence of the availability of free phosphatidylethanolamine (non-NG-PE) in the liposome was determined by linking the ability of Tf to liposomes Tf-NG-cells of the dspe and liposomes obtained by adding distearoylphosphatidylcholine (cells of the dspe) (in the absence of NG). The liposome Tf-NG-cells of the dspe received from DSPC (64 parts), CH (32) and NG-cells of the dspe (4 parts), and the liposome Tf-NG-cells of the dspe + cells of the dspe received from DSPC (64 parts), CH (32) and NG-cells of the dspe (4 parts) and cells of the dspe (10 parts) in the same way as in example 8.

Then NG-cells of the dspe was annexed Tf using NHC and ECD in the amount equal to 10 equivalents, and Tf equal to 0.05 equivalent. Then samples of liposomes in an amount corresponding to 1 mg of lipid was separated using a DDS-Na-PAGE, and bands were visualized by the color of silver, as described in example 17.

The results are shown in Fig. It was found that in the case of liposomes NG-cells of the dspe + cells of the dspe, to which was added 10 mol% of the cells of the dspe, the amount of bound Tf was significantly lower than that in the liposome NG-cells of the dspe, which does not contain the Ala non-NG-cells of the dspe. Apparently, this is due to the fact that the amino group of Tf and the amino group of cells of the dspe compete in a reaction where Tf is attached to the carboxyl group NG-cells of the dspe.

Example 19: Comparison of levels of liposomes in the blood and organs

Using the protocols described in example 12, compared the levels of liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH (Tf-NG-DOPE:NG-DOPE) (obtained as in example 7) and liposomes Tf-PEG-cells of the dspe (obtained as in example 9) in the blood and tumors. Results in the number of liposomes detained in the blood, is shown in Fig. 17, and the number of liposomes identified in tumors, shown in Fig. 18.

The results in Fig. 17 and 18 show that, although liposomes Tf-NG-DOPE:NG-DOPE show less accumulation in the blood (Fig. 17)than liposomes Tf-PEG-cells of the dspe, they had the ability to deliver a greater number oxaliplatin in the tumor (Fig. 18). Less accumulation of liposomes in the blood, probably reduces the adverse systemic effects oxaliplatin.

Example 20: a Comparison of antitumor effects of liposomes in mice carrier Colon tumor 26

Using the protocols described in example 14, compared the action of liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH (obtained as in example 7) and liposomes Tf-PEG-cells of the dspe (obtained as in example 9) in tumors of the colon 26 in mice. The results are shown in Fig.

As you can see in Fig, both liposomes show inhibition of tumor growth by sravnenie the solution oxaliplatin, however, as noted in Fig. 19, the accumulation of smaller quantities of NG-DOPE:Tf-NG-DOPE:DMPC:CH in the blood (plasma) should probably mean that these liposomes are better tolerated by individuals to whom they are administered.

Example 21: the Antitumor effect of liposomes on xenograft models HCT-116 tumors of the colon

Determined the antitumor efficacy of injected liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH (obtained as in example 7) in relation to a subcutaneously implanted xenografts HCT-116 tumors of the colon of a person. Testing was conducted at Southern Research Institute, AL, USA male Nude mice NCr-nu (02/A/08F17T9, Frederick Cancer Research and Development Center, MD, of the control connection). Antitumor activity of liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH summarized on Fig.

Liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH was administered intravenous (IV) every fourth day, a total of four injections (q4d×4) at doses of 15 and 10 mg/kg/injection. Oxaliplatin was administered according to the same scheme at a dose of 15 mg/kg/injection. Control groups were injected with the same pattern media (approximately 10.3% of sucrose) and unloaded the liposome.

The average tumor volume for the model tumor colon HCT-116 after treatment with liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH every 4 days accounted for 28.9% of volume control tumor for a group of 15 mg/kg and 35.9 percent of the volume control of the tumor for a group of 10 mg/kg anti-tumor activity of lipo is ω NG-DOPE:Tf-NG-DOPE:DMPC:CH also compared with oxaliplatin outside of liposomes on the model HCT-116, where liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH showed a higher efficiency in units of relative tumor volume with the introduction of 15 mg/kg (28.9% of volume control tumors) every 4 days (4×). Oxaliplatin outside the liposomes, administered at a dose of 15 mg/kg every 4 days (4×) gave 39.3 per cent of the volume control tumors.

Example 22:The antitumor effect of liposomes on xenograft models HT-29 tumors of the colon

Determined the antitumor efficacy of injected liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH (obtained as in example 7) in relation to a subcutaneously implanted xenografts of HT-29 tumors of the colon of a person. Testing was performed in Panapharm Laboratories Co., Ltd., Japan on female Nude mice BALB/cA Jcl-nu (CLEA Japan, Inc., Japan; 50 mice), and the results are summarized in Fig. 21. Groups of 4 mice were injected liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH doses of 6.7, 10, or 15 mg/kg or control media. Groups that received the medium and 6.7 and 10 mg/kg, received injections of 10, 14 and 19 days, and the group treated with 15 mg/kg, received injections of 10 and 14 days.

The average tumor volume for the model tumor colon HT-29 after treatment with liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH amounted to 66.3% from the volume control of the tumor for a group of 6.7 mg/kg and 39.5% of the volume of the control tumors for a group of 10 mg/kg (p-value≤0,01).

Example 23: the Antitumor effect of liposomes on xenograft models MKN45 is the tumor of the stomach

Determined the antitumor efficacy of injected liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH (obtained as in example 7) in relation to a subcutaneously implanted xenografts MKN45 tumor of a human stomach. Testing was performed in Panapharm Laboratories Co., Ltd., Japan male Nude mice BALB/cA Jcl-nu (CLEA Japan, Inc., Japan; 50 mice), and the results are summarized in Fig.

Groups of 4 mice were injected liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH doses of 6.7, 10, or 15 mg/kg or control media. Groups that received the medium and 6.7 and 10 mg/kg, received injections on days 7, 12 and 24 days, and the group treated with 15 mg/kg, received injections on 7 and 24 days. The average tumor volume for the model tumor gastric MKN45 after treatment with liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH accounted for 65.4% of the volume of the control tumors for a group of 6.7 mg/kg (p-value ≤ 0.05), and 49.6 per cent of the volume control of the tumor for a group of 10 mg/kg (p-value≤0.01) and 48.5 per cent of the volume control of the tumor for a group of 15 mg/kg (p-value≤0,01; introduction every 17 days).

Example 24: the Antitumor effect of liposomes on xenograft models COLO 205 tumors of the colon

Determined the antitumor efficacy of injected liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH (obtained as in example 7) in relation to a subcutaneously implanted xenografts COLO 205 tumors of the colon of a person. Testing was conducted at Southern Research Institute, AL, USA on the male bestias the x mice NCr-nu (01/A/09F3T8, Federic Cancer Research and Development Center, MD, USA), and the results are summarized in Fig.

40 mice were injected liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH by/injection every 4 days, just 3 injections (q4d×4) at doses of 10 and 5 mg/kg/injection. Oxaliplatin was administered at a dose of 5 mg/kg/injection according to the same scheme. The control group received injections according to the same scheme. Tumors late-stage re-treated, starting with 47 days for all groups.

Liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH was administered every second day, for a total of two injections at doses of 15 and 10 mg/kg/injection and then the treatment continued with the introduction of every second day, a total of six injections at doses of 4 and 2 mg/kg/injection, respectively. The control group was treated according to the same scheme.

