Antitumoral agent on basis of immunopolisome biological structure, way of its obtaining and vectorial delivery in central nervous system at tumoral process

FIELD: medicine.

SUBSTANCE: invention concerns biopharmacology and medicine area. The antitumor agent representing a immunoliposome biological structure, including a liposome containing the therapeutic agent, sewed with a vector of peptide nature, thus for treatment of CNS tumors is declared, the liposome contains the therapeutic agent in a water phase, as a vector contains monoclonal antibodies to CD34+, and a linking represents 2-iminotiolan (IT) in 0.1% concentration. As a therapeutic agent, immunoliposome contains the substance chosen from the group: Daunomycin, Carminomycinum, Melphtalan, Methotrexatum, Cytarabinum, Doxorubicinum, Ricine. The method of obtaining of an antitumoral agent and way of inhibition of a tumor of the brain, consisting in agent administering due to item 1 in a pharmaceutically suitable carrier in effective quantity is declared also. Thus preliminary administer parenterally a preparation of hematological stem cells CD34+.

EFFECT: provision of highly effective delivery of an antitumor agent to a CNS tumor.

4 cl, 7 ex, 4 dwg

The invention relates to the field of biopharmacology. In particular, the development of new medicines on the basis of immunoliposomes design, method of its production and vector delivery of such funds in brain tissue during tumor pathology of the Central nervous system (CNS).

Brain tumors are the most malignant diseases. Low treatment efficiency due to asymptomatic early in the disease resistance of these tumors to conventional chemo - and radiation therapies, a high potential for investirovanie healthy tissue, and often the impossibility of an adequate surgical intervention because of the way the localization of the tumor.

Therefore, for the treatment of this pathology it is necessary to develop a fundamentally new tools and approaches for the delivery of therapeutic agent to the tumor cells, localized in the Central nervous system.

One of the promising new approaches in the design of delivery systems for various purposes is the use of lipid vesicles of the nano-range, called liposomes, as a convenient means of transportation, at the same time significantly reducing the toxicity of the drug in comparison with the effect of its free form, in addition, they can serve as a transport system for delivering cytotoxics what their agents, more specific to tumors or other lesions pathology. Medicinal substance; biologically active substance or genetically engineered material can be enclosed in the internal cavity of lipid microcapsules or directly in forming the liposome lipid bilayer (1).

On the other hand, there is a problem not just transportation of the drug to tumor lesion, but also targeted delivery into specific organ affected by the cancer process, in our case it is the tissue of the Central nervous system.

We conducted a patent search confirmed that there are a number of patent publications, confirming the use of liposomes and their various improved forms as medicines for the treatment of neoplastic diseases.

In particular, in 1985, published Japanese patent where the disclosed anti-cancer or carcinostatic agent enclosed in the liposomal membrane, and with the surface of the liposomes is connected antibody to embryonic protein alpha fetoprotein having some affinity with tumor cells (patent JP 60067434, 17.04.1985). The drug is intended to treat hepatocarcinoma.

However, the tool may not be effective in the treatment of tumors of the Central nervous system and, in addition, it is intended to use embryonic material. However, it should be noted, therapy drugs made sambreville fabric, has very serious problems of tissue compatibility, the occurrence of delayed immune conflicts, and moral-ethical, legal and religious restrictions.

Quite promising as a vector of carriers of therapeutic agents to tumors can be considered neutral stem cells (hereinafter NSC), which has a native capacity of the directed migration to the hearth and remote metastasis of tumor tissue (2). therapeutic agent may be a drug of genetically modified NSC expressing biologically active factor, for example, tumor necrosis factor (TNF) (3) or sitoindosides, the enzyme contained in the human body and locally catalyzing non-toxic substance - the prodrug 5-fertilizin or cytostatic agent 5-fluorouracil (4).

However, these tools still for brain tumors barely acceptable due to the specific structure of the glial cells and the presence of the blood-brain barrier (BBB).

