Compounds

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to new purine derivatives possessing the properties of an inhibitor of the enzyme CDK specified in CDK1, CDK2, CDK3, CDK4, CDK6, CDK7, CDK8 and CDK9. In formula (I): R1 and R2 each independently represents H, C1-6alkyl or C1-6halogenalkyl; R3 and R4 each independently represents H, C1-6-alkyl or C1-6-halogenalkyl; R5 represents C1-6-alkyl or C3-12-cycloalkyl, or C3-12-cycloalkyl-C1-6-alkyl each of which may be optionally substituted by one or more OH groups; R6 represents wherein Y represents N, X and Z represents CR9; R7, R8 and R9 optionally represent H, alkyl or C1-6-halogenalkyl; wherein at least one of R7 , R8 and R9 is other than H. The invention also refers to a pharmaceutical composition containing said compounds, using the compounds for treating alopecia, stroke, a proliferative disease, such as cancer, leukaemia, glomerulonephritis, rheumatoid arthritis, psoriasis, viral diseases, such as a disease caused by human cytomegalovirus, type 1 herpex simplex virus, type 1 human immunodeficiency virus, a neurodegenerative disease, a CNS disease, such as Alzheimer's disease.

EFFECT: preparing new purine derivatives possessing the properties of the inhibitor of the enzyme CDK.

30 cl, 8 tbl, 18 ex

 

2-Ola,the Scope of the invention

The invention relates to novel 2,6,9-substituted purine derivatives and their biological application. In particular, the present invention relates to purine derivatives with antiproliferative properties that are suitable for the treatment of proliferative diseases such as cancer, leukemia, psoriasis, and the like.

Background of invention

Initiation, development and completion of the cell cycle in mammals are regulated by different cyclin-dependent kinase (CDK) complexes, which are crucial for cell growth. Such complexes include at least the catalytic (CDK) and a regulatory (cyclin) subunit. Some of the more important complexes for cell cycle regulation include cyclin A (CDK1 - also known as cdc2 and CDK2), cyclin B1-B3 (CDK1), cyclin D1-D3 (CDK2, CDK4, CDK5, CDK6), cyclin E (CDK2). Each of these complexes are involved in a specific phase of the cell cycle. However, not all members of the family of CDK involved solely in the control of cell cycle. For example, CDK 7, 8 and 9 are involved in the regulation of transcription, whereas CDK5 plays a role in the function of neuronal and secretory cells.

The activity of cdks is regulated by excision of temporary connections with other proteins and by changes in their subcellular localization. Developed the e tumor is closely linked to the genetic modification, and impaired regulation of cdks and their regulators, that suggests that CDK inhibitors may be useful anti-cancer therapeutic agents. Of course, the initial results suggest that transformed and normal cells differ in their requirements regarding, for example, cyclina A/CDK2, and that it is possible to develop new anticancer agents that do not have common toxicity to the host organism observed in conventional cytotoxic and cytostatic medicines. Although inhibition associated with cell cycle CDK is clearly important, for example, for use in Oncology, this may not be an argument for inhibition of regulatory RNA polymerase CDK. On the other hand, inhibition of CDK9 function/cyclina T until recently was associated with the prevention of HIV replication, and the opening of the new biology CDK, thus, continues to open new therapeutic indications for CDK inhibitors (Sausville, E.A. Trends Molec. Med. 2002, 8, S32-S37).

Function CDK is the phosphorylation and thus activation or deactivation of certain proteins, including, for example, proteins retinoblastoma, luminy, histone H1 and components of the mitotic spindle. Catalytic stage, mediated by the CDK, involves the reaction of phosphopentose from ATP to the macromolecular substrate of the enzyme. Several groups of compounds (reviewed, for example, Fischer,P.M. Curr. Opin. Drug Discovery Dev. 2001, 4, 623-634), was found to have antiproliferative properties in virtue of CDK-specific antagonism of ATP.

In WO 98/05335 (CV Therapeutics Inc) described 2,6,9-Transnistria purine derivatives which are selective inhibitors of kinases of the cell cycle. Such compounds are useful in the treatment of autoimmune diseases such as rheumatoid arthritis, lupus, type I diabetes, multiple sclerosis; cancer, cardiovascular diseases such as restenosis, the reaction of the host against the graft, gout, polycystic kidney disease and other proliferative diseases, pathogenesis of which is associated with abnormal cell proliferation.

In WO 99/07705 (The Regents of The university of California) described purine analogues that inhibit, inter alia, protein kinases, G-proteins and polymerases. More specifically, the present invention relates to methods of applying such purine analogues for the treatment of diseases with cell proliferation and neurodegenerative diseases.

In WO 97/20842 (CNRS) also described purine derivatives exhibiting anti-proliferative properties that are useful in the treatment of cancer, psoriasis and neurodegenerative disorders. Additional purine derivatives described in WO 03/002565, WO 04/016613 and WO 04/016612.

By this invention, an attempt was made to obtain new 2,6,9-ameenah purine derivatives, especially those that have antiproliferative properties.

The invention

In the first aspect of the present invention relates to the compound of formula (I) or its pharmaceutically acceptable salt

where:

R1and R2each independently represents H, alkyl or halogenated;

R3and R4each independently represents H, alkyl, halogenated or aryl;

R5represents an alkyl or cycloalkyl, or cycloalkenyl, each of which may be optionally substituted by one or more Oh groups;

R6selected from cyclopropylamino, cyclopropanemethylamine, cyclobutylamine, cyclobutylamine and

where one of X, Y and Z represents N and the remaining represent CR9;

R7, R8and each R9independently represent H, alkyl or halogenated; where at least one of R7, R8and each R9is not N.

In a second aspect the present invention relates to the compound of formula II or its pharmaceutically acceptable salt,

where:

at least one of R1', R2', R3'and R4'is halogenation, and the others each independently represents H, alkyl or g is loginall;

R5'represents an alkyl or cycloalkyl, or cycloalkenyl, each of which may be optionally substituted by one or more OH groups;

R6'selected from cyclopropylamino, cyclopropanemethylamine, cyclobutylamine, cyclobutylamine and

where X', Y' and Z'each independently represents CR9'or one of X', Y' and Z' is N and the remaining represent CR9'; and

R7', R8'and each R9', each independently, represents H, halogen, alkyl or halogenated.

The third aspect of the present invention relates to pharmaceutical compositions containing the compound of the present invention and a pharmaceutically acceptable carrier, diluent or excipient.

A fourth aspect of the present invention relates to the use of compounds of the present invention to obtain drugs for the treatment of one or more of the following diseases:

proliferative disease;

viral diseases;

blow;

alopecia;

Central nervous system disorders;

neurodegenerative disorders and

different types of diabetes.

The fifth aspect of the present invention relates to the use of compounds of the present invention as antimitoticescoy tools.

The sixth aspect of the present izaberete the Oia applies to the use of compounds of the present invention for inhibiting protein kinases.

The seventh aspect of the present invention relates to a method of treatment of a proliferative disease, where the method includes the administration to a mammal a therapeutically effective amount of the compounds of the present invention.

The eighth aspect of the present invention relates to the use of compounds of the present invention for analysis to identify additional candidates for compounds that affect the activity of one or more CDK enzymes.

Detailed description of the invention

As described above, the first aspect of the present invention relates to the compound of formula (I)as defined above.

As used herein, the term "alkyl" includes alkyl groups as saturated straight chain and branched. Preferably, the alkyl group is a C1-20alkyl group, more preferably, From1-15even more preferably1-12alkyl group, and most preferably1-6alkyl group, more preferably, From1-3alkyl group. Especially preferred alkyl groups include, for example, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, pentyl and hexyl.

As used herein, the term "cycloalkyl" refers to a cyclic alkyl group. Preferably, cycloalkenyl is a C 3-12cycloalkyl group.

As used herein, the term "cycloalkenyl" refers to a group having cycloalkyl and functional alkyl group.

Preferably, one of R1and R2represents N and the other is alkyl.

More preferably, one of R1and R2represents N and the other is stands, ethyl or isopropyl.

In one preferred embodiment, R1represents ethyl, and R2represents N.

In one of the preferred embodiments of the present invention R3and R4each independently represents H, alkyl, halogenated or aryl; and at least one of R3and R4is not N.

In one of the preferred embodiments of the present invention one of R3and R4represents N and the other is alkyl or halogenation.

In a more preferred embodiment, R3represents N, and R5represents an alkyl or halogenated.

More preferably, R3represents N, and R4represents methyl.

In one of the preferred embodiments of the present invention R6represents a

In one of the preferred embodiments of the present invention Y is N. Preferably for this variant implementation, X is CH, Z is a C-alkyl, R7is N, and R8represents alkyl. More preferably for this variant implementation, X is CH, Z is a C-IU, R7is N, and R8represents Me. In an alternative preferred embodiment, X is CH, Z is a C-IU, R7and R8both are N. In yet another alternative preferred embodiment, X is CH, Z represents-CF3and R7and R8both are N.

In one of the preferred embodiments X is N. Preferably for this variant implementation, Y is C-Me, Z is CH, and R7and R8both are N. In yet another alternative preferred embodiment, Y and Z are CH, R7is N, and R8is IU.

In one of the preferred embodiments of the present invention Z is N. Preferably for this variant implementation, X is CH, Y is C-IU, R7is IU, and R8is N.

In another preferred embodiment, this is about the invention R 6represents cyclopropylamino, cyclopropanemethylamine, cyclobutylamine or cyclobutylmethyl.

In another preferred embodiment of the present invention R5is isopropyl.

In one of the most preferred embodiments, the compound of the present invention are selected from the following:

[1](2R,3S-3-(6-((4,6-dimethylpyridin-3-ylmethylamino)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol
[2]2R,3S-3-(9-isopropyl-6-((6-methylpyridin-3-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol
[6]2R,3S-3-(6-cyclopropylamino)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol
[7]2R,3S-3-(6-(cyclopropylamino)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol
[8]2R,3S-3-(6-(cyclobutylamine)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol
[10]2R,3S-3-(9-isopropyl-6-(2,6-dimethylpyridin-4-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol
[11]2R,3S-3-(9-isopropyl-6-((6-trifluoromethyl)pyridin-3-yl)methylamino-N-purine-2-ylamino)pentane-2-ol
[12]2R,3S-3-(9-isopropyl-6-((6-methylpyridin-2-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol
[13]2R,3S-3-(9-isopropyl-6-((3-methylpyridin-2-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol
[15]1,1,1-Cryptor-3-(9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)propan-2-ol
[16]1,1,1-Cryptor-3-(9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol
[17]1,1,1-Cryptor-3-(9-isopropyl-6-((6-(trifluoromethyl)pyridin-3-yl)methylamino)-N-2-ylamino)pentane-2-ol
[18]1,1,1,3,3,3-hexamer-2-((9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)methyl)propane-2-ol

Another aspect of the present invention relates to the compound of formula II or its pharmaceutically acceptable salt

where:

at least one of R1', R2', R3'and R4'is halogenation, and the others each independently represents H, alkyl or halogenated;

R5'represents an alkyl or cycloalkyl, or cycloalkenyl, each of which can be neo is Astelin substituted by one or more OH groups;

R6'selected from cyclopropylamino, cyclopropanemethylamine, cyclobutylamine, cyclobutylamine and

where X', Y' and Z'each independently represents CR9'or one of X', Y' and Z' is N and the remaining represent CR9'; and

R7', R8'and each R9'independently represent H, halogen, alkyl or halogenated.

Preferably, in this aspect of the present invention R5'represents isopropyl.

Preferably, in this aspect of the present invention R6'represents a

In one of the preferred embodiments, Y' is N, X' and Z' are CH, and R7'and R8'both are N.

In another preferred embodiment of the present invention one of R1'and R2'is N, and the other is alkyl, or R1'and R2'both are N.

In another preferred embodiment of the present invention one of R3'and R4'is N, and the other is CF3.

In one of the most preferred embodiments, the compound of the present invention are selected from the following:

[15]1,1,1-rifter-3-(9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)propan-2-ol
[16]1,1,1-Cryptor-3-(9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol
[18]1,1,1,3,3,3-hexamer-2-((9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)methyl)propane-2-ol

In an additional aspect, the present invention relates to a compound selected from the following:

[1](2R,3S-3-(6-((4,6-dimethylpyridin-3-ylmethylamino)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol
[2]2R,3S-3-(9-isopropyl-6-((6-methylpyridin-3-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol
[3](2R,3S-3-(6-(3-chlorobenzylamino)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol
[4](2R,3S-3-(6-(3-forbindelsen)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol
[5]2R,3S-3-(9-(cyclopropylmethyl)-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol
[6]2R,3S-3-(6-cyclopropylamino)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol
[7]2R3S-3-(6-(cyclopropylamino)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol
[8]2R,3S-3-(6-(cyclobutylamine)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol
[9]2R,3S-3-(9-isopropyl-6-(pyridine-4-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol
[10]2R,3S-3-(9-isopropyl-6-(2,6-dimethylpyridin-4-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol
[11]2R,3S-3-(9-isopropyl-6-((6-trifluoromethyl)pyridin-3-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol
[12]2R,3S-3-(9-isopropyl-6-((6-methylpyridin-2-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol
[13]2R,3S-3-(9-isopropyl-6-((3-methylpyridin-2-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol
[14](R)-1-(9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)propan-2-ol
[15]1,1,1-Cryptor-3-(9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)propan-2-ol
[16]1,1,1-Cryptor-3-(9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol
[17]1,1,1-Cryptor-3(9-isopropyl-6-((6-(trifluoromethyl)pyridin-3-yl)methylamino)-N-2-ylamino)pentane-2-ol
[18]1,1,1,3,3,3-hexamer-2-((9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)methyl)propane-2-ol

In one particularly preferred embodiments, the compound of the present invention are selected from the following:

[3](2R,3S-3-(6-(3-chlorobenzylamino)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol
[4](2R,3S-3-(6-(3-forbindelsen)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol
[5]2R,3S-3-(9-(cyclopropylmethyl)-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol
[9]2R,3S-3-(9-isopropyl-6-(pyridine-4-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol
[14](R)-1-(9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)propan-2-ol

In one particularly preferred embodiment of the present invention the compound is selected from compounds[1], [3], [4], [6], [7], [8], [9], [10] and [11]. More preferably, this compound selected from compounds[1], [3], [4], [6] [11]even more preferably, when the data connection is the connection [1] or [11].

Preferably, the compound of the present invention exhibits at least 3-fold increased activity compared to seliciclib, more preferably at least 4-fold or 5-fold increase in the activity, even more preferably at least 8-fold or 10-fold increased activity.

In one particularly preferred embodiment of the present invention in this connection is (2R,3S-3-(6-((4,6-dimethylpyridin-3-ylmethylamino)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol [1] or its pharmaceutically acceptable salt or ester.

Mainly, the connection [1] shows unexpectedly high activity analyses at the cellular toxicity in a number of different cell lines in comparison with structurally related compounds, known to specialists in this field. Additional details of these analyses are presented in the accompanying examples (see, in particular, table 8).