The average tumor volume for the model tumor colon cancer COLO 205 after treatment with liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH entered first in doses of 10 and 5 mg/kg, with subsequent treatment of late-stage tumors, previously subjected to treatment with liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH at doses of 15 and 10 mg/kg, followed by treatment with liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH at doses of 4 and 2 mg/kg when using different schemes treatment ranged from 53.2 per cent to 69.5 per cent of the volume control tumors (p-value≤0.05 or 0.01 to).

Example 25: Encapsulating oxaliplatin in liposomes directed action

To measure the proportion oxaliplatin encapsulated in liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH (Tf-NG-DOPE:NG-DOPE), obtained as in example 7, and is used the following procedure.

The degree of encapsulation was determined by passing an aliquot of the sample through the rotating column 3,000 MWCO (exclusion molecular weight) (3OK MWCO cellulose ultrafilter membrane column, Cat. No. 42410, Millipore Corp., USA) and the concentration measurement oxaliplatin in the eluate using HPLC with isocratic by elution with 1% acetonitrile in dilute aqueous solution of phosphoric acid (pH 3.0).

Level oxaliplatin was determined after membrane filtration using HPLC analysis to quantify the unencapsulated public (free) medicinal substances. Capture efficiency in 3 batches, obtained as in example 7, was more than 98% (see table 1).

Table 1
The percentage encapsulation oxaliplatin in the liposome
PartyIIIIII
% encapsulation98,899,6of 99.1

Example 26: pH liposomes directed action

pH liposomes directed action, you can determine the location of liposomes of the invention in distilled water and measuring the standard pH-meter, as described below.

<> pH liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH (obtained as in example 7) were determined using pH meter (VWR Model 8000) with the electrode, gel-filled Ag/AgCl. PH for 4 batches of liposomes varied from 7,17 to 7.23, as shown in table 2 below.

Table 2
the pH of the liposomes
Party1234
pH7,177,17of 7.237,20

View of liposomes with different pH are summarized in table 3. These results indicate that low pH leads to aggregation, sedimentation and sediment that may be due to protonation of NG-DOPE and Tf with subsequent aggregation bilayer and denaturation of transferrin.

Table 3
The state of liposomes at different pH values
pHObservations
7,19Liquid, translucent, light pink
6,98Liquid, translucent, unchanged compared with the absence of additives, light pink
6,83Liquid, translucent, unchanged compared with the absence of additives, light pink
6,37Liquid, slight white precipitate after the addition, the enlightenment within minutes to the same extent as without additives, light pink
of 5.53Liquid, a white precipitate after the addition, the enlightenment within minutes, but slightly more whitish in color
5,07Liquid, small white sediment, turbid, white in color
4,33High viscosity, a large amount of white precipitate, very turbid, white in color
3,72Very viscous, the sample does not move when you turn microcentrifuge tubes, white in color from top to bottom, opaque, white color

Example 27: Identification of conjugated transferrin and behavior of transferrin in liposomes directed action when DDS-Na-PAGE

This study was carried out to confirm the conjugation of t is anserina with NG-DOPE in liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH, obtained as in example 7. In this way, when transferrin anywhereman with NG-DOPE, the complex exhibits a high molecular weight in comparison with unconjugated transferrin.

The liposome was dissolved and was denaturiruet at 95°C for 5 min in buffer for sample containing 2.5% VAT-Na and 5% 2-mercaptoethanol. The samples are then put on 5-10% gradient polyacrylamide gel and subjected to electrophoresis in the presence of DDS-Na. Migrating protein bands were visualized using G-colloidal brilliant blue (B2025, SIGMA, USA).

Transferrin in the liposome was identified as transferrin conjugated with NG-DOPE, which showed higher molecular mass compared with intact transferrin (see Fig). A minor band with lower molecular weight were detected as free transferrin.

The ratio of free transferrin to transferrin, anywherefrom with NG-DOPE, DDS-Na-PAGE (Fig) was calculated as the peak area using the program Scion Image (available for free access at the address ). The ratio of free Tf to Tf total liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH amounted to approximately 4.7%.

Example 28: Analysis of osmotic pressure

The osmotic pressure at a given temperature depends on sucrose and salts such as sodium chloride and phosphate buffer. It does not depend on the solute, but depends on the amounts of the nuclear biological chemical (NBC density of ions and the size of molecules in solution. Typically, the osmotic pressure can be measured using an instrument known as osmometer, which determines the osmotic pressure in a suitable pressure units.

The osmotic pressure of liposomes NG-DOPE:Tf-NG-DOPE:DMPC:CH, obtained as in example 7, was measured at room temperature using osmometer (Vapro Vapor Pressure Osmometer Model5520, Wescor, Inc., USA). Values osmolarity for 3 preparations of liposomes varied from 360 to 370 mOsm/kg, as shown in table 4.

Table 4
Solernou delanie
PartyABC
Osmolarity (mOsm/kg)360370368

Example 29: Allocation of Tf-NG-DOPE

900 ml of EtOH was added to 100 ml unloaded liposomes (DMPC/Chol/NG-DOPE/Tf-NG-DOPE) (as prepared in example 10 to liofilizirovanny) and fully stirred. The mixture was then centrifuged (9000 rpm, 10 min, 20°C; CFl 6RX, Hitachi Koki Co., Ltd., Japan) again, and was obtained precipitate. 100 ml of EtOH was added to this residue and fully stirred. The mixture is again centrifuged (9000 rpm, 10 min, 20°C; CFl 6RX, Hitachi Koki Co., Ltd., Japan), and was obtained be awaty (light orange) precipitate. This washing process was repeated once more.

The above precipitate was dried gaseous N2within 30 minutes the Dried substance was then dissolved in 10 ml of distilled water and passed through a sterile filter (0.22 μm) (Millipore Corp., USA).

The filtrate was poured in a container and freeze for about 8 hours on the shelf at -40°C. the pressure in the sample was reduced to approximately 0.1 mm, RT. Art. and left under reduced pressure for 2 days in a stepwise increase in temperature from -40°C to 25°C. Thus was obtained approximately 444 mg Tf-NG-DOPE (the content of transferrin was approximately 45% of the unloaded liposomes).

Example 30: Receiving Tf-NG-cells of the dspe

200 µl of aqueous solution of NHS (Wako Pure Chemical Industries, Ltd., Japan) (0.1 mol/l), 200 μl of an aqueous solution of EDC (N-ethyl-N'-(3-dimethylaminopropyl)-carbodiimide hydrochloride) (Tokyo Chemical Industry Co., Ltd., Japan) (0.25 mol/l) and 1 ml NG-cells of the dspe (2 mmol/l)containing 2% (wt./about.) OG (n-octyl-D-glucopyranoside) (Wako Pure Chemical Industries, Ltd., Japan) in 50 mmol/l MES buffer (pH 5.5) was mixed and which within 10 minutes.

Excess reagents were removed using a column Sephadex G-15 (1.5 cm × 20 cm, 0,1% (wt./about) OG 50 mmol/l HEPES buffer (pH 8.0), GE Healthcare Bio-Sciences Corp., USA) and had fractionally at approximately 1 ml/vial.