Recently made attempts to create anticancer drugs, containing liposome with inclusion inside its shell anticancer and cancerostatic funds, and liposomal particle further comprises a vector component peptide covalently attached to the lipid membrane of the liposomes, and the vector component SEL is an from a group of different families of growth factors (RF patent 2292898, 10.02.2007,). It should be noted that the presented tool is also not suitable for the treatment of brain tumors, because trapnest to tumor cells of the brain does not possess.

Conducting a thorough inspection prior art to assess our work on the criterion of "inventive step", we have investigated the possible use of proteinsathome agents that could be used as a binder agent (crosslinking) between the antibody and cytotoxic agent, or the membrane of the liposomes, our work consisted in the creative choice of such a blending of many well-known (see, e.g., Pat. RF 2270029, 2006): bifunctional derivatives of imidapril, active esters, for example disuccinimidyl, aminothiols, glutaric aldehyde, usaideoire and many others.

The closest analogue for the problem at hand can be considered as the operation described in U.S. Pat. RF 2229287, 2004, There is described a cell line that can Express the molecule, growth inhibitory Central nervous system tumors are encapsulated in immunohistology alginate containing more than 15% guluronic acid, the molecule may have an impact on neurovasculature tumors, can interact with different receptors and growth factors in cancer cells of the brain. However, with the advantages of specificness is to brain tumors, rather, at the physiological level, when the tumour is resectable, this technique does not reveal the effect on the tumor is unresectable, fairly early stage of development, which is particularly important in the treatment of oncological pathologies, especially the Central nervous system.

As a promising vector for delivery of therapeutic agents for the treatment of tumors, mainly glioblastoma, are increasingly seen hematopoietic stem cells (hereinafter GSK). Human and murine HSCs have increased trapnest (positive homing) to the hearth intracranial tumors and are inert with respect to normal tissue of the Central nervous system. Research on organotypic cultures of hippocampus and in vivo, including ours, the fact of the directed migration of HSC to glioma cells.

The advantages of using HSC in therapeutic use for the treatment of intracranial gliomas is caused more by the fact that the traditional therapy of anticancer drugs and radiation therapy, there is a negative impact on migration processes GSK in the hearth of pathology (5). In addition, GSK are not embryonic cells, it is possible to select from the blood of an adult organism, which distinguishes them from the NSC.

Technical task, therefore, was the creation of a new antitumor agents having immunotropic is here to tumor cell glial origin, selected without the use of embryonic elements capable of vysokoeffektivnyi vector delivery to the brain tissue, with the property that penetrates through the BBB, aiming to influence on glial cell, keeping the inertia in relation to healthy cells; the method of obtaining such a means and method of inhibition of tumor in brain tissue.

The problem is solved in that the generated anti-tumor agent, representing immunoliposomes design, including liposome containing antitumor therapeutic agent is made with the vector peptide for the treatment of tumors in the Central nervous system, the liposome containing therapeutic agent in the aqueous phase, as a vector contains monoclonal antibodies to CD34+, and the stitching is a 2-aminothiols (it) in 0.1%concentration. As a therapeutic agent immunoliposome may contain a substance selected from the group: daunomycin, karminomitsin, melphalan, methotrexate, cytarabine, doxorubicin, ricin.

Search vectors for targeted impact on Central nervous system tumors led us, therefore, to the use of hematopoietic stem cells, as there are genetic aspects, explaining the possibility of such homing of these cells, in particular cells of glio the s. Although there is an opinion that the vast majority of gliomas does not Express the gene CXCL12 (6), but in other work using the more sensitive technique of PCR (polymerase chain reaction) researchers showed the presence of mRNA encoding CXCL12, in 13 of the 31 lines gliomas (7), but it should be borne in mind that the interaction of the chemokine CXCL12 (or SDF-1 alpha) receptor CXCR4 leads to active kinase phosphorylation mitogenactivated protein, which in turn initiates DNA synthesis and causes chemotaxis of tumor cells, that is responsible for tumor growth and metastasis.