In addition, experiments have shown that in contrast to the structurally related compounds known to specialists in this field, the compound [1] is not significantly inhibits CYP3A4. And also, these analyses are described in more detail in the accompanying examples (see, in particular, table 5). Of course, the connection [1], as can be seen, did not inhibit CYP3A4 to concentrations of more than 20 μm, which is ~60-Crat who left his cell IC 50. As the value of the IC50for inhibition of CYP3A4 for inhibiting compound [1] is significantly higher than its cellular IC50(see table 8), this shows that at cytotoxic concentrations in this case should not have impact on the activity of CYP3A4. This is important because CYP3A4 is involved in the metabolism of a large number of medicines. If CYP3A4 is inhibited by one drug, it can lead to unexpected toxicity due to reduced metabolism of CYP3A4 substrates, leading thus to markedly elevated levels of such funds.

Similarly, additional experiments showed that in contrast to its structurally related analogues of compound [1] is not a substrate for the six tested CYP isoforms (see, in particular, table 6). This difference corresponds to the observed difference in the inhibition of CYP discussed above. The usual mechanism leading to inhibition of CYP, is evident, if this connection is also a substrate for this CYP.

Accordingly, the compound [1] is not a substrate for CYP3A4 or inhibitor of CYP3A4, which gives an important, unexpected beneficial property, compared with its structurally related analogues.

The pharmaceutical composition

The second aspect of the present invention relates to pharmaceutical compositions, the soda is containing compound of the present invention in a mixture with a pharmaceutically acceptable diluent, excipient or carrier, or mixtures thereof. Although the compounds of the present invention (including their pharmaceutically acceptable salts, esters and pharmaceutically acceptable solvate) can be entered separately, they will usually be applied in a mixture with a pharmaceutical carrier, excipient or diluent, especially for the treatment of humans. The pharmaceutical composition may be intended for use in humans or animals in medicine or veterinary medicine.

Examples of suitable excipients for various forms of the pharmaceutical compositions described herein, can be found in "Handbook of Pharmaceutical Excipients", 2nd Edition, (1994), edited by A. Wade and P.J. Weller.

Acceptable carriers or diluents for therapeutic use are well known to experts in the pharmaceutical field and are described, for example, in Remington's Pharmaceutical Sciences, Mack Publishing Co. (A.R. Gennaro edit. 1985).

Examples of suitable carriers include lactose, starch, glucose, methyl cellulose, magnesium stearate, lures, sorbitol and the like. Examples of suitable solvents include ethanol, glycerol and water.

The choice pharmaceutical carrier, excipient or diluent may be made in accordance with the intended route of administration and a single pharmaceutical practice. The pharmaceutical compositions may contain as carrier, excipient or razbaby the El, or in addition thereto, any suitable binder(s) substance(s), lubricant(s), suspendresume(s) substance(s), substance(a) to cover and solubilizers(s) substance(s).

Examples of suitable binders include starch, gelatin, natural sugars such as glucose, anhydrous lactose, free flowing lactose, beta-lactose, sweeteners made from corn, natural and synthetic gums such as gum acacia, tragakant or sodium alginate, carboxymethylcellulose and polyethylene glycol.

Examples of suitable lubricants include sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride, and the like.

In the pharmaceutical compositions can be presented preservatives, stabilizers, dyes and masking improves the taste and odor substances. Examples of preservatives include sodium benzoate, sorbic acid and esters of p-hydroxybenzoic acid. Can also be used antioxidants and suspendresume tools.

Salts/esters

Compounds of the present invention can be presented in the form of salts or esters, in particular pharmaceutically acceptable salts or esters.

Pharmaceutically acceptable salts of the compounds of the present invention include those suitable additive salts of acids or bases. An overview of suitable pharmaceutical salts see Berge et al., J. Parm. Sci., 66, 1-19 (1977). Salts are formed, for example, with strong inorganic acids such as mineral acids, e.g. sulfuric acid, phosphoric acid or a halogen acid, with strong organic carboxylic acids, such as alcancarao acids with 1-4 carbon atoms, which are unsubstituted or substituted (e.g., halogen), such as acetic acid, with saturated or unsaturated dicarboxylic acids, e.g. oxalic, malonic, succinic, maleic, fumaric, phthalic or tetraphthalate; hydroxycarbonate acids, for example ascorbic, glycolic, lactic, malic, tartaric or citric; amino acids for example aspartic or glutamic; benzoic acid, or with organic sulfonic acids, such as (C1-C4)alkyl - or arylsulfonic acids which are unsubstituted or substituted (e.g., halogen), such as methane - or p-toluensulfonate acid. Esters are formed, or with organic acids, or alcohols/hydroxides, depending on the functional group that you want to atrificial. Organic acids include carboxylic acids, such as alcancarao acid from 1-12 carbon atoms, which are unsubstituted or substituted (e.g., halogen), still is as acetic acid; with a saturated or unsaturated dicarboxylic acid, for example oxalic, malonic, succinic, maleic, fumaric, phthalic or tetraphthalate; hydroxycarbonate acids, for example ascorbic, glycolic, lactic, malic, tartaric or citric acid; and with amino acids, for example aspartic or glutamic acid; benzoic acid; or with organic sulfonic acids, such as (C1-C4)alkyl - or arylsulfonic acids which are unsubstituted or substituted (e.g., halogen), such as methane - or p-toluensulfonate acid. Suitable hydroxides include inorganic hydroxides, such as sodium hydroxide, potassium hydroxide, calcium hydroxide, aluminum hydroxide. The alcohols include alcamovia alcohols of 1-12 carbon atoms which may be unsubstituted or substituted, e.g. by halogen.

Enantiomers/tautomers

In all aspects of the present invention discussed above, the present invention includes, where appropriate, all enantiomers and tautomers of the compounds of the present invention. The person skilled in the art will recognize compounds that have optical properties (one or more chiral carbon atoms) or tautomeric characteristics. The corresponding enantiomers and/or tautomers can be provided is by or obtained by the methods, well-known experts in this field.

Stereo and geometric isomers

Some of the compounds of the present invention can exist as stereoisomers and/or geometric isomers, for example, they may have one or more asymmetric and/or geometric centres and so may exist in two or more stereoisomeric and/or geometric forms. This invention assumes the use of all individual stereoisomers and geometric isomers of such inhibitory agents and mixtures thereof. The terms used in the claims to cover all such forms, provided that these forms retain the appropriate functional activity (though not necessarily to the same degree).

The present invention also includes all suitable isotopic variations of this substance or its pharmaceutically acceptable salt. Isotopic variation means of the present invention or its pharmaceutically acceptable salt is defined as a variation, in which at least one atom is replaced by an atom having the same atomic number, but an atomic mass different from the atomic mass usually found in nature. Examples of isotopes that can be incorporated in the substance and its pharmaceutically acceptable salts, include isotopes of hydrogen, carbon, nitrogen is, oxygen, phosphorus, sulfur, fluorine and chlorine, such as2H,3H,13C,14C,15N17O,18O,31P,32P,35S18F and36Cl, respectively. Certain isotopic variations of the tool and their pharmaceutically acceptable salts, for example, which includes a radioactive isotope, such as3N or14That can be applied in medicine and/or for analyzing the distribution of the substrate in the tissues. Tretirovanie, ie3N, and carbon-14, i.e14With, isotopes are particularly preferred because of their easy access and detection. In addition, substitution with isotopes such as deuterium, i.e2N, may provide some therapeutic advantages in the greater metabolic stability, for example, increased half-life in vivo or reduced dose requirements, and hence may be preferred in some circumstances. Isotopic variations of the compounds of the present invention and pharmaceutically acceptable salts of the present invention can usually be obtained by conventional procedures using appropriate isotopic variations of suitable reagents.

The solvate

The present invention also includes solvated forms of the compounds of the present invention. The terms used in the formula image is etenia, cover all such forms.

Polymorphs

The present invention also relates to compounds of the present invention in their various crystalline forms, polymorphic forms and (without)water forms. In the pharmaceutical industry is well established that chemical compounds can be allocated in any of such forms by small changes in the method of cleaning and/or selection of the solvents used in the synthetic preparation of such compounds.

Prodrugs

The present invention also includes compounds of the present invention in the form of prodrugs. Such prodrugs are typically compounds of the present invention, having one or more respective groups modified so that modification may be reversible when administered to a human or mammal. Such reversibility is usually done with the help of an enzyme that is present in such entity, although it is possible the introduction of a second means together with such prodrug to produce this transformation in vivo. Examples of such modifications include esters (for example, any of those described above), and the transformation can be carried out by esterase, etc. Other such systems are well known to specialists in this field.

Introduction

Pharmaceutical HDMI is tion of the present invention may be adapted for oral, rectal, vaginal, parenteral, intramuscular, intraperitoneal, intrathecal, intrabronchial, subcutaneous, intradermal, intravenous, nasal, cheek, or sublingual routes of administration.

For oral administration, in particular, use compressed tablets, pills, pills, pills, gel, drops and capsules. Preferably, these compositions contain from 1 to 250 mg, and more preferably from 10 to 100 mg of active ingredient per dose.

Other forms for administration include solutions or emulsion that can be administered by injection, intravenously, intraarterially, intrathecally, subcutaneously, intradermally, intraperitoneally or intramuscularly, and which is prepared from sterile or sterilized solutions. The pharmaceutical compositions of the present invention can also be in the form of suppositories, pessaries, suspensions, emulsions, lotions, ointments, creams, gels, medications, aerosols, solutions and powders.

Alternative means of transdermal introduction presents the use of a skin patch. For example, the active ingredient can be incorporated into a cream consisting of an aqueous emulsion of polyethylene glycols or paraffin oil. The active ingredient can be included in a concentration of from 1 to 10% by mass in the ointment, consisting of the basics of bleached beeswax in the ka or white vaseline together with such stabilizers and preservatives, which may be necessary.

Injectable form may contain from 10 to 1000 mg, preferably from 10 to 250 mg of active ingredient per dose.

The composition can be manufactured in a single dosage form, for example, in the form of individual portions containing a single dose or as multiple divided doses single dose.

Dosage

The person skilled in the art can easily determine an appropriate dose of one of these compositions for administration to a patient without undue experimentation. The doctor usually determines the effective dose, which will be most appropriate for a particular patient, and it will depend on a number of factors, including the specific activity of the applied compound, the metabolic stability and length of action of this compound, the age, body weight, General health, sex, diet, mode and time of administration, rate of excretion, combination of drugs, the severity of the particular condition of the individual and the applied therapy. Dosages described herein, are examples for common cases. Of course, there may be individual examples, when shown a higher or lower intervals of doses, and such cases are covered by the scope of the present invention.

Depending on the needs of the tool t is to enter in a dose of from 0.01 to 30 mg/kg of body weight, such as from 0.1 to 10 mg/kg, more preferably from 0.1 to 1 mg/kg of body weight.

In the specific example of a variant of implementation of one or more doses of 10 mg to 150 mg/day will be administered to a patient for the treatment of malignant tumors.

Therapeutic use

Compounds of the present invention was found to have antiproliferative activity, and therefore is believed to be useful in the treatment of proliferative diseases, such as various types of cancer, leukemia and other diseases associated with uncontrolled cellular proliferation such as psoriasis and restenosis.

As defined herein, antiproliferative effect in accordance with the scope of the present invention can be demonstrated by the ability to inhibit the proliferation of cells in the in vitro assays on whole cells, for example, using any of the cell lines A, HeLa, HT-29, MCF7, Saos-2, CCRF-CEM, H460, HL-60 and K-562, or by demonstrating inhibition of the kinase with appropriate analysis. Such analyses, including the methods of their implementation, are described in more detail in the accompanying examples. The use of such tests can determine whether the connection is antiproliferative in the context of the present invention.

One preferred implementation of the present invention, therefore, relates to the note the of one or more compounds of the present invention to obtain drugs for the treatment of proliferative diseases.

In accordance with the description, the phrase "obtaining drugs" includes the use of compounds of the present invention directly as a medicine in addition to its use in the program selection on therapeutic tools in the future or at any stage of the production of such medicines.

The term "proliferative disease" is used herein in a broad sense, so that includes any disease that requires control of the cell cycle, for example, cardiovascular diseases such as restenosis and cardiomyopathy, autoimmune diseases, such as glomerulonephritis and rheumatoid arthritis, dermatological diseases such as psoriasis, fungal, parasitic diseases such as malaria, emphysema and baldness. With these disorders, the compounds of the present invention can induce apoptosis or to maintain stasis in the desired cells, when necessary. Preferably, the proliferative disease is a cancer or leukemia.

In another preferred embodiment, the proliferative disease is psoriasis.

Compounds of the present invention can inhibit any stage or all stages in the cell cycle, for example, the formation of the nuclear envelope, the phase-out of the rest of cleoc the th cycle (G0), G1 development, loosening of the chromosome, the rupture of the nuclear envelope, START, initiation of DNA replication, the development of DNA replication, termination of DNA replication, centrosome duplication, development G2, activation of the mitotic or maticevski functions, chromosome condensation, separation of the centrosome, the nucleation of microtubules, the formation and functioning of the spindle, the interaction with motor proteins of microtubules, chromatide separation and splitting, inactivation of mitotic functions, the formation of the contractile ring and functions of cytokinesis. In particular, the compounds of the present invention may affect some functions of genes, such as the binding of chromatin, formation of complexes of replication licensing replication, phosphorylation, or other activity of secondary modifications, proteolytic degradation, binding to microtubules, actin binding, the binding of septin activity nucleation organizing microtubules centre and linking components and signaling pathways of the cell cycle.

An additional aspect of the present invention relates to a method of treatment of a proliferative disease, where the method includes the administration to a mammal a therapeutically effective amount of the compounds of the present invention.

In the preferred embodiment of this aspect of prolif the operational condition is a cancer or leukemia.

In an even more preferred embodiment of this aspect, the compound is administered in an amount sufficient to inhibit at least one CDK enzyme.

Preferably, the compound of the present invention is administered in a quantity sufficient to inhibit at least one of CDK1, CDK2, CDK3, CDK4, CDK6, CDK7, CDK8 and/or CDK9.

More preferably, the compound of the present invention is used in a quantity sufficient to inhibit at least one of CDK2 and/or CDK4.

Even more preferably, the CDK enzyme is CDK2.

In one of the preferred embodiments of this aspect, the compound is administered orally.

Another aspect of the present invention relates to the use of compounds of the present invention as antimitoticescoy tools.

Another aspect of the present invention relates to the use of compounds of the present invention for the treatment of neurodegenerative disorders.

Preferably neurodegenerative infringement is apoptosis of neurons.

Another aspect of the invention concerns the use of compounds of the present invention as antiviral agents.

Thus, another aspect of the present invention relates to the use of compounds of the present invention to obtain the drug among the STV for the treatment of viral diseases such as a disease caused by human cytomegalovirus (HCMV), herpes simplex virus type 1 (HSV - 1), human immunodeficiency virus type 1 (HIV-1) and varicella zoster virus and herpes zoster (VZV).