5 ml of 1% aqueous solution of transferrin (Sigma, USA) was added dropwise to the factions, with whom containing a series NG-cells of the dspe, and gently stirred for 20 hours at 4°C. Identification of each fraction was performed using MS detection.

The reaction product was then fractionally about 1.7 ml/vial on a column of TOYOPEARL HW-55S (1.5 cm × 45 cm, 0.9% NaCl, Tosoh Bioscience LLC, USA). Tf-NG-cells of the dspe were determined using mass spectrometry (MALDI-TOF/MS) and Na-DDS-PAGE with CBB staining (Coomasie Brilliant Blue, Wako Pure Chemical Industries, Ltd., Japan).

1. Liposome intended for the treatment of cancer, comprising one or more phosphatidylcholines, the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine encapsulated drug substance and at least one additional lipid, which is a cholesterol or a derivative of cholesterol,
where a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine guide includes a ligand that is attached to the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine; and
where the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 1

and the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 3

where R 1, R2, R5and R6independently represent an acyl group,
where the acyl group is, independently, come from saturated or unsaturated aliphatic carboxylic acids having 18 to 22 carbon atoms, and
where R1and R2are the same and R5and R6are the same; and
m and p independently represent an integer from 1 to 10;
and where specified guide the ligand is present in an amount of from about 1 μg guide ligand 1 mg lipid to about 50 μg guide ligand 1 mg lipid, and where the liposome does not include non-modified phosphatidylethanolamine, phosphatidylcholine from egg or hydrophilic polymer used to increase the half-life of liposomes in the circulatory direction, and guide where the ligand is not an intact antibody.

2. Liposome according to claim 1, where at least one additional lipid is a cholesterol.

3. Liposome according to claim 1, where one or more phosphatidylcholine is a DMPC, DSPC, RALO or DPPC.

4. Liposome according to any one of claims 1 to 3, where the liposome comprises DMPC and cholesterol, DSPC and cholesterol, RALO and cholesterol or DPPC and cholesterol.

5. Liposome according to any one of claims 1 to 3, where the liposome comprises DMPC and cholesterol.

6. Liposome according to claim 1, where the guide ligand selected from the group consisting of cord is Rina, folic acid, hyaluronic acid, a sugar chain and a fragment of a monoclonal antibody.

7. Liposome according to claim 6, where the guide ligand selected from the group consisting of transferrin, folic acid, hyaluronic acid and a sugar chain.

8. Liposome according to claim 7, where the guide ligand is a transferrin.

9. The liposome of claim 8, where transferrin is holoform, but not in apopharma.

10. Liposome according to any one of claims 1 to 3 or 6 to 9, where the average diameter of the liposomes is from approximately 50 to approximately 250 nm.

11. Liposome according to any one of claims 1, 2, 6-9, where R1, R2, R5and R6are oleoyl or stearoyl and m and p is equal to 3.

12. Liposome according to claim 1, where m and p each independently is an integer from 2 to 4.

13. Liposome according to claim 1, where m and p are equal and are an integer from 2 to 4.

14. Liposome according to item 13, where m and p are equal and amount to 3.

15. Liposome according to claim 1, where R1, R2, R5and R6independently represent oleoyl or stearoyl.

16. Liposome according to claim 1, where R1, R2, R5and R6are the same.

17. Liposome according to claim 1, where at least one additional lipid is a cholesterol, phosphatidylcholine is a DMPC, m and p are equal and amount to 3, R1, R2, R5and R6represent about evil, the medicinal substance is an oxaliplatin and guide the ligand is a transferrin.

18. Liposome according to claim 1, where the medicinal substance is an anticancer agent.

19. Liposome according to claim 1, where the medicinal substance is a compound of platinum.

20. Liposome according to claim 19, where the platinum compound is biltin, cisplatin, carboplatin, ormaplatin, oxaliplatin, triplatin, angloplats, lobaplatin or spiroplatin.

21. Liposome according to claim 20, where the platinum compound is oxaliplatin.

22. Liposome at 17 or 21, where oxaliplatin dissolved in an aqueous solution of sugar selected from the group consisting of trehalose, maltose, sucrose, mannose, lactose, mannitol, glycerin and dextrose.

23. Liposome at 17 or 21, where oxaliplatin dissolved in an aqueous solution of sugar selected from the group consisting of trehalose, maltose, sucrose, mannose, lactose, mannitol, glycerin and dextrose, sugar is in a concentration of from about 1 to about 20% sugar (about./vol.).

24. Liposome at 17 or 21, where the concentration oxaliplatin is from about 0.1 to about 25 mg/ml inside the liposomes.

25. A method of producing liposomes according to claim 1, which includes stages
a) mixing one or more phosphatidylcholines, the first derivative of N-(ω)-dick is oil acid and phosphatidylethanolamine, modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and at least one additional lipid, representing a cholesterol or a derivative of cholesterol, lipid mixtures;
b) adding a drug lipid mixture formed in stage (a);
c) formation of liposomes.

26. The method according to p. 25, further comprising a phase (a) purification of liposomes from the stage (c).

27. The method according to p. 25 or 26, where the medicinal substance at the stage (b) before mixing is in aqueous solution.

28. The method according to p. 25 or 26, where stage (c) comprises sonication, mixing or extrusion.

29. A method of producing liposomes according to claim 1, which includes stages a) mixing one or more phosphatidylcholines, the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, operations of the air of the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and at least one additional lipid, representing a cholesterol or a derivative of cholesterol, lipid mixtures,
where operations the air of the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 2

where R3and R4represent an acyl group,
where ACI is performance communications group come from saturated or unsaturated aliphatic carboxylic acids, having 18 to 22 carbon atoms, and
where R3and R4are the same;
n independently represents an integer from 1 to 10;
b) adding a drug lipid mixture formed in stage (a);
c) formation of liposomes; and
d) attaching a guide ligand operations to the air of the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine with the formation of a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

30. The method according to clause 29, further comprising a stage (e) purification of liposomes from the stage (d).

31. The method according to clause 29 or 30, where the medicinal substance at the stage (b) before mixing is in aqueous solution.

32. The method according to clause 29 or 30, where stage (C) comprises sonication, mixing or extrusion.

33. A method of treating cancer, comprising a) introduction liposomes according to claim 1 in need of this individual in amounts effective for the treatment of cancer, where the liposome directional incorporates a medicinal substance and the medicinal substance is an anticancer agent.

34. The pharmaceutical composition comprising the liposome according to any one of claims 1 to 24, and one or more pharmaceutically acceptable carriers, excipients, diluents, stabilizers or preservatives.

35. Modified Shuttle service is another derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, where a derivative N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 3

where R5and R6represents an acyl group and R5and R6are the same and p is an integer from 1 to 10,
and transferrin attached to the derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

36. Modified transferrin-derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine in p, where R5and R6are oleoyl or stearoyl and p is 3.

37. Unloaded liposome comprising one or more phosphatidylcholines, the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and at least one additional lipid, representing a cholesterol or a derivative of cholesterol,
where a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine guide includes a ligand that is attached to the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine; and
where the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 1

and the second derivative of N-(ω)-di is arbonboy acid and phosphatidylethanolamine represented by formula 3

where R1, R2, R5and R6each independently represents an acyl group,
where the acyl group is, independently, come from saturated or unsaturated aliphatic carboxylic acids having 18 to 22 carbon atoms, and
where R1and R2are the same, R5and R6are the same and
m and p independently represent an integer from 1 to 10; and
and where specified guide the ligand is present in an amount of from about 1 μg guide ligand 1 mg lipid to about 50 μg guide ligand 1 mg lipid, and
where the liposome does not include non-modified phosphatidylethanolamine, phosphatidylcholine from egg or hydrophilic polymer used to increase the half-life of liposomes in the circulatory direction, and guide where the ligand is not an intact antibody.