The invention is confirmed by a number of experiments conducted both in vitro and in vivo: further experimental confirmation of the ability of therapeutic agent directly penetrate into tumor cells of the brain.

EXAMPLE 1. Create coculture C6 glioma and hematopoietic CD34 stem cells (HSC CD34), (EXPERIENCE).

To obtain a culture of glioblastoma used line C6-glioma

Take 1·10⁶cells C6 gliomas, quickly thawed, washed by centrifugation from cryoconserved (dimethyl sulfoxide), resuspended in 12 ml of medium (DMEM, 20% fetal bovine serum, 2 mm L-glutamine, 25 mm HEPES, antibiotic-antimycotic 10,000 units/ml) and are grown in culture mattresses (Costar). Cultivation of gliomas continue to about the adowanie monolayer. Next glioma collected by enzymatic dissociation (0.05% trypsin-EDTA, 10 min, 37° (C) and used in the second stage of the experiment.

To create coculture use culture insert company Millipore (diameter 12 mm, pore size 0.4 µm). The bottom of wells of a 12-hole culture of the tablet is covered with polyethylenimine (Gibco), followed by laminin (Gibco) according to the manufacturer's recommendations. Next, each well of the tablet is placed culture insert Millipore, where it immobilized a drop of sterile paraffin. Inside the culture insert contribute 0.5·10⁶cells C6-glioma, normal astrocytes rat normal rat fibroblasts, 6, 3 and 2 wells, respectively. One of the inserts leave blank. Tablet with culture inserts incubated for 24 hours. Then at the bottom of the wells are planted hematopoietic CD34+ stem cells to 0.25·10⁶cells stained with a fluorescent marker Vybrant™ CFDA SE Cell Tracer. After 3 hours of incubation is not adherent to the bottom of the wells, and the cells are selected.

Then visually analyze the distribution of hematopoietic CD34+ stem cells in the bottom wells. Counting cells in the area of projection of the membrane of the culture insert is performed using the program densitometric image analysis on the 1st, 2nd, 3rd, 5th and 10th day of cultivation. In the Ana is the study of the obtained data, the number of hematopoietic CD34+ stem cells in the area of projection of the membrane, containing cells in the C6 glioma, increased by 6% - 22% for the first 2 days of cocultivation. For subsequent days of cultivation, the number of HSC in the area of projection of the membrane was not significantly changed.

Figure 1 shows the micrograph of the culture of C6-glioma rats.

The a - lifetime micrograph, phase contrast,

In immunocytochemical staining with anti-GFAP (gliofibrillary acidic protein) mab, rhodamine, nuclei stained DAPI.

EXAMPLE 2. Create coculture normal astrocytes and (HSC CD34+), (CONTROL).

To obtain primary cultures of rat astrocytes were used newborn rats of the Wistar breed. Animals killed by ether anesthesia, the body surface is treated with alcohol. Under sterile conditions from rats separated the head, open the skull, take out the brain and placed in a Petri dish containing Hanks solution. Next, allocate the cerebral cortex, clean it from the vascular membranes, crushed, washed 2 times with Hanks solution and subjected to enzymatic dissociation of 0.05% solution of trypsin-EDTA for 10 minutes at 37°C. Enzymatic dissociation stopped by the addition of 5% FBS (fetal bovine serum) in Hanks solution, washed twice and subjected to mechanical dissociation. The cell suspension is then precipitated by centrifugation, resuspended in complete medium (DMEM/F12, 90%, FBS 10%, L-glutamine 2 mm, glucose and 0.8%, nsulin of 0.2 units/ml, HEPES 25 mm, antibiotic-antimycotic 10,000 units/ml) and planted in a culture mattresses (Costar). The cultivation of astrocytes continue to the formation of the monolayer. Next astrocytes harvested using enzymatic dissociation (0.05%trypsin-EDTA, 10 min, 37° (C) and used in the second stage of the experiment.