In a more preferred embodiment of the invention, the compound of the present invention is used in a quantity sufficient for inhibition of one or more CDK host cell involved in viral replication, i.e. CDK2, CDK7, CDK8 and CDK9. [Wang d, De la Fuente C, Deng L., I. Zilberman, Eadie, C., Healey, M., Stein D., Denny, T., Harrison LE, L. Meijer, Kashanchi F. Inhibition of human immunodeficiency virus type 1 reduced by chemical cyclin-dependent kinase inhibitors. J. Virol. 2001; 75: 7266-7279].

As defined above, an antiviral effect in the scope of the present invention can be demonstrated by the ability to inhibit CDK2, CDK7, CDK8 or CDK9.

In a particularly preferred embodiment, the present invention relates to the use of one or more compounds of the present invention for the treatment of viral disease, which is a dependent or sensitive to CDK. Dependent CDK disease is associated with a level of activity above normal at one or more CDK enzymes. Such diseases are preferably associated with an abnormal level of activity of CDK2, CDK7, CDK8 and/or CDK9. Sensitive to CDK disease is a disease in which abnormal levels of CDK is not about the main reason but is the resulting consequence of the main metabolic abnormalities. In this programme of action, you can say that CDK2, CDK7, CDK8 and/or CDK9 are part of a sensitive metabolic pathways, and the CDK inhibitors so they can be active in the treatment of such diseases.

Compounds of the present invention is also suitable to obtain drugs for the treatment of various eye diseases. Preferably, the eye disease is glaucoma, exudative age-related macular degeneration (AMD) or proliferative diabetic retinopathy (PDR).

A painful condition called glaucoma, is characterized by persistent loss of visual function due to irreversible damage to the optic nerve. Several morphologically or functionally different types of glaucoma are typically characterized by elevated intraocular pressure (IOP), which is reputed to be causally associated with the pathological course of the disease. Ocular hypertension is a condition in which intraocular pressure is high, but the apparent loss of visual function has not occurred; these patients are considered as having a high risk of development of vision loss associated with glaucoma. Inhibitors of GSK-3 are useful for the treatment of eye diseases such as glaucoma. It has been shown that the Wnt with Gallinago path, associated with burn protein (FRP)are differentially expressed in several cell lines glaucomatous trabecular meshwork structure and can tear signaling cascade, leading to an increase in the resistance to outflow and the development of elevated IOP. Hellberg M.R. et al. (US20040186159) showed that through the interaction of GSK-3 with components of the Wnt signaling pathway inhibition of GSK-3 pharmacological means can be overcome mediated FRP antagonism of Wnt signaling pathway, caused by elevated levels of FRP, and to counteract the increasing resistance of the outflow, which is the result of increasing the production of FRP in individuals with glaucoma.

CTGF is secretively cytokine, which, as is known, increases the production of extracellular matrix (ECM), mainly by deposition of collagen I and fibronectin. The overexpression of CTGF was previously linked as the main causal factor with pathological conditions, such as scleroderma, fibroproliferative disease, scarring, etc. in which there is cornacopia components of the ECM. Cornacopia substances extracellular matrix in the area of trabecular mesh (TM) is also a symptom of many forms of glaucoma; this increase is believed, leads to increased resistance to the outflow of intraocular fluid and therefore to increased wew is ipathname pressure. Fleenor D.L. et al. (US20050234075) showed that inhibitors of GSK-3 and CDK inhibitors can inhibit both basic and induced TGF-beta 2 expression of CTGF in human TM cells, therefore, compounds of the present invention can be applied in the treatment of glaucoma.

Compounds of the present invention is also suitable for the treatment of AMD and PDR. Exudative age-related degeneration yellow spots (AMD) and proliferative diabetic retinopathy (PDR) are the main causes of the arising of blindness in developed countries and is characterized by pathological late segment neovascularization in the eye. The precipitating cause both AMD and PDR are still unknown, however, production of various Pro-angiogenic growth factors, apparently, is a common stimulus. Soluble growth factors such as vascular endothelial growth factor (VEGF), a growth factor derived from platelets (PDGF), basic fibroblast growth factor (bFGF or FGF-2), insulin-like growth factor 1 (IGF-1), angiopoietin etc. were found in the tissues and fluids taken from patients with pathological angiogenesis in the eye. Inhibition or blockade of the activity of these growth factors and other intracellular enzymes, such as aurora kinase, as shown, have antiangiogenic effects. Thus, the compounds of the present invention can be applied in the treatment of GLA the diseases, illnesses, characterized by neovascularization.

Another aspect of the present invention relates to the use of compounds of the present invention or their pharmaceutically acceptable salts for getting medicines for diabetes.

In a particularly preferred embodiment, the diabetes is type II diabetes.

GSK-3 is one of several protein kinases, which phosphorylate glikogensintetazy (GS). Stimulation of glycogen synthesis by insulin in skeletal muscle occurs as a result of dephosphorylation and activation of GS. The effect of GSK-3 GS, thus, leads to deactivation of the latter and, thus, inhibition of transformation of glucose into glycogen in the muscles.

The type II diabetes (insulin dependent diabetes mellitus) is a multi-factorial disease. Hyperglycemia associated with insulin resistance in liver, muscle and other tissues in combination with impaired secretion of insulin. Skeletal muscle is the main site-stimulated insulin glucose uptake, here she is, or is removed from the bloodstream, and / or converted to glycogen. The deposition of glycogen in the muscles is a major determinant of glucose homeostasis and diabetes type II is defective deposition of glycogen in the muscles. There is some evidence that the increase in the activity of GSK3 is important in diabetes type II [Chen, Y.H.; Hansen, L.; Chen, M.x.; Bjorbaek, C.; Vestergaard, H.; Hansen, T.; Cohen, P.T.; Pedersen, O. Diabetes, 1994, 43, 1234]. In addition, it was shown that diabetes type II is the overexpression of GSK3 in muscle cells, and that there is an inverse correlation between activityGSK3in skeletal muscle and insulin action [Nikoulina, S.E.; Ciaraldi, T.P.; Mudaliar, S.; Mohideen, P.; Carter, L.; Henry, R.R. Diabetes, 2000, 49, 263].

Inhibition of GSK3 therefore has therapeutic value in the treatment of diabetes, particularly type II, and diabetic neuropathy.

Interestingly, GSK3, as you know, phosphorylates many substrates, in addition to GS, and, thus, participates in the regulation of many biochemical pathways. For example, high expression of GSK occurs in the Central and peripheral nervous systems.

In another aspect the present invention relates to the use of compounds of the present invention or its pharmaceutically acceptable salts to obtain drugs for the treatment of CNS diseases, for example neurodegenerative diseases.

Preferably, the CNS disease is Alzheimer's disease.

Tau protein is a substrate of GSK-3, which is associated with the etiology of Alzheimer's disease. In healthy nerve cells Tau acompanhada with tubulin in microtubules. However, in Alzheimer's disease Tau forms a large plexus of fibers, which destroys the structure of microtubules in nerve cell is, breaking, thus, the transport of nutrients and signaling of nerve cells.

Without wanting to be bound by theory, believe that GSK3 inhibitors may be able to prevent and/or correct abnormal phosphorylation is associated with the microtubule protein Tau, which is an invariant feature of Alzheimer's disease and other neurodegenerative diseases, such as progressive supranuclear paralysis, corticobasal degeneration and disease Peak. Mutations in the Tau gene cause congenital forms frontotemporal dementia, additionally focusing on the value of the dysfunction of Tau protein for neurodegenerative process [Goedert, M. Curr. Opin. Gen. Dev., 2001, 11, 343].

Another aspect of the present invention relates to the use of compounds of the present invention or their pharmaceutically acceptable salts to obtain drugs for the treatment of bipolar disorder.

Another aspect of the present invention relates to the use of compounds of the present invention or their pharmaceutically acceptable salts to obtain drugs for the treatment of shock.

Reduced neuronal apoptosis is an important therapeutic goal in the context of head trauma, stroke, epilepsy and motor neuron disease [Mattson, M.P. Nat. Rev. Mol. Cell. Biol., 2000, 1, 120]. Therefore, GSK3 as the e proapoptotic factor in neural cells makes this protein kinase an attractive therapeutic target for inhibiting create medicines for the treatment of such diseases.

Another aspect of the present invention relates to the use of compounds of the present invention or its pharmaceutically acceptable salts to obtain drugs for the treatment of alopecia.

Hair growth is regulated by Wnt signaling by, in particular Wnt3. In model systems of the skin in the form of tissue culture expression not destroy mutants of β-catenin leads to a noticeable increase in the estimated population of stem cells that have extensive proliferative potential [Zhu, A.J.; Watt, F.M. Development, 1999, 126, 2285]. This population of stem cells expresses not associated with Katherina β-catenin at a higher level [DasGupta, R.; Fuchs, E. Development, 1999, 126, 4557], which may contribute to their high proliferative potential. In addition, transgenic mice with overexpression of a truncated β-catenin in the skin in a new way is the morphogenesis of the hair follicle, which is normally set only during embryogenesis. Ectopic application of GSK3 inhibitors may therefore be therapeutically useful in the treatment of alopecia and for hair regrowth after caused by chemotherapy alopecia.

An additional aspect of the present invention relates to a method of treatment of GSK3-dependent disease, where the method includes the administration to a patient in need this connection this is part II of the invention or its pharmaceutically acceptable salt, as defined above, in a quantity sufficient for inhibition of GSK3.

Preferably, the compound of the present invention or its pharmaceutically acceptable salt is administered in an amount sufficient for inhibition of GSK3.

In one of the embodiments of the compound of the present invention is administered in an amount sufficient for inhibiting at least one enzyme PLK.

Prepodobnye kinase (PLK) form a family of serine/treoninove protein kinases. Mitotic mutants of Drosophila melanogaster Polo locus show anomalies spindle [Sunkel et al., J. Cell Sci., 1988, 89, 25] and Polo, as found, encodes a mitotic kinase [Llamazares et al., Genes Dev., 1991, 5, 2153]. In humans there are three closely related PLK [Glover et al., Genes Dev., 1998, 12, 3777]. They contain highly homologous to aminocentesis catalytic kinase domain, and a carboxyl ends contain two or three conservative field, the Polo boxes. Function boxes Polo remains not fully understood, but they are involved in the direction of PLK in subcellular compartments [Lee et al., Proc. Natl. Acad. Sci. USA, 1998, 95, 9301; Leung et al., Nat. Struct. Biol., 2002, 9, 719], mediating interactions with other proteins [Kauselmann et al., EMBO J. 1999, 18, 5528] or can form part of the autoregulatory domain [Nigg, Curr. Opin. Cell Biol., 1998, 10, 776]. In addition, dependent on Boxing Polo PLK1 activity is required for proper transition the metaphase/anafa the a and cytokinesis [Yuan et al., Cancer Res., 2002, 62, 4186; Seong et al., J. Biol. Chem., 2002, 277, 32282].

Studies have shown that the human PLK regulate some fundamental aspects of mitosis [Lane et al., J. Cell. Biol., 1996, 135, 1701; Cogswell et al., Cell Growth Differ., 2000, 11, 615]. In particular, the activity of PLK1 is considered necessary for the functional maturation of centrosomes in late G2/early prophase and subsequent formation of the bipolar spindle. The decrease in cell PLK1 method of small interfering RNA (siRNA) also confirmed that this protein is required for many mitotic processes and the completion of cytokinesis [Liu et al., Proc. Natl. Acad. Sci. USA, 2002, 99, 8672].

In a more preferred embodiment of the present invention, the compound of the present invention is administered in an amount sufficient for inhibition of PLK1.

Of the three human PLK PLK1 is best characterized; it regulates some activities of the division cell cycle, including the beginning of mitosis [Toyoshima-Moritomo et al., Nature, 2001, 410, 215; Roshak et al., Cell Signaling, 2000, 12, 405], the activation of checkpoint DNA damage [Smits et al., Nat. Cell Biol., 2000, 2, 672; van Vugt et al., J. Biol Chem., 2001, 276, 41656], regulation of activating the anaphase complex [Sumara et al., Mol. Cell, 2002, 9, 515; Golan et al., J. Biol. Chem., 277, 15552; Kotani et al., Mol. Cell, 1998, 1, 371], the phosphorylation of the proteasome [Feng et al., Cell Growth Differ., 2001, 12 29] and the doubling and the maturation of the centrosome [Dai et al., Oncogene, 2002, 21, 6195].

In particular, the initiation of mitosis requires the activation of the call to the non M-phase factor (MPF), complex between cyclin-dependent kinase CDK1 and cyclename-type [Nurse, Nature, 1990, 344, 503]. Last accumulates during S and G2 phases of the cell cycle and causes the vast phosphorylation of MPF complex kinases WEE1, MIK1 and MYT1. At the end of the G2 phase corresponding dephosphorylation using phosphatase with dual specificity CDC25C triggers activation of MPF [Nigg, Nat. Rev. Mol. Cell Biol., 2001, 2, 21]. In the interphase cyclin-In is localized in the cytoplasm [Hagting et al., EMBO J. 1998, 17, 4127], then it becomes phosphorylated during prophesy, and this phenomenon causes nuclear translocation [Hagting et al., Curr. Biol., 1999, 9, 680; Yang et al., J. Biol. Chem., 2001, 276, 3604]. Nuclear accumulation of active MPF during profasi believed to be important for the initiation of events M-profasi [Takizawa et al., Curr. Opin. Cell Biol., 2000, 12, 658]. However, nuclear MPF remains inactive through WEE1, if it does not prevent CDC25C. Itself phosphatase CDC25C, localized in the cytoplasm during interphase, accumulates in the nucleus in prophase [Seki et al., Mol. Biol. Cell, 1992, 3, 1373; Heald et al., Cell, 1993, 74, 463; Dalal et al., Mol. Cell. Biol., 1999, 19, 4465]. Admission to the kernel as cyclina In [Toyoshima-Moritomo et al., Nature, 2001, 410, 215], and CDC25C [Toyoshima-Moritomo et al., EMBO Rep., 2002, 3, 341] is stimulated by phosphorylation by PLK1 [Roshak et al., Cell Signalling, 2000, 12, 405]. This kinase is an important regulator of initiation of M-phase.

In one particularly preferred embodiment, compounds of the present invention are ATP-antag the Communist inhibitors of PLK1.

In this context ATP antagonism refers to the ability of the vast compound to reduce or prevent the catalytic activity of PLK, i.e. posterino from ATP to the macromolecular substrate PLK, due to reversible or irreversible binding in the active site of the enzyme in such a way that attenuated or cancelled binding of ATP.

In another preferred embodiment, the compound of the present invention is used in a quantity sufficient to inhibit PLK2 and/or PLK3.

PLK2 (also known as SNK) and PLK3 (also known as PRK and FNK) mammals, as was originally shown, are direct early gene products. Kinase activity of PLK3, apparently, reaches its maximum during S phase and G2. It is activated during activation at a checkpoint, DNA damage and severe oxidative stress. PLK3 also plays an important role in the regulation of the dynamics of microtubules and functions of the centrosome in the cell, and misaligned expression of PLK3 leads to cell cycle arrest and apoptosis [Wang et al., Mol. Cell. Biol., 2002, 22, 3450]. PLK2 is at least understandable homologue three PLK. Both PLK2 and PLK3 may have additional important postmitotic functions [Kauselmann et al., EMBO J., 1999, 18, 5528].