38. Unloaded liposome according to clause 37, where the one or more phosphatidylcholine is a DMPC, DSPC, RALO or DPPC.

39. Unloaded liposome according to clause 37 or 38, where the mixture comprises DMPC and cholesterol, DSPC and cholesterol, RALO and cholesterol or DPPC and cholesterol.

40. Unloaded liposome according to § 39, where the mixture comprises DMPC and cholesterol.

41. Unloaded liposome according to clause 37, where the guide ligand selected from the group consisting of transferrin, Folie the Oh of the acid, hyaluronic acid, a sugar chain and a fragment of a monoclonal antibody.

42. Unloaded liposome according to paragraph 41, where the guide ligand selected from the group consisting of transferrin, folic acid, hyaluronic acid and a sugar chain.

43. Unloaded liposome according to § 42, where the guide ligand is a transferrin.

44. Unloaded liposome according to item 43, where transferrin is holoform, but not in apopharma.

45. Unloaded liposome according to any one of p, 38 or 41 to 44, where the average diameter of the liposomes is from approximately 50 to approximately 250 nm.

46. Unloaded liposome according to any one of p or 41-44, where R1, R2, R5and R6are oleoyl or stearoyl and m and p is equal to 3.

47. Unloaded liposome according to clause 37, where m and p each independently is an integer from 2 to 4.

48. Unloaded liposome according to clause 37, where m and p are equal and are an integer from 2 to 4.

49. Unloaded liposome according p, where m and p are equal and amount to 3.

50. Unloaded liposome according to clause 37, where R1, R2, R5and R6independently represent oleoyl or stearoyl.

51. Unloaded liposome according to clause 37, where R1, R2, R5and R6are the same.

52. Unloaded liposome according to clause 37, where R1, R2, R5and R6represent the second oleoyl, m and p are equal and are 3, one or more phosphatidylcholines represent DMPC, there is at least one additional lipid, representing cholesterol, and guide the ligand is a transferrin.

53. The method of obtaining the unloaded liposomes according to clause 37, which includes stages
a) mixing one or more phosphatidylcholines, the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine, and at least one additional lipid, representing a cholesterol or a derivative of cholesterol for the formation of a lipid mixture; and
b) the formation of liposomes.

54. The method according to item 53, further comprising a stage (c) purification of liposomes from stage (b).

55. The method according to item 53 or 54, where stage (b) comprises sonication, mixing or extrusion.

56. The method of obtaining the unloaded liposomes according to clause 37, which includes stages
a) mixing one or more phosphatidylcholines, the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and operations of air of the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and at least one additional lipid, representing a cholesterol or a derivative of cholesterol is Rina, for the formation of the lipid mixture,
where operations the air of the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 2

where R3and R4represent an acyl group,
where the acyl group come from saturated or unsaturated aliphatic carboxylic acids having 18 to 22 carbon atoms, and
where R3and R4are the same,
n independently represents an integer from 1 to 10;
b) formation of liposomes and
c) attaching a guide ligand operations to the air of the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine with the formation of a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine.

57. The method according to p, further comprising a stage (d) purification of liposomes from the stage (c).

58. The method according to p or 57, where stage (b) comprises sonication, mixing or extrusion.

59. The way to obtain therapeutic liposomes, including the state
a) encapsulating the drug in an unloaded the liposome according to any one of p-52.

60. The lipid mixture to obtain a liposome according to claim 1 or 37, comprising a mixture of one or more phosphatidylcholines, the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, succinyl the underwater ether of the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and at least one additional lipid, representing a cholesterol or a derivative of cholesterol, where
the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 1

and operations the air of the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 2

where R1, R2, R3and R4independently represent an acyl group,
where the acyl group is, independently, come from saturated or unsaturated aliphatic carboxylic acids having 18 to 22 carbon atoms, and
where R1and R2are the same, R3and R4are the same and
m and n independently represent an integer from 1 to 10; and
where the mixture includes a non-modified phosphatidylethanolamine, phosphatidylcholine from egg or hydrophilic polymer used to increase the half-life of liposomes in the circulatory direction, and guide where the ligand is not an intact antibody.

61. The lipid mixture according to p, where one or more phosphatidylcholine is a DMPC, DSPC, RALO or DPPC, where m and n are equal and amount to 3 and where R1, R2, R3and R4are oleoyl or stearoyl.

62. The lipid mixture to obtain l is bosomy according to claim 1 or 37, comprising a mixture of one or more phosphatidylcholines, the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and at least one additional lipid, representing a cholesterol or a derivative of cholesterol, where
the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 1

and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine guide includes a ligand that is attached to the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine; and
where the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 3

where R1, R2, R5and R6independently represents an acyl group,
where the acyl group is, independently, come from saturated or unsaturated aliphatic carboxylic acids having 18 to 22 carbon atoms, and
where R1and R2are the same, R5and R6are the same;
m and p independently represent an integer from 1 to 10; and
where specified guide the ligand is present in an amount of from about µg guide ligand 1 mg lipid to about 50 μg guide ligand 1 mg lipid and
where the mixture includes a non-modified phosphatidylethanolamine, phosphatidylcholine from egg or hydrophilic polymer used to increase the half-life of liposomes in the circulatory direction, and guide where the ligand is not an intact antibody.

63. The lipid mixture according to item 62, where one or more phosphatidylcholine is a DMPC, DSPC, RALO or DPPC, where m and p are equal and amount to 3, where R1, R2, R5and R6are the same and represent oleoyl or stearoyl and guide where the ligand is a transferrin.

64. A method of obtaining a lipid mixture p, comprising a stage of mixing one or more phosphatidylcholines, the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and operations of air of the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and at least one additional lipid, representing a cholesterol or a derivative of cholesterol.

65. The method of obtaining the lipid mixture according to item 62, comprising a stage of mixing one or more phosphatidylcholines, the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and at least one additional lipid is, representing a cholesterol or a derivative of cholesterol.

66. Containing liposome composition to obtain liposomes according to claim 1, including a non-homogeneous mixture of lipid, aqueous solution and liposomes comprising one or more phosphatidylcholines, the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine, operations of the air of the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and at least one additional lipid, representing a cholesterol or a derivative of cholesterol, where
the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 1

and operations the air of the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 2

where R1, R2, R3and R4independently represent an acyl group,
where the acyl group is, independently, come from saturated or unsaturated aliphatic carboxylic acids having 18 to 22 carbon atoms, and
where R1and R2are the same, R3and R4are the same;
m and n independently represent an integer from 1 to 10; and
where the composition does not include non-modified phosphatidylethanolamine, phosphatidylcholine from egg or hydrophilic polymer used in isoamyl to increase the half-life of liposomes in the circulatory line, and guide where the ligand is not an intact antibody;
and where the composition optionally includes the encapsulated drug.

67. Containing liposome composition on p, where one or more phosphatidylcholine is a DMPC, DSPC, RALO or DPPC, where m and n are equal and amount to 3, where R1, R2, R3and R4are the same and represent oleoyl or stearoyl.