To create coculture use culture insert company Millipore, as in example 1 (diameter 12 mm, pore size 0.4 µm). The bottom of the hole 12 hole cultural tablet cover with polyethylenimine (Gibco), followed by laminin (Gibco) according to the manufacturer's recommendations. Next, each well of the tablet is placed culture insert Millipore, where it immobilized a drop of sterile paraffin. Inside the culture insert contribute 0.5·10⁶cells C6-glioma, normal rat astrocytes in the wells, respectively. One of the inserts leave blank. Tablet with culture inserts incubated for 24 hours. Then at the bottom of the wells are planted hematopoietic CD34+ stem cells to 0.25·10⁶cells stained with a fluorescent marker Vybrant™ CFDA SE Cell Tracer. After 3 hours of incubation is not adherent to the bottom of the wells, and the cells are selected.

Then visually analyze the distribution of hematopoietic CD34+ stem cells in the bottom wells. Counting cells in the area of projection of the membrane of the culture insert assests who are using densitometric image analysis Photo-Capt v. 12.4 on the 1st, 2nd, 3rd, 5th and 10th day of cultivation. The analysis of the obtained data, the number of hematopoietic CD34+ stem cells in the field of projection containing astrocytes, significant changes in the number of hematopoietic CD34+ stem cells in this control was not observed.

Figure 2 presents the micrograph culture of normal rat astrocytes.

The a - lifetime micrograph, phase contrast,

In immunocytochemical staining of the cultures of astrocytes anti-GFAP mab.

EXAMPLE 3. Create coculture fibroblasts and (HSC CD34+), (CONTROL).

To obtain primary cultures of rat fibroblasts using rat 12-day embryos. Female rats killed by ether anesthesia, the body is rubbed with alcohol. Next, sterile open the abdominal cavity, from the horns of the uterus are removed embryos. The embryos are placed in the Hanks solution with the addition of the antibiotic. The embryo is removed limbs, head and internal organs. The remaining mass is crushed and subjected to enzymatic dissociation (0.25% trypsin, 40 minutes, 37°). Cells precipitated by centrifugation (1000 g, 5 min). Sediment resuspended in complete medium (DMEM/F12, 10% FBS, 2 mm L-glutamine, 0.8% of glucose, insulin and 0.2 units/ml, 25 mm HEPES, antibiotic-antimycotic 10,000 units/ml) and planted in a culture mattresses (Costar). Next fibroblasts collected using the enzymatic dissocia the AI (0.05% trypsin-EDTA, 10 min, 37° (C) and used in the second stage of the experiment.

To create coculture use culture insert company Millipore, as in example 1 (diameter 12 mm, pore size 0.4 µm). The bottom of the hole 12 hole cultural tablet cover with polyethylenimine (Gibco), followed by laminin (Gibco) according to the manufacturer's recommendations. Next, each well of the tablet is placed culture insert Millipore, where it immobilized a drop of sterile paraffin. Inside the culture insert contribute 0.5·10⁶cells C6-glioma, normal astrocytes rat normal rat fibroblasts, 6, 3 and 2 wells, respectively. One of the inserts leave blank. Tablet with culture inserts incubated for 24 hours. Then at the bottom of the wells are planted hematopoietic CD34+ stem cells to 0.25·10⁶cells stained with a fluorescent marker Vybrant™ CFDA SE Cell Tracer. After 3 hours of incubation is not adherent to the bottom of the wells, and the cells are selected.

Then visually analyze the distribution of hematopoietic CD34+ stem cells in the bottom wells. Counting cells in the area of projection of the membrane of the culture insert is performed using the program densitometric image analysis Photo-Capt v. 12.4 on the 1st, 2nd, 3rd, 5th and 10th day of cultivation. The analysis of the obtained data in the area of projection of the membrane, the content is soup fibroblasts significant changes in the number of hematopoietic CD34+ stem cells in this control was not observed.

The results of this test are visualized in figure 3 (micrograph culture of normal rat fibroblasts).