Another aspect of the present invention relates to the use of compounds of the present izopet is ment for inhibiting protein kinases.

In a preferred embodiment, in this aspect, the protein kinase is a cyclin-dependent kinase. Preferably, the protein kinase is CDK1, CDK2, CDK3, CDK4, CDK6, CDK7, CDK8 or CDK9, more preferably, CDK2.

An additional aspect of the present invention relates to a method for inhibiting protein kinases, where the method includes contacting the specified protein kinase with the compound of the present invention.

In a preferred embodiment, in this aspect, the protein kinase is a cyclin-dependent kinase, more preferably, CDK2.

Tests

Another aspect of the present invention relates to the use of compounds as defined above in the analysis for identifying candidate compounds that affect the activity of one or more cyclin-dependent kinases, aurora kinases, GSK and/or PLK enzyme.

Preferably, the analysis allows the identification of candidate compounds capable of inhibiting one or more cyclin-dependent kinases, aurora kinases and GSK and/or PLK enzyme.

More preferably, the analysis is the analysis of competitive binding.

More preferably, the connection candidate receive conventional SAR modification of the compounds of the present invention.

As used herein, the term "conventional SAR modify the relative is carried out by standard methods, well-known experts in this field, to change the connection by chemical receiving its derivatives.

Thus, in one aspect of the identified compound can act as a model (e.g., matrix) to obtain other compounds. The compounds used in such a test may be free in solution, fixed to a solid substrate placed on the cell surface or located intracellularly. Elimination of activity or the formation of binding complexes between the joint and the tool that you want to experience, can be quantified.

Analysis according to the present invention can be selected, which have a number of funds. In one aspect of the method of analysis according to the present invention is a highly productive selection.

The present invention assumes the use of assays for screening drugs for competition, in which neutralizing antibodies capable of binding compounds specifically compete with a test compound for binding to the connection.

Another method of selection provides selection of high throughput (HTS) of the funds with the appropriate binding affinity of these substances, and is based on a method described in detail in WO 84/03564

Assumes that the analysis methods of the present invention will be useful in screening small volume and large-scale screening of test compounds, as well as quantitative analyses.

Preferably, the analysis of competitive binding includes contacting compounds of the present invention with cyclin-dependent kinase, aurora kinase, GSK or PLK enzyme in the presence of a known substrate of the indicated CDK enzyme and detecting any change in the interaction between a specified kinase and specified a known substrate.

An additional aspect of the present invention is a method of detecting binding of the ligand with cyclin-dependent kinase, aurora kinase, GSK or PLK enzyme, where the method involves the following stages:

(i) contacting a ligand with cyclin-dependent kinase, aurora kinase, GSK or PLK enzyme in the presence of a known substrate of the indicated kinase;

(ii) detecting any change in the interaction between a specified kinase and specified a known substrate;

and where specified ligand is a compound of the present invention.

One aspect of the present invention relates to a method, which includes stages:

(a) the method of analysis described above;

(b) determining one or more ligands the binding with a ligand-binding domain; and

(C) obtaining a quantity of the specified one or more ligands.

Another aspect of the present invention is a method including steps:

(a) the method of analysis described above;

(b) determining one or more ligands capable of binding with a ligand-binding domain; and

(C) obtaining a pharmaceutical composition containing the specified one or more ligands.

Another aspect of the present invention is a method including steps:

(a) the method of analysis described above;

(b) determining one or more ligands capable of binding with a ligand-binding domain; and

(C) modification of the specified one or more ligands capable of binding with a ligand-binding domain;

(d) the method of analysis described above;

(e) optionally, pharmaceutical compositions containing the specified one or more ligands.

The present invention also relates to a ligand identified by the method described above.

Another aspect of the present invention relates to pharmaceutical compositions containing a ligand identified by the method described above.

Another aspect of the present invention relates to the use of a ligand identified by the method, description of the authorized above, to obtain a pharmaceutical composition for use in the treatment of proliferative diseases.

The above methods can be used for selection of the ligand used as an inhibitor of one or more CDK enzymes.

The present invention is described below using the following examples and with reference to the following figure, where:

figure 1 shows the downward regulation of Mcl-1 compounds of the present invention. Cells In processed within 24 hours by different concentrations of compounds and analyzed by Western blotting for changes in the level of Mcl-1;

figure 2 shows some of the following compounds on the levels of Mcl-1 in cells N. Cells were treated with several concentrations of each drug and analyzed after 5 hours.

Examples

General information

Chemicals and solvents were purchased from commercial sources and used upon receipt, unless otherwise specified. THF and Et2O dried at boiling point under reflux with sodium benzophenone in an atmosphere of N2and separated by distillation. Toluene was dried at boiling temperature under reflux over sodium in the atmosphere of N2. CH2Cl2were dried at boiling temperature under reflux over San2in the atmosphere attaipendeake microwave generator was the model CEM Discover design in a single round cavity, for focusing microwave radiation on the tube for the sample. TLC (thin layer chromatography) was performed using glass plates coated with silica gel G60 (0.25 cm). The developed plates were air-dried and analyzed under a UV lamp (254/365 nm). Anhydrous MgSO4used as the standard method for drying organic solutions, unless otherwise indicated. Column flash chromatography was performed using silica gel Fluorochem (35-70 μm). The melting temperature (etc) were determined using capillary apparatus for determining the melting temperature Electrothermal 9100 and not corrected. Reduction (decomp.) means the decomposition temperature. Spectra1H-NMR were recorded on a spectrophotometer Bruker Avance 300 (300,1 MHz) or Varian Gemini 2000 (300 MHz) using the deuterated solvent as the lock and the residual solvent as an internal standard in all cases. Spectra13C-NMR using sequence PENDANT were recorded on a spectrophotometer Bruker Avance 300 (75.5 MHz). All other spectra13With were recorded on a spectrophotometer Varian Gemini 2000 (75.5 MHz), using complex pulse separation1N. The coupling constant (J) was estimated with an accuracy of 0.1 Hz. Used the following abbreviations: s - singlet; d - doublet; t - triplet; sq Quartet; Queen. quintet; m - multiplet and the Il. - wide. Elemental analyses were performed Mrs S. Williamson, Scholl of Chemistry, Purdie Building, University of St Andrews, UK. The obtained results were within 0.4% of calculated values. Mass spectra with elektrorazpredelenie (ESI) were recorded on a mass spectrometer Micromass LCT, in combination with HPLC Waters 2975. Analytical RP-HPLC was performed using an automatic sample injector Dionex ASI-100, connected to a pump Dionex P580. For analytical purposes used column Phenomenex (150×4,60 mm Synergi 4 ám hydro-RP 80 Å), sustained at 25°C. the Installation HPLC regulated using software Chromeleon. Linear gradient elution was performed using a systems N2O/MeCN (containing 0.1% CF3COOH) at a flow rate of 1 ml/min Purity was assessed by integration of the chromatogram (λ=254 nm).

Synthesis

(2R,3S)-3-Aminoindan-2-ol was obtained by one or other of two ways, different protecting group used for Amina.

Path 1 using trityl as a protective group

(S)-2-(trailmen)butane-1-ol

To a stirred solution of (S)-(+)-2-(trailmen)butane-1-ol (10 g, 112,18 mmol) in dichloromethane (DCM, 250 ml) in an argon atmosphere at room temperature was added diisopropylethylamine (DIEA, and 19.4 ml, 112,18 mmol), followed by the addition of Fritillaria (31,2 g, 112,18 mmol. The reaction mixture was stirred at this temperature for 48 hours, until TLC (hexane:ether:Meon; 55:40:5) showed that the reaction went to completion. The solvent is evaporatedin the vacuumand the residue was taken in ethyl acetate. The organic solution was washed with water (2×), dried over sodium sulfate. The solvent was removed, to obtain (S)-2-(trailmen)butane-1-ol as a pale yellow oil; yield: 33 g (89%).

1H-NMR (CDCl3, 250 MHz): δ to 0.72 (3H, t, J=7.5 Hz, -NHCH(CH2CH3)CH2OH), 1,15-1,10 (m, 2H, -NHCH(CH2CH3)CH2OH), was 2.05 (1H, s, of user., NH), 2,24 (1H, s, of user., OH), 2,62-of 2.54 (m, 1H, -NHCH(CH2CH3)CH2OH), 3,17-is 3.08 (1H, m, -NHCH(CH2CH3)CHHOH), 3,35-3,29 (1H, m, NHCH(CH2CH3)CHHOH), 7,37 to 7.2 (12H, m, ArH), 7,65-7,58 (3H, m, ArH); δC(250 MHz, CDCl3) 146,86 (C), 129,43 (6×CH), 127,90 (6×CH), 126,48 (3×CH), 71,27 (C), 62,72 (CH2), 48,91 (CH), RUB 24.55 (CH2), 10,47 (CH3).

(S)-2-(Trailmen)Butyraldehyde

To a stirred solution of dry DMSO (2.4 ml, 2.8 EQ., 33,82 mmol) in dry dichloromethane (30 ml) in an argon atmosphere at-78ºC was added dropwise oxalicacid (2M solution in DCM, to 8.45 ml, 1,40 EQ. of 16.9 mmol). The reaction mixture was stirred at-78º for 1 hour, then added dropwise with stirring was added (S)-2-(trailmen)butane-1-ol (4 g, 1 EQ, 12,07 mmol) in DCM (30 ml). The reaction mixture was stirred is ri this temperature for 2 hours, then the solution was added triethylamine (TEA, and 8.4 ml, 5 EQ., 60,27 mmol) in DCM and the solution was allowed to warm to room temperature for 1 hour. The reaction mixture was diluted with additional DCM (100 ml) and washed with water (250 ml). The aqueous phase was extracted with DCM (3×50 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (Na2SO4) and evaporatedin the vacuum. The residue was purified flash chromatography on silica (ethyl acetate:hexane 1:4), obtaining (S)-2-(trailmen)Butyraldehyde in the form of a light yellow oil; yield: of 3.64 g (91%).

1H-NMR (CDCl3, 250 MHz): δ of 0.95 (3H, t, J=7.5 Hz, -NHCH(CH2CH3)CHO), 1,72-1,52 [2H, m, NHCH(CH2CH3)CHO], was 2.76 (1H, s, of user., -NH), 3,41-3,36 [1H, m, NHCH(CH2CH3)CHO], 7,35-7,17 (12H, m, ArH), to 7.67-7,51 (3H, m, ArH), 9,05 (1H, s, NHCH(CH2CH3)CHO). δC(250 MHz, CDCl3) 202,95 (CO), 146,23 (C), 129,23 (6×CH), 127,96 (6×CH), 126,85 (3×CH), 71,13 (C), 62,62 (CH), 24,78 (CH2), 10,48 (CH3).

(2R,3S)-3-(Trailmen)pentane-2-ol

To a stirred suspension of CuBr·SMe2(3 g, 14.6 mmol) in anhydrous ether (100 ml) in an argon atmosphere at-78º was added dropwise motility (1.6 m in ether, 16,5 ml, 4.0 EQ., of 26.5 mmol)and the solution was allowed to warm to room temperature over 1 hour. The mixture was again cooled to-78º and was added dropwise a solution of (S)-2-(trailmen)Butyraldehyde (2.2 g, 6,62 m is ol) in ether (25 ml) under stirring. The reaction mixture was stirred at this temperature for 2 hours, then was allowed to warm to room temperature over 1 hour. Was added a saturated aqueous solution of NH4Cl (50 ml) and the two layers were separated. The organic phase is washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified column flash chromatography on silica gel with elution by the mixture hexane:ethyl acetate (80:20), to obtain (2R,3S)-3-(trailmen)pentane-2-ol as a pale yellow oil; yield: 1.5 g (66%). (75% de 2R,3S: 25% de 2S,3S).

1H-NMR (d6-DMSO, 250 MHz): δ 0, 0,47+0,55 (2×t, J=7,50+7,26 Hz,- NHCH(CH2CH3)CH(CH3)OH), 0,99 by 1.12 (m, 5H, -NHCH(CH2CH3CH) (CH3)OH), a 2.01 (1H, m, -NHCH(CH2CH3)CH(CH3)OH), 3,22-of 3.43 (m, 1H, -NHCH(CH2CH3)CH(CH3)OH), to 4.41 [1H, d, J=3.3,thenNHCH(CH2CH3)CH(CH3)OH], 7,14-7,56 (15H, m, ArH). δC(250 MHz, CDCl3) 146,88 (C), 128,97 (6×CH), 127,83 (6×CH), 126,43 (3×CH), 71,03 (C), 68,13 (CH), 58,77 (CH), 23,09 (CH2), 17,88 (CH3), 10,47 (CH3).

(2R,3S)-3-Aminoindan-2-ol

To a stirred solution of (2R,3S)-3-(trailmen)pentane-2-ol (1,64, of 4.75 mmol) in dichloromethane (20 ml) in an argon atmosphere at room temperature was added dropwise triperoxonane acid (10 ml) and the solution was stirred at this temperature for 1 hour. The solvent vapour is Wali in the vacuumand the residue was besieged from ether (15 ml), hexane (150 ml) under stirring, to obtain a yellow oil. The solvent decantation of the oil, the oil was washed with hexane (30 ml) and driedin the vacuumwith the receipt of (2R,3S)-3-aminoindan-2-ol as a pale yellow oil; yield: 0,30 g (98%). (75% de 2R,3S: 25% de 2S,3S).

1H-NMR (d6-DMSO, 250 MHz): δ 0,913+0,923 (2×t, 3H, J=7,50+7,50 Hz, NH2CH(CH2CH3)CH(CH3)OH), 1,11+1,18 (3H, 2×d, J=6,48+6,48 Hz, NH2CH(CH2CH3CH) (CH3)OH), 1.41 to of 1.65 (2H, m, NH2CH(CH2CH3)CH(CH3)OH), was 2.76+2,93 [2×1H, m, NH2CH(CH2CH3)CH(CH3)OH], 3,61-3,69+3,80-3,90 [2×1H, m, NH2CH(CH2CH3)CH(CH3)OH], 7,73 (2H, s, of user.,NH2).

When path 2 Amin defended by dibenzylamine.

(S)-2-(Dibenzylamino)butane-1-ol

To a stirred solution of (S)-(+)-2-aminobutane-1-ol (5 g, 56,18 mmol) in dry acetonitrile (100 ml) was added dry powdered potassium carbonate (31 g, 224,72 mmol), followed by the addition of benzylbromide (19 g, 111,11 mmol). The reaction mixture was stirred at room temperature for 24 hours. The solvent was removed in vacuum and the residue was taken in ethyl acetate (100 ml) and water (100 ml). The organic phase is again washed with water, dried (Na2SO4) and concentrated, to obtain the pure product as pale as the addition of oil (14.5 g, 97,3%).