68. Containing liposome composition to obtain liposomes according to claim 1, including a non-homogeneous mixture of lipid, aqueous solution and liposomes comprising one or more phosphatidylcholines, the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and at least one additional lipid, representing a cholesterol or a derivative of cholesterol, where
the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 1

modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine guide includes a ligand that is attached to the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine; and where the second derivative of N-(ω)-dicarboxylic acid and phosphatic is lateralline represented by formula 3

where R1, R2, R5and R6independently represents an acyl group,
where the acyl group is, independently, come from saturated or unsaturated aliphatic carboxylic acids having 18 to 22 carbon atoms, and
where R1and R2are the same, R5and R6are the same and
m and p independently represent an integer from 1 to 10; and
and where specified guide the ligand is present in an amount of from about 1 μg guide ligand 1 mg lipid to about 50 μg guide ligand 1 mg lipid and
where the composition does not include non-modified phosphatidylethanolamine, phosphatidylcholine from egg or hydrophilic polymer used to increase the half-life of liposomes in the circulatory direction, and guide where the ligand is not an intact antibody;
and where the composition optionally includes the encapsulated drug.

69. Containing liposome composition on p, where one or more phosphatidylcholine is a DMPC, DSPC, RALO or DPPC,
where m and p are equal and amount to 3, where R1, R2, R5and R6are the same and represent oleoyl or stearoyl and guide where the ligand is a transferrin.

70. Contains whether the Osama composition on p, comprising the medicinal substance.

71. Containing liposome composition on p comprising the medicinal substance.

72. The method of obtaining containing liposome composition according p or 68, which includes stages
a) mixing one or more phosphatidylcholines and the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and, when present, operations of the air of the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine or modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and at least one additional lipid, representing a cholesterol or a derivative of cholesterol, a lipid mixture; and
b) adding a drug lipid mixture formed in stage (a); and
c) education containing liposomes of the composition.

73. The method of obtaining containing liposome composition according p, which includes stages
a) mixing one or more phosphatidylcholines, the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine and at least one additional lipid, representing a cholesterol or a derivative of cholesterol, a lipid mixture; and
b) after the Department of solvent to the mixture, formed in stage (a), with the formation containing liposomes of the composition.

74. Liposome according to claim 1, which contains from about 10 μg guide ligand/mg lipid to about 50 μg guide ligand/mg lipid.

75. Liposome according to claim 1, where the drug is oxaliplatin, R1, R2, R5and R6are oleoyl, m and p is equal to 3, and the guide ligand is a transferrin, one or more phosphatidylcholine is a DMPC and liposome directional contains from about 10 μg guide ligand/mg lipid to about 50 μg guide ligand/mg lipid.

76. Liposome according to claim 1, which contains from about 10 μg guide ligand/mg lipid to about 25 μg guide ligand/mg lipid.

77. Liposome according to claim 1, where the drug is oxaliplatin, R1, R2, R5and R6are oleoyl, m and p is equal to 3, and the guide ligand is a transferrin, one or more phosphatidylcholine is a DMPC and liposome directional contains from about 10 μg guide ligand/mg lipid to about 25 μg guide ligand/mg lipid.

78. Liposome according to claim 1, where the total content of the first derivative of N-(ω)-d the carboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is from about 2 to about 6 mol % of the total lipid content.

79. Liposome according to claim 1, where the drug is oxaliplatin, R1, R2, R5and R6are oleoyl, m and p is equal to 3, the guide ligand is a transferrin, one or more phosphatidylcholine is a DMPC, and the total content of the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is from about 2 to about 6 mol % of total lipid content.

80. Liposome according p, where the total content of the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is from about 2 to about 6 mol % of total lipid content.

81. Liposome according p, where the total content of the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is from about 2 to about 6 mol % of total lipid content.

82. Liposome according p, where the total content of the first derivative of N-(ω)-dicarboxylic acid and phosphatidylserine the ins and modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine is from about 2 to about 6 mol % of total lipid content.

83. Liposome intended for the treatment of cancer, comprising one or more phosphatidylcholines, the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine encapsulated drug substance and at least one additional lipid, which is a cholesterol or a derivative of cholesterol,
where a modified guiding factor derived N-(ω)-dicarboxylic acid and phosphatidylethanolamine guide includes a ligand that is attached to the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine; and
where the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 1

and the second derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine represented by formula 3

where R1, R2, R5and R6independently represent an acyl group,
where the acyl group is, independently, come from saturated or unsaturated aliphatic carboxylic acids having 12-22 carbon atom,
where liposomes directional contain from about 10 μg guide ligand 1 mg lipid to about 50 μg guide League of the Yes on 1 mg of lipid and
m and p independently represent an integer from 1 to 10; and
where the liposome does not include non-modified phosphatidylethanolamine, phosphatidylcholine from egg or hydrophilic polymer used to increase the half-life of liposomes in the circulatory direction, and guide where the ligand is not an intact antibody.

84. Liposome according p, where at least one additional lipid is a cholesterol.

85. Liposome according p or 84, where one or more phosphatidylcholine is a DMPC, DSPC, RALO or DPPC.

86. Liposome according p, where the liposome comprises DMPC and cholesterol.

87. Liposome according to any one of p, 84 and 86, where the guide ligand selected from the group consisting of transferrin, folic acid, hyaluronic acid, a sugar chain and a fragment of a monoclonal antibody.

88. Liposome according p, where the guide ligand selected from the group consisting of transferrin, folic acid, hyaluronic acid and a sugar chain.

89. Liposome according p, where the guide ligand is a transferrin.

90. Liposome according p, where transferrin is holoform, but not in apopharma.

91. Liposome according to any one of p, 84, 86, 88-90, where the average diameter of the liposomes is from approximately 50 to approximately 250 nm.

92. Liposome according to any one of p, 84, 86, 88-90, where R1, R2, R5and Rsup> 6are oleoyl or stearoyl and m and p is equal to 3.

93. Liposome according p, where R1, R2, R5and R6are oleoyl.

94. Liposome according to any one of p, 84, 86, 88-90, or 93 where the medicinal substance is an anticancer agent.

95. Liposome according p, where the medicinal substance is a compound of platinum.

96. Liposome according p, where the platinum compound is biltin, cisplatin, carboplatin, ormaplatin, oxaliplatin, triplatin, angloplats, lobaplatin or spiroplatin.

97. Liposome according p, where the platinum compound is oxaliplatin.

98. Liposome according p, where the drug is oxaliplatin, R1, R2, R5and R6are oleoyl, each of m and p is equal to 3, the guide ligand is a transferrin, one or more phosphatidylcholines represent DMPC at least one additional lipid is a cholesterol.

99. Liposome according p, where transferrin is holoform.

100. Liposome according to any one of p-99, where oxaliplatin dissolved in an aqueous solution of sugar selected from the group consisting of trehalose, maltose, sucrose, mannose, lactose, mannitol, glycerin and dextrose.

101. Liposome according to item 100, where the sugar is present at a concentration of priblizitelen is 300 mm.

102. Liposome according to any one of p-99 and 101, where the concentration oxaliplatin is from about 0.1 mg/ml to about 25 mg/ml inside the liposomes.

103. A method of treating cancer, including
a) introduction liposomes on p needy in this individual in amounts effective for the treatment of cancer, where the liposome comprises a medicinal substance and the medicinal substance is an anticancer agent.