Examples confirm the high trapnest HSC CD34+ pianim cells. We conclude that antibodies to CD34+ can be tested as vectors for targeted delivery of adequate anti-cancer drugs to the brain in the presence of the latter corresponding to the pathological process. As the original form of therapeutic tools we took a liposomal structure.

Another object of the invention is a method of obtaining the above-described anti-cancer drugs to combat tumors of the Central nervous system. The method consists in the following:

The way to obtain anti-cancer agents, including mixing 5 parts cholesterol, 15 parts of lecithin, 5.0 parts of distearoylphosphatidylcholine conjugated to polyethylene glycol, then all the components are dissolved in chloroform-methanol (1:9) mixture and dried in the form of a film, to the resulting lipid film add cyclohexane, frozen and lyophilized, lyophilized lipids dissolved in the aqueous phase of 0.1 M sodium phosphate buffer (pH=7,4), adding 0.5 to 2.5 weight. part of therapeutic agent, receiving liposomes, with the holding antitumor agent, then 10 parts by volume MA CD34+ mixed with 1 volume part of a solution of 2-aminothieno taken at 0.1%concentration, incubated for about one hour at room temperature in the dark, then separated from unbound 2-aminothieno through column chromatography (size 1×15 cm)with Sephadex G-25, equilibrated sodium phosphate buffer, then mixed suirvey etiolirovannye protein with 10 volume parts of liposomes and incubated 12 hours at a temperature of +4°With results in immunoliposomes structure containing the water portion of therapeutic agent, and as a vector - monoclonal antibodies to CD34+.

In the following examples are explained receiving antineoplastic agents in the form of immunoliposomes design.

EXAMPLE 4. Synthesis of liposomes.

Liposomes are prepared according to the method described in the article (8) with our modifications.

All manipulations except for short-term (weighing, centrifugation) was carried out in argon atmosphere. In liposomes composed of 5 mg cholesterol, 15 mg of lecithin, 5.0 mg of distearoylphosphatidylcholine conjugated to polyethylene glycol (cells of the dspe-PEG), 3.0 mg cells of the dspe-PEG conjugated to maleimide, 0.33 mg of fluorescent labels (in this case, Dil) for visualization of the resulting effect. All components are dissolved in chloroform-methanol (ratio shall osenia 1:9) mixture and dried in a rotary evaporator (about 1.5 hours). To the lipid film add cyclohexane, frozen in liquid nitrogen and lyophilized. Lyophilized lipids dissolved in 1.25 ml of 0.1 M sodium phosphate buffer (pH=7,4)containing 40 μg of fluorescent labels - Sytox Green (Molecular Probes), which is in this case the simulator therapeutic agent is hydrophilic fluorescent nucleic acid dye, which is unable to penetrate intact cell. Therefore, staining of the nucleus can occur only in the case of fusion of the liposomes with the cell-target and migrate into the content of the aqueous phase. Emulsion incubated, usually on a rotary evaporator at room temperature for about 12 hours.

Lipid emulsion further ekstragiruyut sequentially through a membrane with a pore diameter of 400, 200, 100 and 50 nm for receiving liposomal particles with included visualizing the label or therapeutic agent for subsequent in vivo tests.

Simulator therapeutic agent in this method may be replaced with a therapeutic agent, in our case selected from the group: daunomycin, karminomitsin, melphalan, methotrexate, cytarabine, doxorubicin, and ricin. As therapeutic agents can be used, and other suitable substances.

EXAMPLE 5. Getting immunoliposomes biological design CD34+.

Received what I immunoliposomes design using liposomes prepared as described in example 4.

Antibodies to CD34 antigen were obtained by the method of hybridoma technology. At the same time as antigen using recombinant peptide, identical to extracellar domain CD34 protein.

200 µg of monoclonal antibodies to CD34 mixed with 20 μl of a solution of 2-aminothieno (1 mg/ml), incubated for 1 hour at room temperature in the dark, then separated from unbound 2-aminothieno through column chromatography with Sephadex G-25 (1×15 cm), equilibrated sodium phosphate buffer. Control is carried out spectrophotometrically at λ=280 nm. Then mix suirvey etiolirovannye protein with 200 µl of liposomes and incubated 12 hours at a temperature of +4°C.