δH(250 MHz, CDCl3) and 0.98 (3H, t, J=7,5, CHCH2CH3), 1,38-1,2 (1H, m, CHCHHCH3), 1,94-of 1.78 (1H, m, CHHCH3), 2,83-a 2.71 (1H, m, CHCHHCH3), up 3.22 (1H, s, of user., OH), 3,65-3,4 (2H, m, CH2OH), 3,47 (2H, d, J=17,5, 2×CHHPh), of 3.94 (2H, d, J=17,5, 2×CHHPh), 7,46-7,26 (10H, m, 2×C6H5); δC(250 MHz, CDCl3) 139,42 (2×C), 129,1 (2×CH), 128,52 (2×CH), 127,25 (2×CH), 61,97 (CH), 60,67 (CH2), 53,23 (CH2), 17,92 (CH2), 11,83 (CH3); m/z 270,2 (M+H).

(S)-2-(Dibenzylamino)butanal

2M solution of oxalicacid in dichloromethane (3,18 ml, 6,36 mmol) was cooled to -78°C and was diluted with dry dichloromethane (20 ml) in a dry nitrogen atmosphere. To a cooled stirred solution was added dropwise a solution of dimethylsulfoxide (1 g, 12,72 mmol) in anhydrous dichloromethane. The reaction mixture was stirred for a further 1 hour after the addition. Solution was added (S)-2-(dibenzylamino)butane-1-ol (1,43 g, 5.3 mmol) in dichloromethane for 5 minutes. After 10 minutes, was added diisopropylethylamine (2,73 g of 21.2 mmol). The reaction mixture was allowed to warm to room temperature and left to mix for 1 hour. It was cooled to 0°C was added a mixture of ethyl acetate/water (50 ml:50 ml). The organic layer was washed with water (50 ml), saturated salt solution (50 ml), dried (MgSO4) and concentrated. The product was purified flash column-chromatography utilized:hexane 1:4), to obtain the pure product (1.28 g, 90.5 per cent).

δH(250 MHz, CDCl3) to 0.88 (3H, t, J=7,5, CHCH2CH3), 1,77-and 1.54 (2H, m, CH2CH3), to 2.99 (1H, t, J=7,5, CHCH2CH3), 3,74 is 3.57 (4H, m, 2×CH2Ph), 7,31-7,11 (10H, m, 2×C6H5) for 9.64 (1H, s, CHO); δC(250 MHz, CDCl3) the amount of 203.9 (CO), 139,33 (2×C), 128,99 (4×CH), 128,45 (4×CH), RUB 127.3 (2×CH), 68,46 (CH), 54,85 (CH2), 17,44 (CH2), 11,83 (CH3); m/z 268,2 (M+H).

(2R,3S)-2-(Dibenzylamino)butane-2-ol

To a stirred suspension of CuBr·SMe2(1.54 g, 7.5 mmol) in anhydrous ether (100 ml) in an argon atmosphere at-78º was added dropwise motility (1.6 m in ether, 9.4 ml, 15 mmol). After the addition was completed, the reaction mixture was allowed to warm to room temperature. The reaction mixture was again cooled to -78°C. and was added dropwise a solution of (S)-2-(dibenzylamino)butanal (1 g, 3.75 mmol) in ether (20 ml). After the addition stirring was continued for 2 hours. Then the reaction mixture was extinguished saturated aqueous NH4Cl (10 ml). The reaction mixture was extracted with ether (2×30 ml) and the combined organic phase was washed with saturated salt solution (20 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified column gradient flash chromatography on silica gel with elution by the mixture hexane:ethyl acetate(100:0→80:20), with the receipt of the product in the form of velo yellow oil (0.95 g, 89%) as a single isomer.

δH(250 MHz, CDCl3) of 1.05 (3H, t, J=7,5, CHCH2CH3), 1,25 [3H, d, J=7,5, CH(CH3)OH], 1,6-1,49 (1H, m, CHHCH3), a 1.88-of 1.73 (1H, m, CHHCH3), is 2.41 (1H, s, of user., OH), 2,66 at 2.59 (1H, m, CHCH2CH3), 3,85-the 3.65 (4H, m, 2×CH2Ph), 4,05 to-3.9 (1H, m, CHOH), 7,41-of 7.25 (10H, m, ArH). δC(250 MHz, CDCl3) 140,05 (2×C), 128,98 (4×CH), 128,37 (4×CH), RUB 127.3 (2×CH), 66,81 (CH), 63,65 (CH), 55,41 (CH2), 20,63 (CH3), 18,44 (CH2), 12.5cm (CH3).

Example 1

2-chloro-4,6-diethylnicotinamide

4,6-Dimethyl-2-oxo-1,2-dihydropyridines-3-carbonitrile (5 g, 34 mmol) was added to phosphorus oxychloride (20 ml). The reaction mixture was stirred while boiling under reflux for 2 hours, and then observed that it ended. Volatiles were removed and the residue triturated with petrol. The obtained solid was filtered, washed with hexane and dried, to obtain a pure white solid (5.1 g, 90%).

δH(250 MHz, CDCl3) to 2.55 (3H, s, CH3), to 2.57 (3H, s, CH3), to 7.09 (1H, s, ArH); δWith(250 MHz, CDCl3) 162,64 (C), 154,39 (C), 152,26 (C), 123,22 (CH), 114,28 (C), 108,31 (C)24,5 (CH3), 20,54 (CH3); m/z 189 (M+Na).

Tert-butyl ether 4,6-dimethylpyridin-3-iletilerinindeki acid

2-Chloro-4,6-diethylnicotinamide (5 g, to 30.1 mmol) was dissolved in a mixture of 10% acetic who iSlate/ethanol (30 ml). Added catalyst 10% palladium on carbon (0.5 g) and the reaction mixture was stirred in an atmosphere of hydrogen for 24 hours at 60°C. the Mixture was filtered through a loose layer of celite. Volatiles were removed and the crude residue was dissolved in dichloromethane (30 ml). To the stirred solution was then added triethylamine (5 ml), followed by addition of di-tert-BUTYLCARBAMATE (6.5 g, 30 mmol). After 3 hours the solvent was removed and the residue was dissolved in ethyl acetate. The resulting solution was washed with water (50 ml), saturated bicarbonate solution (50 ml), dried and evaporated. The crude product was purified column flash chromatography on silica gel (ethyl acetate:hexane 1:2), to obtain 1.4 g of pure indicated in the title compound (20% yield).

δH(250 MHz, CDCl3) was 1.43 (9H, s, 3×CH3), are 2.19 (3H, s, CH3), of 2.38 (3H, s, CH3), 4,19 (2H, s, of user., ArCH2NH), at 6.84 (1H, s, ArH), of 8.15 (1H, s, ArH); δC(250 MHz, CDCl3) 157,41 (CO), 155,63 (C), 148,93 (CH), 145,91 (C), 129,51 (C), 124,76 (CH), 79,44 (C), 46,12 (CH2), weighing 28.32 (3×CH3), 23,74 (CH3), 18,97 (CH3); m/z of 237.2 (M+H).

(4,6-Dimethylpyridin-3-ylmethyl)-(2-fluoro-N-purine-6-yl)Amin

To a stirred solution of 6-chloro-2-porporino (0,83 g, 4.9 mmol) in n-BuOH (50 ml) in an argon atmosphere at 0°C was added DIEA (2.5 ml, 14.7 mmol), followed by addition of (4,6-dimethylpyridin-3-yl)methanamine (1 g, of 7.35 mmol). Re clonney the mixture was stirred at this temperature for 1 hour, then was allowed to return to room temperature and was stirred for 4 hours, it was observed that the reaction is not yet completed, and therefore the reaction mixture was heated to 100°C and left at this temperature for 2 hours. The solvent is evaporatedin the vacuumand the residue was purified gradient column flash chromatography on silica gel with elution CHCl3:MeOH(100:0→90:10), obtaining the product as a white solid; yield: 0,86 g (65%).

δH(250 MHz, CDCl3) to 2.35 (3H, s, CH3), 2,39 (3H, s, CH3), br4.61 (2H, s, of user., NHCH2), 7,07 (1H, s, ArH), 8,13 (1H, s, ArH), with 8.33 (1H, s, ArH), 8,69 (1H, s, of user., NH); δc (250 MHz, CDCl3) 161,2 (C), 158,57 (C), 156,08 (C), 150 (C), 148,08 (CH), 148,14 (CH), 147,9 (CH), 145,93 (C), 129,92 (C), 129,76 (C), 124,37 (CH), and 41.7 (CH2), 23,17 (CH3), 18,14 (CH3); m/z 273,2 (M+H).

(4,6-Dimethylpyridin-3-ylmethyl)-(2-fluoro-9-isopropyl-N-purine-6-yl)Amin

To a stirred solution of (4,6-dimethylpyridin-3-ylmethyl)-(2-fluoro-N-purine-6-yl)amine (0.6 g, 1.9 mmol) in DMF (10 ml) in an argon atmosphere while K.T. was added K2CO3(powder, anhydrous, or 1.77 g, 5 EQ., 13 mmol), followed by the addition of 2-bromopropane (1.8 ml, 10 EQ., 19 mmol). The reaction mixture was stirred at K.T. within 24 hours, until TLC (CHCl3:MeOH; 90:10) showed that the reaction went to completion. The solvent is evaporatedin the vacuumand the residue was distributed settled between water (50 ml) and ethyl acetate (50 ml), the aqueous phase was separated and additionally was extracted with EtOAc (2×50 ml). The combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4), evaporatedin the vacuumand the residue was purified gradient column chromatography on silica gel with elution CHCl3:MeOH(100:1→95:5), obtaining the product as a yellow film (0.4 g, 59%).

δH(250 MHz, CDCl3) 1,52 [6H, d, J=7,5 CHCCH3)2], and 2.27 (3H, s, CH3), a 2.45 (3H, s, CH3), 4,73-to 4.62 (3H, m, NHCH2and CH[CH3]2), 6,91 (1H, s, ArH), 7,12 (1H, NH), 7,47 (1H, s, ArH), 8,32 (1H, s, ArH); δC(250 MHz, CDCl3) 160,77 (C), 157,89 (C), 157,43 (C), 156,12 (C), 155,79 (C), 149,14 (CH), 137,7 (CH), 128,7 (C), 129,76 (C), 124,83 (CH), 47,2 (CH), 40,14 (CH2), 23,9 (CH3), 22,47 (2×CH3), 18,54 (CH3); m/z 315,3 (M+H).

(2R,3S-3-(6-((4,6-dimethylpyridin-3-ylmethylamino)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol [1]

To a stirred solution of (4,6-dimethylpyridin-3-ylmethyl)-(2-fluoro-9-isopropyl-N-purine-6-yl)amine (300 mg, 0.84 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (1.7 ml, 10 EQ., 8.4 mmol), followed by addition of (2R,3S)-3-aminoindan-2-ol (0.5 g, 4.8 mmol). The flask was supplied with the condenser and the reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 72 hours. The reaction mixture gave axladitsa room temperature and the solvent evaporated in the vacuum. The residue was distributed between ethyl acetate (50 ml) and water (50 ml), the aqueous phase was extracted with EtOAc (2×25 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified gradient column chromatography on silica gel with elution CHCl3:MeOH(100:1→95:5), to obtain 55 mg of pure compound indicated in heading (12%).

δH(250 MHz, CDCl3) of 0.95 (3H, t, J=7,5, CHCH2CH3), was 1.06 (3H, d, J=7,5, CHCH3OH), 1,48 [6H, d, J=7,5 CHCCH3)2], of 2.24 (3H, s, CH3), 2,4 (3H, s, CH3), 3,92-3,82 (2H, m, NHCH2), 4,67 is 4.45 (3H, m, CHEtCHMeOH), x 6.15 (1H, s, of user., NH), 6.87 in (1H, s, ArH), 7,37 (1H, ArH), 8,31 (1H, s, ArH); δC(250 MHz, CDCl3) 160,11 (C), 157,68 (C), 154,57 (C), 149,42(CH), 146,38 (C), 134,54 (CH), 129,24 (C), 124,84 (CH), 71,52 (CH), 59,65(CH), 46,47 (CH), 40,33 (CH2), 24,94 (CH2), 23,89 (CH3), 23,52 (2×CH3), 17,37 (CH3), 12,57 (CH3); m/z 398,3 (M+H).

Example 2

2-Fluoro-N-((6-methylpyridin-3-yl)methyl)-N-purine-6-amine

To a stirred solution of 6-chloro-2-porporino (0.4 g, 2.3 mmol) in n-BuOH (50 ml) in an argon atmosphere at 0°C was added DIEA (2.5 ml, 14.7 mmol), followed by addition of (6-methylpyridin-3-yl)methylamine (0.36 g, 2,95 mmol). The reaction mixture was stirred at this temperature for 1 hour, then was allowed to return to room temperature and was stirred in ECENA 4 hours, observed that the reaction was still not complete, so the reaction mixture was heated to 100°C and left at this temperature for 8 hours. The solvent is evaporatedin the vacuumand the residue was purified gradient column chromatography on silica gel with elution CHCl3:MeOH(100:0→90:10), with obtaining the product as a white solid; yield: 0,38 g (65%).

δHCDCl3, 250 MHz) of 2.44 (3H, s, CH3), 3,66 is 3.57 (2H, m, NHCH2), 4,63 (1H, s, of user., NH), 7,25 (1H, d, J=7,5, ArH), 7,71 (1H, DD, J=2,5, And 7.5, ArH), to 8.14 (1H, s, ArH), 8,49 (1H, s, ArH), 9,07 (1H, s, of user., NH); δWith(CDCl3, 250 MHz) 159,12 (C), 158,62 (C), 157,61 (C), 155,56 (C), 147,44 (CH), 146,99 (CH), 136,32 (C), 123,05 (2×CH), 119,42 (C), 41,64 (CH2), 18,47 (CH3); m/z 259,2 (M+H).

2-Fluoro-9-isopropyl-N-((6-methylpyridin-3-yl)methyl)-N-purine-6 - amine

To a stirred solution of 2-fluoro-N-(6-methylpyridin-3-yl)methyl-N-purine-6-amine (0.3 g, at 1.17 mmol) in dimethylformamide (10 ml) at room temperature in an argon atmosphere was added anhydrous powder To2CO3(0.8 g, 5 EQ., to 5.85 mmol), followed by the addition of 2-bromopropane (1,15 ml, 11.7 mmol). The reaction mixture was stirred at room temperature for 24 hours, until the mixture DCM:ether:Meon (55:40:5) showed that the reaction went to completion. The solvent is evaporatedin the vacuumand the residue was distributed between EtOAc (100 ml) and water (100 ml). The aqueous phase will add the flax was extracted with EtOAc (2×50 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified column flash chromatography on silica gel with elution CHCl3:MeOH (98:2), obtaining specified in the title compound as a slightly yellow film (195 mg, 55%).

δH(CDCl3, 250 MHz) of 1.52 (6H, d, J=7,5, CH[CH3]2), 2,5 (3H, s, CH3), 4,76-4,6 (3H, m, NHCH2and CHMe2), 7,06 (1H, d, J=2,5, ArH), 7,35 (1H, s, of user., NH), 7,56 (2H, s, of user., ArH), of 8.47 (1H, s, of user., ArH); δC(CDCl3, 250 MHz) 157,6 (C), 156,32 (C)156 (C), 148,47 (CH), 137,72 (CH), 136,08 (CH), 130,83 (C), 123,11 (CH), 118,2 (C), 47,38 (CH), 43,2 (CH2), 23,99 (CH3), 22,5 (2×CH3); m/z 301,2 (M+H).