104. The pharmaceutical composition comprising the liposome according p and one or more pharmaceutically acceptable carriers, excipients, diluents, stabilizers or preservatives.

105. Liposome according p, where the guide ligand is a transferrin at least one additional lipid is a cholesterol where the specified liposome directional contains transferrin in an amount of from about 10 μg transferrin, 1 mg lipid to about 50 μg transferrin, 1 mg of lipid, each R1, R2, R5and R6represents an acyl group of a saturated or unsaturated aliphatic carboxylic acids having 18 to 22 carbon atoms, R1and R2are the same, R5and R6are the same.

106. Liposome according p, where R1, R2, R5and R6are oleoyl, stearoyl, Palmitoyl or meristo and m and p is equal to 3.

107. Liposome according p, where the medicinal substance is an anticancer agent.

108. Liposome according p, where one or more phosphatidylcholines represent DMPC or DSPC.

109. Liposome according p, where the guide ligand is a transferrin and at least one additional lipid is a cholesterol where the specified liposome directional contains transferrin in an amount of from about 10 μg transferrin, 1 mg lipid to about 50 μg transferrin, 1 mg of lipid, R1and R2are the same, R5and R6are the same and R1, R2, R5and R6are not Palmitoyl or myristoyl.

110. Liposome according p, where the guide ligand is a transferrin and at least one additional lipid is a cholesterol where the specified liposome directional contains transferrin in an amount of from about 10 μg transferrin, 1 mg lipid to about 50 μg transferrin, 1 mg of lipid content phosphatidylcholine is 40-60 mol %; the content of cholesterol is 35-55 mol %; the total content of the first derivative of N-(ω)-dicarboxylic acid and phosphatidylethanolamine and modified guiding factor derived N-(ω)-dicarboxylic acid and the phosphate is of diethanolamine is 2-8 mol % and the total content of phosphatidylcholine, cholesterol and derivatives of phosphatidylethanolamine is 100 mole %; each R1, R2, R5and R6represents an acyl group of a saturated or unsaturated aliphatic carboxylic acids having 18 to 22 carbon atoms, R1and R2are the same, R5and R6are the same.

111. Liposome according p, where R1, R2, R5and R6are oleoyl or stearoyl and m and p is equal to 3.

112. Liposome according to § 111, where the medicinal substance is an anticancer agent.

113. Liposome according to § 111, where one or more phosphatidylcholines represent DMPC or DSPC.



 

Same patents:

FIELD: chemistry.

SUBSTANCE: invention relates to peptide-based compounds containing three-member rings containing a heteroatom, which efficiently and selectively inhibit specific activity of N-terminal nucleophilic (Ntn) hydrolase, bonded with a proteasome. The peptide-based compounds contain epoxide and are functionalised at the N-end.

EFFECT: peptide-based compounds exhibit anti-inflammatory properties and cell proliferation inhibition, oral administration of said peptide-based proteasome inhibitors is possible owing to bioavailability thereof.

23 cl, 14 ex

FIELD: chemistry.

SUBSTANCE: invention relates to novel derivatives of the anti-tumour antibiotic of the aureolic acid olivomycin A group, having anti-tumour activity and low toxicity, and synthesis method thereof. The invention also relates to a method of producing derivatives of the antibiotic of the aureolic acid olivomycin A group, involving selective oxidation of the side chain of olivomycin A aglycone through reaction with sodium periodate, followed by amidation of the obtained key intermediate 1'-des-(2,3-dihydroxy-n-butyroyl)-1'-carboxyolivomycin A with corresponding amines in the presence of a condensing agent.

EFFECT: disclosed compounds have marked anti-tumour activity and low toxicity compared with the original olivomycin A.

2 cl, 9 ex, 1 dwg, 6 tbl

FIELD: chemistry.

SUBSTANCE: invention relates to compounds of formula (I), having histone deacetylase (HDAC) enzyme inhibiting action, stereoisomers, hydrates, solvates and pharmaceutically acceptable salts thereof, compounds of formula (II), compounds selected from a list, a method of producing compounds of formula (I), a pharmaceutical composition, an inhibition method and methods of treating using compounds of formula (I). In formulae

and

R denotes substituted or unsubstituted groups selected from (C6-C10)aryl, (C3-C12)cycloalkyl, heteroaryl, (C6-C10)aryl (C1-C6)alkyl and heterocyclyl; where the heterocyclyl hereinafter is a 5-10-member ring radical which consists of carbon atoms and 1-5 heteroatoms selected from nitrogen, oxygen and sulphur, and heteroaryl hereinafter is an aromatic heterocyclyl, and each aryl, cycloalkyl, heteroaryl, arylalkyl and heterocyclyl can be substituted with one or more substitutes selected from halogens, including fluorine, chlorine, bromine, iodine, (C1-C6)alkyl, (C1-C6)alkoxy, (C6-C10)aryl, halogen(C1-C6)alkyl, (C6-C10)aryl(C1-C6)alkoxy, -O-(C3-C12)cycloalkyl, -O-CH2-(C3-C12)cycloalkyl, hydroxyl, NRaRb and ORa, where Ra and Rb independently denote (C1-C6)alkyl and aryl; R1 denotes (C6-C10)aryl; R2 and R3 independently denote hydrogen, (C1-C6)alkyl, -COOR5, -CONR5R6, -CH2NR5R6, -CH2CH2NR5R6, -CH2CH2OH or -CH2OH; provided that one of R5 or R6 is hydrogen or unsubstituted alkyl, the other is neither hydrogen nor unsubstituted alkyl; R5 and R6 independently denote hydrogen, (C1-C6)alkyl, (C3-C12)cycloalkyl, (C6-C10)aryl, (C6-C10)aryl(C1-C6)alkyl, heteroaryl or heteroaryl(C1-C6)alkyl, which can be unsubstituted or substituted; or R5 and R6 can be combined to form a saturated or unsaturated 3-8-member ring having 0-2 heteroatoms, including N, O or S; where the heteroaryl hereinafter is a 5-10-member ring radical consisting of carbon atoms and 1-5 heteroatoms selected from nitrogen, oxygen and sulphur, and each alkyl, cycloalkyl, aryl, arylalkyl, heteroaryl or heteroarylalkyl can be substituted with one or more substitutes selected from halogen, including chlorine, fluorine, bromine or iodine,(C1-C6)alkoxy and NRaRb; R4 denotes OH, (C6-C10)aryl, ortho-substituted aniline or amino (C6-C10) aryl, which can be optionally substituted with one or more groups selected from halogens, including fluorine, chlorine, bromine, iodine, hydroxyl, amino groups or (C6-C10)aryl; X denotes -NR7-, -CONR7- or -N R7CO; R7 denotes hydrogen or (C1-C6)alkyl; Y denotes (C6-C10)aryl or (C6-C10)aryl(C2-C6)alkenyl; m is an integer from 0 to 1; n is an integer from 0 to 1; o is an integer from 0 to 7; and p is an integer from 0 to 1.

EFFECT: improved method.

FIELD: medicine.