To stop the reaction conjugation with antibodies to liposomes add a 100-fold molar excess of a solution of N-ethyl-maleimide (1 mg/ml), concentrate on the cone to 0.5 ml and applied to a gel filtration column (1×47 cm) with separate CL4B. The elution spend sodium phosphate buffer. On the column of immunoliposome separated from the unbound N-ethyl-maleimide and not included inside the dye. Inspection is carried out visually by staining Dil and when λ=280 nm. Collect Paglierani liposomes in 3 ml of buffer solution and concentrate to the required volume. The resulting suspension is incubated with GSK for 30 minutes on an orbital necks the ore at 200 rpm When this occurs, the binding vector (anti-CD34 Mab)conjugated with liposome surface GSK.

Scheme immunoliposome presented in figure 4.

The obtained cell-liposomal tool with trapnest to tumor cells of the brain and contains within liposomes antitumor agent in the form of doxorubicin (in aqueous phase) and some other toxins in our test, with a highly efficient vector in the form sewn to the liposome antibodies to CD34, can be used for targeted delivery into brain tissue. This is confirmed by our experiments.

As a cytotoxic agent in the composition immunoliposome can be used drugs with antitumor activity and bearing free amino groups, such as daunomycin, karminomitsin, melphalan, methotrexate, cytarabine, doxorubicin, and cytotoxic drugs protein nature, such as toxins (diphtheria or ricin), and other suitable to fight brain tumors. Here is the example of an experiment conducted on animals.

EXAMPLE 6. Confirmation of the possibility of targeted delivery of antitumor immunoliposome in the brain.

Wistar rats (body weight 80 g, 8 animals per group) administered intraperitoneally injected (0.1 ml/10 g body weight) immunoliposomes design, made according to example 5, with antibodies is m, labeled¹²⁵I (1 MCI/ml)dissolved in a mixture of 1.5% (wt./about.) copolymer Pluronic R and 2.5% (wt./about.) copolymer Pluronic L64, dissolved in RPMI medium 1640.¹²⁵I-labeled polypeptide dissolved in medium RPMI 1640, was administered in the same concentration as the control series).

Three days later the animals were killed by anesthesia and received tissue samples for radioactive analysis of the distribution of labels in the tissues. The distribution of radioactivity was quantified using liquid scintillation counting.

The experiments were repeated at least twice and the reproducibility of the results was more than 90%. These results, expressed as the ratio of radioactivity in brain tissue to radioactivity in other tissues, are presented in the following table 1.

The table shows that the use of the drug in vivo in rats for brain tissue considerably exceed the possibility of its application in other tissues, and tests coculture proves the possibility of targeted delivery of anticancer drug in a tumor cell.

Another object of the present invention is a method of inhibiting tumor in the brain in experimental animals. Method of inhibiting neoplastic process is as follows.

Method of inhibiting tumor growth through vector delivery immunoliposomes design in the tumor of the CNS, characterized by the fact that the antitumor agent described previously, and obtained by the method described in examples 4 and 5, is administered parenterally in a pharmaceutically suitable carrier in an effective amount, with pre-enter product hematological stem cells CD34+.

EXAMPLE 7. Inhibition of tumor growth in models of tumor in the brain.

In the experiment used the Wistar rats. Model of brain tumor get put the m introduction cell culture C6-glioma. After a few days in rats developing glioblastoma. The groups used rats as in example 6 (weight 80 g, 8 animals per group). Used immunoliposomes design with included in the aqueous phase of the liposome doxorubicin. The drugs were injected at doses of 0.2 μg/kg of doxorubicin scheme: once in 2 days, a total of five injections, starting from the 3rd day after inoculation of the tumor. Pre-injected intravenous drug CCD34+ based 1,3(0,005)×10⁶cells/kg with native potential for homing in the hearth of the tumor process.