2R,3S-3-(9-isopropyl-6-((6-methylpyridin-3-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol [2]

To a stirred solution of 2-fluoro-9-isopropyl-N-(6-methylpyridin-3-ylmethyl)-N-purine-6-amine (180 mg, 0.59 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (1 ml, 10 EQ., 5.6 mmol), followed by addition of (2R,3S)-3-aminoindan-2-ol (0.34 g, 6 mmol). The flask was supplied with the condenser and the reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 72 hours. The reaction mixture was allowed to cool to room temperature and the solvent evaporatedin the vacuum. The residue was distributed between EtOAc (50 ml) and water (50 ml), the aqueous phase is additionally extraheavy and EtOAc (2×25 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified gradient column flash chromatography on silica gel with elution CHCl3:MeOH(100:1→95:5), obtaining specified in the title compound (40 mg, 19%).

δH(CDCl3, 250 MHz) of 0.95 (3H, t, J=7,5, CHCH2CH3), to 1.14 (3H, d, J=5, CHCH3OH), 154 (6H, d, J=7,5, CH[CH3]2), 1,62 was 1.43 (2H, m, CHCH2CH3), is 2.44 (3H, s, ArCH3), 3,93 (1H, m, CHMe2), 4,77-4,58 (1H, m, CHCH3OH), 4,8-4,6 (2H, m, NHCH2Ar), and 5.8 (1H, s, of user., NH), PC 6.82 (1H, s, of user., NH), to 7.09 (1H, d, J=10, ArH), 7,31-of 7.23 (2H, m, ArH), 8,49 (1H, s, of user., ArH); δC(CDCl3, 250 MHz) 157,71 (C), 156,28 (C), 155,95 (C), 148,58 (CH), 137,73 (CH), 129,01 (C), 128,52 (C), 128,42 (CH), 1231,14 (CH), 68,84 (CH), 50,45 (CH2), 47,25 (CH), 23,27 (CH3), 22,53 (2×CH3), to 20.9 (CH2), 19,46 (CH3), 10,45 (CH3); m/z 384,3 (M+H).

Example 3

(3 Chlorbenzyl)-(2-fluoro-N-purine-6-yl)Amin

To a stirred solution of 6-chloro-2-porporino (1 g, 1 EQ., 5.9 mmol) in n-BuOH (50 ml) in an argon atmosphere was added DIEA (2.6 ml, 2.5 EQ., 14,75 mmol), followed by addition of 3-chlorobenzylamino (1,25 g, 1.5 EQ., cent to 8.85 mmol). The reaction mixture was heated to 100°C and kept at this temperature for 3 hours, then the reaction was completed. The solvent is evaporatedin the vacuumand the residue was purified gradient column chromatography on silica gel with elution with a mixture of DCM:ether:EON(55:45:0→55:43:2), obtaining specified in the title compound as a white solid; yield: 1,15 g (70%).

δH(250 MHz, d6-DMSO) 4,78-4,6 (2H, m, NHCH2Ar), 7,41-7,28 (4H, m, ArH), EUR 7.57 (1H, s, of user., ArH), of 8.15 (1H, s, of user., NH), 8,88 (1H, s, of user., NH); δWith(250 MHz, d6-DMSO) 142,3 (C), 133,02 (C), 132,92 (C), 130,25 (CH), 130,17 (C), 127,82 (CH), 127,27 (CH), 127,03 (CH), 126,76 (C), 125,92 (CH), 116,4 (C), 115 (C), 42,67(CH2); m/z 278 (M+H).

(3 Chlorbenzyl)-(2-fluoro-9-isopropyl-N-purine-6-yl)Amin

To a stirred solution of (3-Chlorobenzyl)-(2-fluoro-N-purine-6-yl)amine (0.6 g, of 2.16 mmol) in dimethylformamide (10 ml) at room temperature in an argon atmosphere was added powdered anhydrous potassium carbonate (1.47 g, 5 EQ., the 10.8 mmol), followed by the addition of 2-bromopropane (2.2 ml, 10 EQ., 21.6 mmol). The reaction mixture was stirred at room temperature for 24 hours, while elution with a mixture of DCM:ether:Meon (55:40:5) showed that the reaction went to completion. The solvent is evaporatedin the vacuumand the residue was distributed between ethyl acetate (50 ml) and water (50 ml). The aqueous phase was additionally extracted with ethyl acetate (2×50 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified column flash chromatography on silica gel with elution CHCl3with obtaining specified in the header is soedineniya in the form of a slightly yellow resin: yield 0.45 g (65%).

δH(250 MHz, CDCl3) of 1.57 (6H, d, J=7,5, CH[CH3]2), 4,81-4,63 (3H, m, NHCH2Ar and CHMe2), 5,98 (1H, s, of user., NH), 7,29-7,19 (3H, m, ArH), 7,34 (1H, s, of user., ArH), 7,42 (1H, d, J=7,5, ArH), 7,51 (1H, s, of user., NH); δC(250 MHz, CDCl3) 161,93 (C), 156,37 (C), 156,05 (C), 141,33 (C), 140,42 (C), 137,75 (CH), 134,49 (C), 129,92 (CH), 127,69 (CH), 125,81 (CH), 125,64 (CH), 47,43 (CH), 43,82 (CH2), 22,56 (2×CH3); m/z 320,3 (M+H).

2R,3S-3-(6-(3-Chlorobenzylamino-9-isopropyl-N-purine-2-ylamino)pentane-2-ol [3]

To a stirred solution of (3-Chlorobenzyl)-(2-fluoro-9-isopropyl-N-purine-6-yl)amine (0.3 g, of 0.93 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (0.6 ml, 10 EQ., 10 mmol), followed by addition of (2R,3S)-3-aminoindan-2-ol (0.5 g, 4.8 mmol). The reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 72 hours, until TLC with a mixture of chloroform:methanol (95:5) showed that the reaction went to completion. The reaction mixture was allowed to cool to room temperature and the solvent evaporatedin the vacuum. The residue was distributed between dichloromethane (100 ml) and water (100 ml), the aqueous phase was additionally extracted with dichloromethane (3×50 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified gradient column the lash-by chromatography on silica gel with elution with a mixture of DCM:ether:MeOH(60:40:0→60:40:2), obtaining specified in the title compounds as a colorless film; yield 90 mg (22.5 percent).

δH(250 MHz, CDCl3) to 0.97 (3H, t, J=7,5, CHCH2CH3), a 1.08 (3H, d, J=7,5, CHCH3OH), a 1.45 (6H, d, J=7,5, CH[CH3]2), 1,6-of 1.35 (2H, m, CHCH2CH3), 3,93-3,8 (2H, m, CHEt and NH), 4,55-4,45 [1H, m, CH(CH3)OH], 4,74 with 4.64 (3H, m, NHCH2Ar and CHMe2), and 5.5 (1H, s, of user., OH), 6,53 (1H, s, of user., NH), 7,19-7,13 (3H, m, ArH), 7,22 (1H, s, ArH), 7,32 (1H, s, ArH); δC(250 MHz, CDCl3) 175,06 (C), 160,17 (C), 154,75 (C), 141,22 (C), 134,57 (CH), 134,29 (C), 129,73 (CH), 127,75 (CH), 127,30 (CH), 125,7 (CH), 114,56 (C), 71,53 (CH), 59,58 (CH), 46,44(CH), 43,78 (CH2), 23,96 (2×CH3), 18,65 (CH3), 18,32 (CH3), 12,56 (CH3); m/z 403,2 (M+H).

Example 4

(3-Terbisil-2-fluoro-N-purine-6-yl)Amin

To a stirred solution of 6-chloro-2-porporino (1 g, 5.9 mmol) in n-BuOH (30 ml) in an argon atmosphere was added DIEA (2.6 ml, 2.5 EQ., 14,75 mmol), followed by addition of 3 - carbencillin (1.1 g, 1.5 EQ., cent to 8.85 mmol). The reaction mixture was heated to 100°C and kept at this temperature for 3 hours, then the reaction was completed. The solvent is evaporatedin the vacuumand the residue was purified gradient column flash chromatography on silica gel with elution with a mixture of DCM:ether:Meon(55:45:0→55:43:2), obtaining specified in the title compound as a white solid; yield: 1.1 g (71,7%).

δH(250 MHz, d6 -DMSO) 4,79-of 4.66 (2H, s, of user., NHCH2Ar), 7,41-7,28 (4H, m, ArH), EUR 7.57 (1H, s, of user., ArH), of 8.15 (1H, s, of user., NH), 8,87 (1H, s, of user., NH); δC(250 MHz, d6-DMSO) 142,5 (C), 133,02 (C), 132,92 (C), 130,24 (CH), 130,17 (C), 128,41 (CH), 127,75 (C), 127,04 (CH), 126,76 (CH), 125,92 (CH), to 116.2 (C), 115 (C), 42,68 (CH2); m/z 262,1 (M+H).

(3-Terbisil-(2-fluoro-9-isopropyl-N-purine-6-yl)Amin

To a stirred solution of (3-terbisil)-(2-fluoro-N-purine-6-yl)amine (0.6 g, 2,24 mmol) in dimethylformamide (10 ml) at room temperature in an argon atmosphere was added powdered anhydrous potassium carbonate (1.52 g, 5 EQ., of 11.2 mmol), followed by the addition of 2-bromopropane (2.3 ml, 10 EQ., of 22.4 mmol). The reaction mixture was stirred at room temperature for 24 hours until TLC with a mixture of DCM:ether:Meon (55:40:5) showed that the reaction went to completion. The solvent is evaporatedin the vacuumand the residue was distributed between ethyl acetate (100 ml) and water (100 ml). The aqueous phase was additionally extracted with ethyl acetate (2×50 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified column flash chromatography on silica gel with elution by chloroform, obtaining specified in the title compound as a slightly yellow resin: the yield of 0.37 g (54%).

δH(250 MHz, CDCl3) of 1.55 (6H, d, J=7,5, CH[CH3]2), 4,85 to 4.5 (3H, m, NHCH2Ar and CHMe2), 5,96 (1H, s, of user., NH), 7,31-6,94 (4H, m, ArH), to 7.59 (1H, s, of user., ArH); δC(250 MHz, CDCl3) 164,93 (C), 161,01 (C), 156,04 (C), 139,77 (C), 137,76 (CH), 130,13 (CH), 129,92 (C), 123,19 (CH), 114,59 (CH), 114,26 (CH), 47,22 (CH), 43,93 (CH2), 22,56 (2×CH3); m/z 304,2 (M+H).

2R,3S-3-[6-(3-Forbindelsen)-9-isopropyl-N-purine-2-ylamino]pentane-2-ol [4]

To a stirred solution of (3-terbisil)-(2-fluoro-9-isopropyl-N-purine-6-yl)amine (0.3 g, 0,99 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (0.6 ml, 10 EQ., 10 mmol), followed by addition of (2R,3S)-3-aminoindan-2-ol (0.5 g, 4.8 mmol). The reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 72 hours, until TLC with a mixture of chloroform:methanol (95:5) showed that the reaction went to completion. The reaction mixture was allowed to cool to room temperature and the solvent evaporatedin the vacuum. The residue was distributed between dichloromethane (100 ml) and water (200 ml), the aqueous phase was additionally extracted with dichloromethane (3×25 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified gradient column flash chromatography on silica gel with elution with a mixture of DCM:ether:MeOH(60:40:0→60:40:2), obtaining specified in C is the head of product as a colorless film; yield: 80 mg (20%).

δH(250 MHz, CDCl3) of 1.05 (3H, t, J=7,5, CHCH2CH3)and 1.15 (3H, d, J=7,5, CHCH3OH), of 1.55 (6H, d, J=7,5, CH[CH3]2), 1,65-1,4 (2H, m, CHCH2CH3), 4-3,91 (2H, m, CHEt and CHMe2), 4,66-4,55 (1H, m, CHMeOH), and 4.8 (2H, d, J=5, NHCH2Ar)6,41 (1H, s, of user., NH), 7,33-6,92 (4H, m, ArH), 7,46 (1H, s, of user., ArH); δC(250 MHz, CDCl3) 165,2 (C), 164,92 (C), 160,18 (C), 154,77 (C), 141,67 (C), 141,56 (C), 134,59 (CH), 130,08 (CH), 123,14 (CH), 114,71 (CH), 113,95 (CH), 71,61 (CH), 59,65 (CH), 46,48 (CH), 43,94 (CH2), 25,02 (CH2), 22,52 (2×CH3), 17,24 (CH3), 10,59 (CH3); m/z 387,3 (M+H).

Example 5

(9-Cyclopropylmethyl-2-fluoro-N-purine-6-yl)pyridine-3-ylmethylamino

To a stirred solution of 2-fluoro-6-[(pyridine-3-ylmethyl)amino]purine (1 g, 4,10 mmol) in DMF (12 ml) in an argon atmosphere while K.T. was added To the2CO3(anhydrous powder, 2,84 g, 5 EQ., to 20.52 mmol), followed by the addition (bromacil)cyclopropane (of 5.53 ml, 10 EQ., 41 mmol). The reaction mixture was stirred at K.T. within 24 hours, until TLC (CHCl3:MeOH; 90:10) showed that the reaction went to completion. The solvent is evaporatedin the vacuumand the residue was distributed between water (50 ml) and EtOAc (50 ml); the aqueous phase was separated and additionally was extracted with EtOAc (2×50 ml). The combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuumand the residue was purified gradient column flash chromatography on silica gel with elution CHCl 3:MeOH(100:0→95:5), obtaining the product as a colourless resin; yield: 0.8 g (68%).

δH(250 MHz, CDCl3) of 0.24 to 0.15 (2H, m, CHH and CHH from Cp), 0,48 is 0.38 (2H, m, CHH and CHHfrom Cp), 1,11-of 0.95 (1H, m, CHfrom Cp), of 2.56 (1H, s, of user., NH), of 3.69 (2H, d, J=7,5, NCH2Cp), 4,58 (2H, s, of user., NHCH2Ar),? 7.04 baby mortality-of 6.96 (1H, m, ArH), 7,51 was 7.45 (2H, m, ArH), compared to 8.26-8,24 (1H, m, ArH), of 8.37 (1H, s, of user., ArH); δC(250 MHz, CDCl3) 156,32 (C), 156,01 (C), 149,29 (CH), 148,97 (CH), 139,85 (CH), 139,8 (C), 135,54 (CH), 133,81 (C), 123,51 (CH), 117,86 (C), 48,52 (CH2), 42,09 (CH2), 11,06 (CH3), 5,28 (2×CH2); m/z 299,2 (M+H).