SUBSTANCE: invention refers to medicine, namely oncology, and is applicable for treating rectal cancer with synchronous remote metastases. That is ensured by radiation therapy together with radio modifiers and combined with chemotherapy followed by radical resection of a primary tumour and remote metastases. The radio modifiers are presented by the intrarectal introduction of metronidazole as a part of a composite mixture containing sodium alginate and 2% dimethylsulphoxide, and also local microwave hyperthermia. The chemotherapy is presented by a two-day intensive course of chemotherapy FOLFOX6 involving high doses of platinum and 5-fluorouracil in the form of the 48-hour infusion.

EFFECT: method allows reducing total length of treatment, enabling a complete course of effective preoperative radiation chemotherapy with minimum toxicity in a relatively short time, performing earlier sphincter-preserving operations with well-timed resection of the primary tumour, regional and remote metastases ensured by implementing mechanisms, chemosensitisation and synergic effect of said radio sensitising agents.

2 ex

FIELD: medicine.

SUBSTANCE: invention refers to cell biology. Dopamine and/or its synthetic analogues, particularly substituted 3,4-dihydro-2(1H)-pyrimidinthione is applied as a cytotoxic agent having an effect on human cancer cells in culture.

EFFECT: presented substances can find application in medicine as a base for developing dosage forms used for therapy of malignant growths.

5 dwg, 3 ex

FIELD: chemistry.

SUBSTANCE: invention relates to a compound having chemical structure of formula II , all salts and stereoisomers thereof, where the value of radicals D, A2 and B are as described in paragraph 1 of the claim. The invention also relates to a composition having activity as a c-kit and c-fms modulator, a method of treating a subject suffering from a disease or condition mediated by c-kit and c-fms and a kit for modulating c-kit and c-fms.

EFFECT: novel compounds which can be useful in treating c-kit-mediated diseases or conditions and/or c-fms-mediated diseases or conditions are obtained and described.

21 cl, 44 ex

FIELD: chemistry.

SUBSTANCE: invention relates to substituted heteroarylpiperidine derivatives of formula (I) and enantiomers, diastereomers, tautomers, solvates and pharmaceutically acceptable salts thereof, where R1 denotes -N(R10)-(C(R6)2)m-T, (C(R6)2)1-T or -O-(C(R6)2)m-T; R6 is independently selected from H, OCH3, C1-6-alkyl, possibly substituted with 1-3 substitutes which are halogen, and C3-6-cycloalkyl, possibly substituted with 1-3 substitutes which are halogen, T denotes NR7R8, , , , or ; R7 and R8 are independently selected from H, C1-6-alkyl; R9 is independently selected from OH, C1-6-alkyl, O-C1-6-alkyl, or NR12R13; R10 denotes H or C1-6-alkyl; R12 and R13 are independently selected from C1-6-alkyl, possibly substituted with OH, C2-6-alkylene-O-C1-6-alkyl and W denotes CH, O or NR10; B denotes CR2 or N; G denotes CR2 or N; D denotes CR2 or N; E denotes CR2 or N; provided that one or more of variables B, G, D and E must be N; R2 is independently selected from H, F, Cl, CH3, OCH3 and CF3; R3 denotes: H, CI, F or CH3; R4 denotes Cl, F or CH3, R5 denotes , morpholine, possibly substituted with 1-3 identical or different substitutes R14, a 4-7-member saturated or partially unsaturated heterocycle containing one nitrogen atom in the ring and possibly an additional heteroatom selected from O, N and S, where the heterocycle is possibly substituted with 1-4 identical or different substitutes R11, or NR12R13; R11 is indendently selected from halogen, OH, C1-6-alkyl, possibly substituted with 1-3 substitutes which are halogen, C2-6-alkynyl, -C0-6-alkyl-C3-6-cycloalkyl, -OC(O)C1-6-alkyl, -NH2, -NH(C1-6-alkyl) and -N(C1-6-alkyl)2; A denotes a 3-7-member saturated ring; R12 and R13 are independently selected from C1-6-alkyl, possibly substituted with OH, C2-6-alkylene-O-C1-6-alkyl; R14 denotes C1-6-alkyl; 1 equals 0, 1, 2, 3 or 4; m equals 0, 1, 2, 3 or 4; o equals 0, 1 or 2; p equals 0, 1, 2, 3 or 4; r equals 0, 1, 2, 3 or 4; s equals 1 or 2 and t equals 0 or 1. The invention also relates to use the compound of formula I to produce a drug for treating or preventing disorders, diseases or conditions responsible for inactivation or activation of the melanocortin-4 receptor in mammals, and to a pharmaceutical composition based on said compounds.

EFFECT: novel compounds which can be used as melanocortin-4 receptor modulators are obtained and described.

10 cl, 134 ex, 16 tbl

FIELD: medicine, pharmaceutics.

SUBSTANCE: present invention refers to pharmacology and medicine and concerns a combination containing a compound of formula (1) and one or more pharmaceutically active agents for treating cancer, a pharmaceutical composition containing said combination, a method of treating, and a market pack containing said combination.

EFFECT: invention provides high clinical effectiveness.

9 cl, 6 dwg, 2 tbl, 1 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: present invention refers to a pharmaceutical composition and to a drug of 1,3-diethylbenzimizadolium salts of general formula: wherein X is an anion.

EFFECT: there are produced the compositions exhibiting high anti-cancer activity.

2 cl, 3 tbl, 5 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: what is presented is application of an effective amount of the composition containing nanoparticles containing taxane and a carrier protein for preparing a drug for a proliferative disease in a combination with an effective amount of at least one other chemotherapeutic agent specified in a group consisting of antimetabolite, platinum agents, alkylating agents, tyrosine kinase inhibitors, anthracycline antibiotics, vinca alkaloids, proteasome inhibitors, macrolides, a therapeutic antibody, an antiangiogenic agent, geldanamycin, 17-AAG and topoisomerase inhibitors; a related composition and a kit.

EFFECT: shown higher clinical effectiveness in combined treatment of even hard-to-treat cancers, including cancer of lungs and pancreatic cancer with no new toxicity cases detected.

46 cl, 9 dwg, 16 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to pharmacology. A liposome suspension composition for prevention and treatment of respiratory infections, particularly tuberculosis, contains liposomes, propylene glycol pine wood extract, propylene glycol sage extract, marigold, bee balm and eucalyptus essences, Carbopol, glycerin, blanose, epo-phen, kathon, sodium hydroxide and water. In other version, the liposome suspension composition for prevention and treatment of respiratory infections, particularly tuberculosis, contains liposomes as a basis, propylene glycol pine wood extract, propylene glycol dandelion, burdock and cornflower extracts, lavender, bergamot and schizandra essences, Carbopol, glycerin, blanose, epo-phen, kathon, sodium hydroxide and water. A method for prevention and treatment of respiratory infections, particularly tuberculosis by inhalations; it involves aerosol processing of a room by the presented liposome suspension composition by making 10 applications of a dosing cock with the area of 20 sq. m.

EFFECT: using the offered invention enables widening the spectrum of ecologically safe high-effective immune-enhancing agents with antimicrobial, anti-viral and fungicidal effect for massive prevention and treatment of respiratory infections, including tuberculosis.

3 cl, 2 ex, 2 tbl, 2 dwg

FIELD: medicine, pharmaceutics.