For control was introduced in experimental animals, inoculated with tumor doxorubicin subcutaneously. Counting the surviving animals showed that in the experimental groups, the life span of the animals was significantly increased within the 25 to 35%.

Similar results were obtained when testing daunomycin, karminomitsin, melphalan, methotrexate, ricin and other therapeutic agents against tumors.

The creation of antineoplastic agents in the form of a liposomal vector constructs containing therapeutic protivoopujolevy agent with which it is possible to implement the method of the directed transport of such agents in brain tissue affected by a malignant process, using as a vector of monoclonal antibodies to CD34+derived from hemopet the ical stem cells, provides a broad perspective of the creation of medicines vector type for the treatment of various severe pathologies of the Central nervous system.

LITERATURE

1. Tetsuya Hamaguchi, Yasuhiro Matsumura, Yukihiro Nakanishi,K. ei Muro, Yasuhide Yamada, et al. Cancer Sci, July 2004 vol., 95 no. 7.

2. Karen S. Aboody, Alice Brown, Nikolai G. Rainov, Kate A. Bower, Shaoxiong Liu et al. PNAS, 2000.

3. Connors So A.J. Gene Ther. 1995; 2: 702-709.

4. Bentires A.M., Hellin A.S., Lechanteur S., et al. J. Cancer Gene Ther. 2000; 7, 20-26.

5. Tabatabai G, Frank C., Mohle R, Weller M, Wick W. Brain. 2006.

6. Zhou Y., P. H. Larsen, C. Hao, Yong V. W. J. Biol. Chem. 2002 Dec 20; 277(51): 49481-7, Epub 2002 Oct 17.

7. A. Bajetto, F. Barbieri, Dorcaratto A. et al. Newochem Int. 2006 Apr 16.

8. N. Shi, W. Pardridge (PNAS, 2000, 97(13), 7567-72).

1. Antitumor agent representing immunoliposomes biological structure, comprising a liposome containing an antitumor therapeutic agent is made with the vector peptide, characterized in that for the treatment of tumors in the Central nervous system (CNS) liposome containing therapeutic agent in the aqueous phase, as a vector contains monoclonal antibodies to CD34+ (MA CD34+), and the stitching is a 2-aminothiols (it) in 0.1%concentration.

2. Antitumor agent according to claim 1, characterized in that as a therapeutic agent immunoliposome contains a substance selected from the group: daunomycin, karminomitsin, melphalan, methotrexate, cytarabine, doxorubicin, ricin.

3. The method of obtaining the anti-Christ. pwholesale means, includes mixing 5 parts cholesterol, 15 parts of lecithin, 5.0 parts of distearoylphosphatidylcholine conjugated to polyethylene glycol, then all the components are dissolved in chloroform-methanol (1:9) mixture and dried in the form of a film, to the resulting lipid film add cyclohexane, frozen and lyophilized, lyophilized lipids dissolved in the aqueous phase of 0.1 M sodium phosphate buffer pH 7.4 by adding 0.5 to 2.5 weight. including a therapeutic agent, receiving liposomes containing anti-tumor agent, and then about 10. H. MA CD34+ is mixed with about 1. PM solution of 2-aminothieno taken at 0.1%concentration, incubated for about one hour at room temperature in the dark, then separated from unbound 2-aminothieno through column chromatography with Sephadex G-25, balanced sodium phosphate buffer, then mixed suirvey etiolirovannye protein with about 10. including liposomes and incubated 12 h at a temperature of 4°With results in immunoliposomes structures containing in the water portion of therapeutic agent, and as a vector MA CD34+.

4. Method of inhibiting tumor growth, characterized by the fact that the tumor CNS vector is delivered immunoliposomes design in the form of anti-cancer agents, as described in claim 1 or 2, obtained by the method described is in clause 3, administered parenterally in a pharmaceutically suitable carrier in an effective amount, with pre-parenteral drug hematological stem cells CD34+.