2R,3S-3-(9-(Cyclopropylmethyl)-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol [5]

To a stirred solution of (9-cyclopropylmethyl-2-fluoro-N-purine-6-yl)pyridine-3-ylmethylamino (300 mg, 1 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (1.9 ml, 10 EQ., 10.5 mmol), followed by addition of (2R,3S)-3-aminoindan-2-ol (500 mg, 4.8 equiv., 4.8 mmol). The flask was supplied reflux, the reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 72 hours. The reaction mixture was allowed to cool to room temperature and the solvent evaporatedin the vacuum. The residue was distributed between ethyl acetate (50 ml) and water (50 ml), the aqueous phase was additionally extracted with ethyl acetate (2×25 ml) and the combined organic the definition phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified gradient column flash chromatography on silica gel with elution CHCl3:MeOH(100:0→95:5), obtaining specified in the header of the product as a colorless film. Output (85 mg, 22%).

δH(250 MHz, CDCl3) of 0.45 and 0.35 (2H, m, CHHand CHHfrom Cp), 0,7-0,6 (2H, m, CHH and CHH from Cp), of 1.02 (3H, t, J=7,5, CHCH2CH3), of 1.16 (3H, d, J=7,5, CHCH3OH), 1,35-1,2 (1H, m, CHCHHCH3), 1,68-to 1.38 (2H, m, CHCHHCH3and CH from Cp), 3,9-3,8 (2H, m, CHOH and CHEt), of 4.05 (2H, d, J=7,5, NCH2Cp), was 4.76 (2H, s, of user., NHCH2Ar), to 4.87 (1H, d, J=5, OH), 5,77 (1H, s, of user., NH), to 6.67 (1H, s, of user., NH), 7,22-7,17 (1H, m, ArH), 7,53 (1H, s, ArH), the 7.65 (1H, DD, J=2,5, And 7.5, ArH), and 8.5 (1H, d, J=5, ArH), 8,61 (1H, s, ArH), 1,11-of 0.95 (1H, m, CHfrom Cp), of 2.56 (1H, s, of user., NH), of 3.69 (2H, d, J=7,5, NCH2Cp), 4,58 (2H, s, of user., NHCH2Ar),? 7.04 baby mortality-of 6.96 (1H, m, ArH), 7,51 was 7.45 (2H, m, ArH), compared to 8.26-8,24 (1H, m, ArH), of 8.37 (1H, s, of user., ArH); δC(250 MHz, CDCl3) 160,3 (C), 154,71 (C), 151,05 (C), 151,05 (C), 149,28 (CH), 148,64 (CH), 136,85 (CH), 135,30 (CH), 134,56 (C), to 129.2 (C), 123,37 (CH), 122,1 (C), 114,21 (C), 77,27 (CH), 59,5 (CH), 48,02 (CH2), 41,99 (CH2), to 24.84 (CH2), 17,4 (CH3), 11,53 (CH3), br11.01 (CH3), 4.2V (CH2); m/z 382,3 (M+H).

Example 6

6-Chloro-2-fluoro-9-isopropyl-N-purine

A mixture of 2-fluoro-6-chloropurine (2 g, 11.7 mmol) and powdered potassium carbonate (4 g, 28 mmol) was vigorously stirred in 30 ml of DMF. Isopropylated (6 ml, 60 mmol) very slowly obavljale within 2 hours. The reaction mixture was stirred for another 5 hours. DMF was removed and the crude product was taken into ethyl acetate, washed with water (50 ml), saturated salt solution (50 ml), dried (MgSO4) and concentrated. The crude product was purified column chromatography on silica gel (30% ethyl acetate in hexane), to obtain the pure product as white solid (1.1 g, 44%).

δH(CD3OD, 250 MHz) 1,65 [6H, d, J=7,5, CHC(CH3)2], 4,92 [1H, m, CH(CH3)2], 8,66 (1H, s, ArH); δWith(CD3OD, 250 MHz) 154,7 (C), 153,88 (C), 152,2 (C), 147,65 (CH), 132,44 (C), 50,66 (CH), 22,72 (2×CH3); m/z 215,2 (M+H).

N-Cyclopropyl-2-fluoro-9-isopropyl-N-purine-6-amine

6-Chloro-2-fluoro-9-isopropyl-N-purine (0.3 g, 1.4 mmol), diisopropylethylamine (0.2 g, 1.55 mmol) and cyclopropylamine (0.2 g, of 2.66 mmol) were stirred together in ethanol (30 ml) at room temperature for 6 hours. Volatiles were removed and the residue was taken in ethyl acetate, washed with water (50 ml), saturated salt solution (50 ml), dried (MgSO4) and concentrated. The crude product was purified column flash chromatography on silica gel (ethyl acetate:hexane 3:2), to obtain the pure product (232 mg, 70,5%).

δH(CDCl3, 500 MHz) 0,63 is 0.55 (2H, m, CHHCHH from Cp), 0,85 - 0,78 (2H, m, CHHCHHfrom Cp), and 1.54 (6H, d, J=7,5, CH[CH3]2), of 2.97 (1H, s, of user., CH from Cp), 4,73-to 4.62 (1H, m, Cu> HMe2), 6,72 (1H, s, of user., NH) and 7.7 (1H, s, ArH); δC(CDCl3, 500 MHz) 159,29 (C), 157,63 (C), 156,8 (C), 156,64 (C), 137,69 (CH), 47,35 (CH), 22,6 (2×CH3), 21,06 (CH), 7,28 (2×CH2); m/z 236,2 (M+H).

2R,3S-3-(6-Cyclopropylamino)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol [6]

To a stirred solution of N-cyclopropyl-2-fluoro-9-isopropyl-N-purine-6-amine (232 mg, 0,99 mmol) in N-methylpyrrolidinone (10 ml) at room temperature in an argon atmosphere was added DIEA (0.2 ml, 10,96 EQ., to 1.14 mmol), followed by addition of (2R,3S)-3-aminoindan-2-ol (540 mg, 5 EQ., the 5.25 mmol). The reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 24 hours. The reaction mixture was allowed to cool to room temperature and was added an excess of water, whereupon the product separated in the form of oil. Added ethyl acetate and the organic layer was carefully washed with water (3×50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified column flash chromatography on silica gel with elution by ethyl acetate, to obtain specified in the header of the product as a pale brown solid (46 mg, 15%).

δH(CDCl3, 500 MHz) of 0.58-0,49 (2H, m, CHHCHH from Cp), 0,83 is 0.81 (2H, m, CHHCHHfrom Cp), of 1.03 (3H, t, J=7,5, CHCH2CH3), of 1.13 (3H, d, J=7,5, CHCH3OH), of 1.55 (6H, d, J=7,5, CH[CH3]2),1,57-to 1.45 (2H, m, CHCH2CH3), only 2.91 (1H, s, of user., CH from Cp), of 3.95 (2H, s, of user., CHMe2and CHEt), 4,6-of 4.57 (1H, m, CHMeOH), 4,84 (1H, s, of user., NH), and 6.25 (1H, s, of user., NH), 7,49 (1H, s, ArH); δC(CDCl3, 500 MHz) 160,16 (C), 155,91 (C), 150,24 (C), 134,57 (CH), 114,63 (C), 71,57 (CH), 59,88 (CH), 46,27 (CH), to 25.15 (CH2), 22,59 (CH3), 17,22 (CH3), to 11.61 (CH3), 7,34 (CH2), 47,35 (CH), 22,6 (2×CH3), 21,06 (CH), 7,28(2×CH2); m/z 319,3 (M+H).

Example 7

N-(Cyclopropylmethyl)-2-fluoro-9-isopropyl-N-purine-6-amine

6-Chloro-2-fluoro-9-isopropyl-N-purine (0.3 g, 1.4 mmol), diisopropylethylamine (0.2 g, 1.55 mmol) and cyclopropylamine (0.24 g, 2.7 mmol) was stirred with ethanol (30 ml) at room temperature for 6 hours. Volatiles were removed and the residue was taken in ethyl acetate, washed with water (50 ml), saturated salt solution (50 ml), dried (MgSO4) and concentrated. The crude product was purified column flash chromatography on silica gel (ethyl acetate:hexane 3:2), to obtain the pure product (290 mg, 83%) as a colourless resin.

δH(CDCl3, 500 MHz) of 0.29 to 0.27 (2H, m, CHHCHH from Cp), up 0,56 0,54 (2H, m, CHHCHHfrom Cp), 1,13 (1H, s, of user., CH from Cp), was 1.58 (6H, d, J=7,5, CH[CH3]2), 3,44 (2H, s, of user., NHCH2Cp) of 2.97 (1H, s, of user., CH from Cp), 4,74-4,72 (1H, m, CHMe2), to 6.22 (1H, s, of user., NH), 7,76 (1H, s, ArH); δWith(CDCl3, 500 MHz) 160,09 (C), 156,14 (C), of 147.4 (C), 137,38 (CH), 118,26 (C), 47,14 (CH), 45,78 (CH2), 22,46 (2×CH3), 10,06 (CH), 3,5 (2×CH2).

2R,3S-3-(6-(Cyclopropylamino)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol [7]

To a stirred solution of N-(cyclopropylmethyl)-2-fluoro-9-isopropyl-N-purine-6-amine (290 mg, 1.1 mmol) in N-methylpyrrolidinone (10 ml) at room temperature in an argon atmosphere was added DIEA (1.42 g, 1.11 mmol), followed by addition of (2R,3S)-3-aminoindan-2-ol (566 mg, 5 EQ., 5.5 mmol). The reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 24 hours. The reaction mixture was allowed to cool to room temperature and was added an excess of water, whereupon the product separated in the form of oil. Added ethyl acetate and the organic layer was carefully washed with water (4×50 ml). The organic layer was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified column flash chromatography on silica gel with elution by ethyl acetate, to obtain specified in the header of the product as a pale brown solid (120 mg, 31%).

δH(CDCl3, 500 MHz) is 0.22 to 0.19 (2H, m, CHHCHHfrom Cp), 0,36-0,34 (2H, m, CHHCHHfrom Cp), and 0.8 (3H, t, J=7,5, CHCH2CH3), to 0.89 (3H, d, J=7,5, CHCH3OH), 1,13 (1H, s, of user., CH from Cp), of 1.33 (6H, d, J=7,5, CH[CH3]2), 3,24 (2H, s, of user., NHCH2Cp) 3,7 (2H, with usher., CHEt and CHMe2), of 6.52 (1H, s, of user., NH), 7,26 (1H, s, ArH); δC(CDCl3, 500 MHz) 160,2 (C), 154,89 (C), 134,35 (CH), 71,72 (CH), 59,76 (CH), 46,238 (CH), 25,23 (CH2), 22,59 (CH3), 17,15 (CH3), to 11.61 (CH3), 3,48 (2×CH2); m/z to 333.3 (M+H).

Example 8

N-Cyclobutyl-2-fluoro-9-isopropyl-N-purine-6-amine

6-Chloro-2-fluoro-9-isopropyl-N-purine (0.3 g, 1.4 mmol), diisopropylethylamine (0.2 g, 1.55 mmol) and cyclobutylamine (0.2 g, 2.8 mmol) was stirred with ethanol (30 ml) at room temperature for 6 hours. Volatiles were removed and the residue was taken in ethyl acetate, washed with water (50 ml), saturated salt solution (50 ml), dried (MgSO4) and concentrated. The crude product was purified column flash chromatography on silica gel (ethyl acetate:hexane 3:2), to obtain the pure product (237 mg, 68%).

δH(CDCl3, 500 MHz) of 1.56 (6H, d, J=7,5, CH[CH3]2), 1,76-of 1.74 (2H, m, CH2cyclobutyl), 1,98 -1,94 (2H, m, CH2cyclobutyl), a 2.45 (2H, s, of user., CH2cyclobutyl)) 4,72-4,7 (2H, m, CH of cyclobutyl and CHMe2), 6.35mm (1H, s, of user., NH), of 7.75 (1H, s, ArH); δC(CDCl3, 500 MHz) 160,08 (C), 158,43 (C), 155,34 (C), 155,18 (C), 150,09 (C), 137,44 (CH), 47,26 (CH), 31,56 (2×CH2), 22,38 (2×CH3), 15,09 (CH2); m/z 250,2 (M+H).

2R,3S-3-(6-(Cyclobutylamine)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol [8]

To a stirred solution of N-cycle is butyl-2-fluoro-9-isopropyl-N-purine-6-amine (237 mg, 0.95 mmol) in N-methylpyrrolidinone (10 ml) at room temperature in an argon atmosphere was added DIEA (1.28 g, 10 mmol), followed by addition of (2R,3S)-3-aminoindan-2-ol (566 mg, 5 EQ., 5.5 mmol). The reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 24 hours. The reaction mixture was allowed to cool to room temperature and was added an excess of water, whereupon the product separated in the form of oil. Added ethyl acetate and the organic layer was carefully washed with water (4×50 ml). The organic phase is washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified column flash chromatography on silica gel with elution by ethyl acetate, to obtain specified in the header of the product as a pale brown solid (120 mg, 31%).

δH(CDCl3, 500 MHz)of 0.97 (3H, t, J=7,5, CHCH2CH3), with 1.07 (3H, d, J=7,5, CHCH3OH), to 1.47 (6H, d, J=7,5, CH[CH3]2), 1,53-of 1.42 (2H, m, CHCH2CH3), 1,71 by 1.68 (2H, m, CH2cyclobutyl), 1,89-to 1.87 (2H, m, CH2cyclobutyl), 2,39-is 2.37 (2H, m, CH2cyclobutyl), the 3.89 (1H, d, J=5, NH cyclobutyl)), 4,53 to 4.5 (3H, m, CHMe2and CHEt and CH cyclobutyl), 5,62 (1H, m, CHMeOH), x 6.15 (1H, s, of user., NH), the 7.43 (1H, s, ArH); δC(CDCl3, 500 MHz) 161,16 (C), 154,76 (C), 152,2 (C), 134,43 (CH), 114,67 (C)of 71.7 (CH), 59,74 (CH), 46,36 (CH), 31,68 (CH2in ), 2.25 (2×CH 2), 22,59 (CH3), 17.11 per bbl (CH3), 15,04 (CH2), to 11.61 (CH3); m/z to 333.3 (M+H).

Example 9

(2-fluoro-9-H-purine-6-yl)pyridine-4-ylmethylamino

To a stirred solution of 6-chloro-2-porporino (1 g, 5.8 mmol) in n-BuOH (50 ml) in an argon atmosphere at 0°C was added diisopropylethylamine (3 ml of 17.4 mmol), followed by addition of 4-(aminomethyl)pyridine (0.9 ml, 1.5 EQ., 8,7 mmol). The reaction mixture was stirred at this temperature for 1 hour, then was allowed to return to room temperature and was stirred for 4 hours, until TLC (CHCl3:MeOH; 90:10) showed that the reaction went to completion. The solvent is evaporatedin the vacuumand the residue was purified gradient column chromatography on silica gel with elution CHCl3:MeOH(100:0→90:10), with obtaining the product as a white solid; yield: 0,85 g (60%). TPL 200-202°C.

1H-NMR (d6-DMSO, 250 MHz): δ of 4.67 (2H, d, J=5, -HNCH2-Pyr), 7,34-7,28 (2H, m, Pyr-H), 8,48-8,42 (2H, m, Pyr-H), 8,54 (1H, s, -N=CH-NH-), 8,84 (1H, s, of user., -HNCH2-Pyr), 13,13 (1H, s, of user., -N=CH-NH-);m/z245 ([M+H]+.