SUBSTANCE: micellar composition of an amphiphilic block copolymer contains taxane, an amphiphilic block copolymer containing a hydrophilic block (A) and a hydrophobic block (B), and an agent for osmolality regulation. The amphiphilic block copolymer is a block copolymer of A-B, A-B-A or B-A-B type. The hydrophilic block (A) has number-average molecular weight 500-20000 dalton, and the hydrophobic block (B) has number-average molecular weight 500-20000 dalton. The amphiphilic block copolymer contains 40-70 wt % of the hydrophilic block (A) of weight of the copolymer. Hydroxyl terminal groups of the hydrophobic block (B) can be protected by fatty-acid groups. The agent for osmolality regulation represents an inorganic salt. What is also described is a method for preparing the given composition. The composition exhibits good stability that allows preventing fast release of a medical preparation of taxane and can improve required pharmacological action.

EFFECT: method ensures high effective preparation of the composition.

14 cl, 2 dwg, 3 tbl, 4 ex

Colloidal solution // 2449776

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention relates to field of dermatology and represents colloidal solution suitable for dermal and/or local application on skin, as base for compositions of dispersed foam, containing, at least, one active ingredient, at least, one membrane-forming molecule, at least, one foam-generating component and, at least, one solvent, with active ingredient representing dexpanthenol, membrane-forming molecule represents phosphatidylcholine and foam-generating component being selected from opiate, cocoamphoceate, capryl/capramidopropyl betaine, mixture of PEG-5 lauryl citrate/sulfosuccinate/laureatesulfate, mixture of lauryl glucose carboxylate/lauryl glucoside or their mixture.

EFFECT: invention ensures high stability of particles in colloidal solution at 50°C for 2 months without changing their size, as well as ensures formation of stable foam when solution is applied as base for dispersed foam.

16 cl, 3 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: group of inventions relates to medicine. Substance of the inventions involves monolamellar liposomes of the size of 100-200 nm consisting of a mixture containing 1 weight fraction of tetramannosyl-tri-L-lysine-diopeoyl glycerol and 99 weight fraction of 2,3-dipalmitole-Sn-glycero-phosphatidylcholine, and containing three oligopeptides with their sequence relevant to the sequences of the fragments: 46-62, 124-139 and 147-170 myelin basic protein. What is offered as an agent for multiple sclerosis is a pharmaceutical composition containing monolamellar liposomes as an active ingredient, as well as a method of treating multiple sclerosis involving the introduction of said pharmaceutical composition into the patient.

EFFECT: preparing the liposomes containing oligopeptides - fragments of myelin basic protein, the pharmaceutical composition and developing the method of treating multiple sclerosis.

5 cl, 7 ex, 1 tbl

FIELD: medicine.

SUBSTANCE: invention refers to a liposomal preparative form for dermatological and external pharmacological application. A liposomal nanocapsule represents a hollow sphere formed by a bilayer lipid membrane containing inner and outer hydrophilic layers comprising an aqueous extract of peloids with a hydrophobic area of the bilayer lipid membrane in between, comprising a lipid extract of peloids and polar charged molecules being arranged on the surfaces of the inner and outer hydrophilic layers. The liposomal nanocapsule comprises: an aqueous extract of peloids - 64%, an lipid extract of peloids - 25%, an aqueous complex of humic acids - 10%, a phospholipid-based stabiliser - 1%.

EFFECT: invention provides enhanced therapeutic and cosmetic effect of the liposomal nanocapsule.

1 dwg, 1 tbl, 2 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to medicine and chemical-pharmaceutical industry. A composition contains a corticosteroid and an antihistamine, phospholipid liposomes, a pharmaceutically acceptable aqueous carrier and a pharmaceutically acceptable buffer likely to control pH approximately 4 to 8. Herewith the antihistamine is cetirizine and its salts, and the corticosteroid is budesonide, ciclesonide, fluticasone, triamcinolone, mometasone and their salts.

EFFECT: preparing the homogenous pharmaceutical composition for treating, eg rhinitis, asthma and/or chronic obstructive pulmonary disease.

58 cl, 6 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to pharmacology and represents a dried pharmaceutical composition containing a lyophilised active agent containing vesicles comprising: a) at least one lipid, b) at least one active agent which appears to be protein or its active fragment, c) a fusion promoting agent, where the fusion promoting agent is an alkaline amino acid selected from arginine, histidine, lysine or citrulline, and d) being free from a membrane stabilising agent, where rehydration of the dried pharmaceutical composition with using an aqueous solution causes formation of multilamellar liposomes having an average liposome diameter exceeding 1 mcm with these liposomes encapsulating the active agent.

EFFECT: invention provides high efficacy and stability of the active agent in the liposomes having the diameter exceeding 1 mcm.

28 cl, 9 ex, 2 tbl, 3 dwg

FIELD: medicine.

SUBSTANCE: group of inventions is referred to the area of medicine, namely to the area of virusology, and is related to virosomes containing hemagglutinin extracted from influenza virus produced in the cell lines, compositions, containing said virosomes, means of manufacturing and applications. The essence of the invention including virosomes containing hemagglutinin extracted from influenza virus produced in the bird cell lines, compositions, containing said virosomes, application of virosome as a vessel, set, methods of vaccination, methods of treatment and methods of virosome production.

EFFECT: production of virosomes having improved merging capability and increased immunogenicity.

21 cl, 3 tbl, 5 dwg

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to medicine, particularly immunology, namely immunocorrection drugs, and can be used as an inducer of a granulocyte-macrophage colony-forming factor in cells of a mononuclear phagocyte system in vitro and for efferent therapy in pathological conditions accompanied by decrease in cell-mediated immunity. The drug represents oxidised dextrane of average molecular weight 35 - 65 kDa. The drug can be presented in the form of a solution or a nanoliposomal emulsion of the concentration of oxidised dextrane 1-5 wt %. The drug is applied by introduction in a cell culture of the mononuclear phagocyte system in an amount containing oxidised dextrane 125-250 mcg per culture medium 1 ml.

EFFECT: drug under the invention exhibits high biocompatibility.

8 cl, 1 tbl, 4 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to medicine, specifically pharmaceutics. What is created is a liposome preparative form of aminoglycoside with a lipid double layer and encapsulated aminoglycoside, in which a bulk amount of lipid makes less than or equal to an amount of aminoglycoside with the lipid double layer containing neutral phospholipid and sterol, and liposome has an average diameter 0.1 to 0.5 micron. Also, the present invention refers to a method of treating a patient with a pulmonary infection which involves administration of a therapeutically effective amount of the lipid anti-infectious preparative form in the patient, and to a method of treating a patient suffering mucoviscidosis which is based on administration of a therapeutically effective amount of the lipid anti-infectious preparative form according to the present invention in the patient.

EFFECT: use of the declared liposome anti-infectious agent provides a prolonged systemic therapeutic effect by prolonged release.

79 cl, 6 ex, 13 tbl, 9 dwg

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to the use of a composition for inhibiting lipogenesis and a method for inhibiting lipogenesis. The composition contains lactoferrin and a trivalent chromium compound with lactoferrin being connected with trivalent chromium ions in the complex of lactoferrin-trivalent chromium. The trivalent chromium compound is specified in a group consisting of chromium chloride hexahydrate (III), chromium chloride (III), chromium acetate (III), chromium sulphate (III), chromium picolinate, chromium nicotinate, GTF chromium, complex of chromium and yeast extract and their combination.

EFFECT: invention provides glucose transfer from cells to muscle tissue thereby decreasing stored fat transformed of glucose that leads to achieve a goal of body weight control.

14 cl, 5 tbl, 5 ex

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