(2-Fluoro-9-isopropyl-N-purine-6-yl)pyridine-4-ylmethylamino

To a stirred solution of (2-fluoro-N-purine-6-yl)pyridine-4-ylmethylamino (0.6 g, the 2.46 mmol) in DMF (10 ml) in an argon atmosphere while K.T. was added To the2CO3(powder is anhydrous, 1.7 g, 5 EQ., 12.3 mmol), followed by the addition of 2-bromopropane (2.3 ml, 10 EQ., 24.6 mmol). The reaction mixture was stirred at K.T. within 24 hours, until TLC (CHCl3:MeOH; 90:10) showed that the reaction went to completion. The solvent is evaporatedin the vacuumand the residue was distributed between water (200 ml) and EtOAc (50 ml). The aqueous phase was separated and additionally was extracted with EtOAc (2×50 ml). The combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuumthe residue was purified gradient column chromatography on silica gel with elution with a mixture of chloroform:methanol(100:0→95:5), obtaining the product as a white solid; yield: 0.40 g (57%). TPL 170-173°C.

1H-NMR (d6-DMSO, 250 MHz): δ for 1.49 (6H, d, J=7,5, -CH(CH3)2), 4,63 (3H, m, -CH(CH3)2+-HNCH2-Pyr), 7,30, of 8.47 (4H, 2×m, Pyr-H), of 8.28 (s, 1H, -N=CH-N-), 8,97 (1H, s, of user., -HNCH2-Pyr); m/z: 287 [M+H].

2R,3S-3-(9-isopropyl-6 - (pyridine-4-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol [9]

To a stirred solution of (2-fluoro-9-isopropyl-N-purine-6-yl)pyridine-4-ylmethylamino (300 mg, 1.05 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (2 ml, 10 EQ., 10.5 mmol), followed by addition of (2R,3S)-3-aminoindan-2-ol (600 mg, 5.5 equiv., 5.8 mmol). The flask is supplied back what holodilniki, the reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 72 hours. The reaction mixture was allowed to cool to room temperature and the solvent evaporatedin the vacuum. The residue was distributed between EtOAc (50 ml) and water (50 ml), the aqueous phase was additionally extracted with EtOAc (2×25 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified gradient column flash chromatography on silica gel with elution CHCl3:MeOH(100:0→95:5), obtaining the product as a colorless film. Output (225 mg, 58%);

δH(CDCl3, 500 MHz) to 0.92 (3H, t, J=7,5, CHCH2CH3), of 1.05 (3H, d, J=7,5, CHCH3OH), 1,4 (6H, d, J=7,5, CH[CH3]2), 1,47-of 1.36 (2H, m, CHCH2CH3), 3,88-of 3.85 (2H, m, CHEt and NH), 4.53-in-4,5 (1H, m, CHCH3OH), 4,72 (2H, s, of user., NHCH2Ar), and 6.5 (1H, s, of user., NH), 7,2 (2H, s, of user., ArH), 7,45-7,42 (1H, m, ArH), 8,46-8,43 (2H, m, ArH); δC(CDCl3, 500 MHz) 154,66 (C), 149,86 (2×CH), 149,69 (C), 148,26 (C), 134,84 (CH), 122,22 (2×CH), 71,51 (CH), 59,57 (CH), 46,58 (CH), 43,45 (CH2), 24,9 (CH2), 22,59 (2×CH3), 17,27 (CH3), 11,53 (CH3); m/z 370,2 (M+H).

Example 10

2,6-Dimethylethanolamine

To stir the amount of 2,6-lutidine-1-oxide (12.3 g, 100 mmol) was slowly added dimethylsulfate (12, g, 100 mmol) at such a speed that the temperature of the reaction mixture was kept at 80°C during the addition. When the addition was complete (approximately one hour), the solution was stirred at the same temperature for another 2 hours. Salt crystallized upon cooling and was recrystallized from anhydrous acetone gave white prisms; TPL 96-97°C. the Yield 18 g (73%). To a solution of the obtained 1-methoxy-2,6-dimethylaminoethylacrylate (11,65 g, 50 mmol) in water (50 ml) under nitrogen atmosphere was added a solution of potassium cyanide (10 g, 150 mmol), dissolved in 50 ml of water. The solution was left to stand at room temperature for 2 days, at this time, nitrile, which was separated from the solution in the form of long white needles were removed by filtration, obtaining 2.8 g of pure product (42%).

δH(CDCl3, 500 MHz) of 2.44 (6H, s, 2×CH3)and 4.2 (2H, d, J=5, NHCH2), 6,78 (2H, s, ArH); δC(CDCl3, 500 MHz) 157,83 (2×C), 122,71 (C), 118,2 (2×CH), 24,28 (2×CH3).

tert-Butyl(2,6-dimethylpyridin-4-yl)methylcarbamate

2,6-dimethylethanolamine (2 g, br15.15 mmol) was dissolved in a mixture of 10% acetic acid/ethanol (30 ml). Added catalyst 10% palladium on carbon (0.5 g) and the reaction mixture was stirred in an atmosphere of hydrogen for 24 hours at 60°C. the Mixture was filtered through a loose layer of cellite. Volatiles were removed and neojidanni the residue was dissolved in dichloromethane (30 ml). To the stirred solution was then added triethylamine (5 ml), followed by addition of di-tert-BUTYLCARBAMATE (3.3 grams, br15.15 mmol). After 3 hours the solvent was removed and the residue was dissolved in ethyl acetate. The resulting solution was washed with water (50 ml), saturated bicarbonate solution (50 ml), dried and evaporated. The crude product was purified column flash chromatography on silica gel (ethyl acetate:hexane 1:2), to obtain 0.6 g of the net specified in the connection header (17,24% yield).

δH(CDCl3, 500 MHz) of 1.42 (9H, s, 3×CH3), is 2.44 (6H, s, 2×CH3)and 4.2 (2H, d, J=5, NHCH2in ), 5.25 (1H, s, of user., NH), for 6.81 (2H, s, ArH); δC(CDCl3, 500 MHz) 157,83 (C), 156,01 (CO), 148,71 (C), 118,58 (2×CH), 79,77 (C), 43,41 (CH2), 28.34 points (3×CH3), 24,28 (2×CH3); m/z of 237.2 (M+H).

N-((2,6-Dimethylpyridin-4-yl)methyl)-2-fluoro-N-purine-6-amine

To a stirred solution of 6-chloro-2-porporino (0.4 g, 2.9 mmol) in n-BuOH (50 ml) in an argon atmosphere at 0°C was added DIEA (2.5 ml, 14.7 mmol), followed by addition of (2,6-dimethylpyridin-4-yl)methanamine (0.54 g, 4 mmol). The reaction mixture was stirred at this temperature for 1 hour, then was allowed to return to room temperature and was stirred for 4 hours, it was observed that the reaction is not yet completed, and therefore the reaction mixture was heated to 100°C and left at this temperature for 8 hours. Dissolve Italy evaporated in the vacuumand the residue was purified gradient column flash chromatography on silica gel with elution CHCl3:MeOH(100:0→90:10), obtaining the product as a white solid; yield: 0.5 g (63%).

δH(CDCl3, 250 MHz) 2,48 (6H, s, 2×CH3), the 3.65-of 3.54 (2H, m, NHCH2), br4.61 (1H, s, of user., NH), 7,32 (2H, s, ArH), to 7.84 (1H, s, ArH),), 9,05 (1H, s, of user., NH); δC(CDCl3, 250 MHz) 159,2(C), 156,62 (C), 156,61 (C), 155,46 (C), 146,92 (CH), 136,3 (C), 122,05 (2×CH), 119,48 (C), to 41.6 (CH2), 18,06 (2×CH3).

N-((2,6-Dimethylpyridin-4-yl)methyl)-2-fluoro-9-isopropyl-N-purine-6-amine

To a stirred solution of 2-fluoro-N-(2,6-dimethylpyridin-4-yl)methyl-N-purine-6-amine (0.4 g, 1.48 mmol) in dimethylformamide (10 ml) at room temperature in an argon atmosphere was added powdered anhydrous K2CO3(1 g, 5 EQ., 7.4 mmol), followed by the addition of 2-bromopropane (1.2 ml, 12.2 mmol). The reaction mixture was stirred at room temperature for 24 hours until TLC with a mixture of DCM:ether:Meon (55:40:5) showed that the reaction went to completion. The solvent is evaporatedin the vacuumand the residue was distributed between EtOAc (50 ml) and water (50 ml). The aqueous phase was additionally extracted with EtOAc (2×50 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified column chromatography naselesele when the elution CHCl 3:MeOH (98:2), obtaining specified in the title compounds as a colorless film (250 mg, 54%).

δH(CDCl3, 500 MHz) of 1.46 (6H, d, J=7,5, CH[CH3]2), of 2.38 (3H, s, ArCH3), 2,39 (3H, s, ArCH3), 4,73-4,6 (3H, m, NHCH2and CHMe2), 6,85 (2H, s, ArH), 7,35 (1H, s, of user., NH), and 7.5 (1H, s, of user., ArH); δc(CDCl3, 500 MHz) 157,8 (C), 158,6 (C), 154,32 (C), 151 (C), 148,42 (CH), 122,2 (2×CH), 121,08 (C), 47,52 (CH), 43,6 (CH2), 23,68 (2×CH3), 22,3 (2×CH3); m/z 315,2 (M+H).

2R,3S-3-(9-isopropyl-6-(2,6-dimethylpyridin-4-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol [10]

To a stirred solution of 2-fluoro-9-isopropyl-N-(2,6-dimethylpyridin-4-ylmethyl)-N-purine-6-amine (250 mg, 0.8 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (1.4 ml, 10 EQ., 8 mmol), followed by addition of (2R,3S)-3-aminoindan-2-ol (0.35 g, 3.4 mmol). The flask was supplied reflux, the reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 72 hours. The reaction mixture was allowed to cool to room temperature and the solvent evaporatedin the vacuum. The residue was distributed between EtOAc (50 ml) and water (50 ml), the aqueous phase was additionally extracted with EtOAc (2×25 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin in the cosmology vacuum . The residue was purified gradient column flash chromatography on silica gel with elution CHCl3:MeOH(100:0→95:5), obtaining specified in the title compound as a colourless oil (56 mg, 18%).

δH(CDCl3, 500 MHz) of 0.95 (3H, t, J=7,5, CHCH2CH3), of 1.05 (3H, d, J=5, CHCH3OH), was 1.43 (6H, d, J=10, CH[CH3]2), 1,55-1,4 (2H, m, CHCH2CH3), 2,43 (3H, s, ArCH3), is 2.44 (3H, s, ArCH3), 3,9-3,8 (2H, m, CHEt and CHMe2), 4,57-to 4.46 (1H, m, CHCH3OH), 4.63 to-4,58 (2H, m, NHCH2Ar), 6,28 (1H, s, of user., NH), 6,8 (2H, s, of user., ArH), was 7.45 (1H, s, ArH); δC(CDCl3, 250 MHz) 160,08 (C), 157,76 (C), 155,36 (C), 148,97 (C), 135,36 (C), 134,75 (CH), 118,95 (2×CH), 71,52 (CH), 59,59 (CH), 46,65 (CH), 44,92 (CH2), 24,91 (CH2), 24,01 (2×CH3), 22,5 (2×CH3), 17,27 (CH3), 11,51 (CH3); m/z 398,3 (M+H).

Example 11

2-Fluoro-N-((6-trifluoromethyl)pyridin-3-yl)methyl)-N-purine-6-amine

To a stirred solution of 6-chloro-2-porporino (344 g, 2 mmol) and [6-(trifluoromethyl)pyridin-3-yl]methanamine (0.4 g, of 2.27 mmol) in n-BuOH (50 ml) in an argon atmosphere at 0°C was added DIEA (2.5 ml, 14.7 mmol). The reaction mixture was stirred at this temperature for 1 hour, then was allowed to return to room temperature and was stirred for 4 hours, it was observed that the reaction is not yet completed, and therefore the reaction mixture was heated to 100°C and left at this temperature is 5 hours. The solvent is evaporatedin the vacuumand the residue was purified gradient column chromatography on silica gel with elution CHCl3:MeOH(100:0→90:10), obtaining the product as a white solid; yield: 0,46 g (75%).

δH(CDCl3, 500 MHz) 3,62 is 3.57 (2H, m, NHCH2), 4,69 (1H, s, of user., NH), to 7.2, and 7.1 (2H, s, of user., ArH and NH), of 7.69 (1H, d, J=7,5, ArH), of 8.27 (1H, s, ArH), 8,71 (1H, s, ArH); δC(CDCl3, 500 MHz) 158,69 (C), 158,18 (C), 157,67 (C), 154,56 (C), 147,48(CH), 146,99 (CH), 144,32 (C), 133,87 (CH), 121,67 (CH), 118,88 (C), 43,48 (CH2); m/z 313,1 (M+H).

2-Fluoro-9-isopropyl-N-((6-(trifluoromethyl)pyridin-3-yl)methyl)-N-purine-6-amine

To a stirred solution of 2-fluoro-N-[(6-(trifluoromethyl)pyridin-3-yl)methyl-N-purine-6-amine (0.4 g, of 1.27 mmol) in dimethylformamide (10 ml) at room temperature in an argon atmosphere was added powdered anhydrous K2CO3(0,86 g, 5 EQ., is 6.54 mmol), followed by the addition of 2-bromopropane (1 ml of 10.3 mmol). The reaction mixture was stirred at room temperature for 24 hours until TLC with a mixture of DCM:ether:Meon (55:40:5) showed that the reaction went to completion. The solvent is evaporatedin the vacuumand the residue was distributed between EtOAc (100 ml) and water (100 ml). The aqueous phase was additionally extracted with EtOAc (2×50 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. OS is atok was purified column chromatography on silica gel with elution CHCl 3:MeOH (98:2), obtaining specified in the title compounds as a colorless film (350 mg, 77%).

δH(CDCl3, 500 MHz) 1,5 (6H, d, J=7,5, CH[CH3]2), 4,7-4,6 (3H, m, NHCH2and CHMe2), 7,16 (1H, d, J=2,5, ArH), 7,25 (1H, s, of user., NH), 7,66 (2H, s, of user., ArH), 8,67 (1H, s, of user., ArH); δC(CDCl3, 500 MHz) 158,8 (C), 156,72 (C), 156.3 m (C), 147,42 (CH), 137,92 (CH), 138,08 (CH), 131,73 (C), 123,31 (CH), 119,25 (C), 48.38 per (CH), 44.1kHz (CH2), 22,72 (2×CH3); m/z 355,1 (M+H).

2R,3S-3-(9-Isopropyl-6-((6-trifluoromethyl)pyridin-3-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol [11]

To a stirred solution of 2-fluoro-9-isopropyl-N-(6-(triptorelin-3-ylmethyl)-N-purine-6-amine (200 mg, of 0.56 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (1.4 ml, 10 EQ., 8 mmol), followed by addition of (2R,3S)-3-aminoindan-2-ol (0.25 g, 2.4 mmol). The flask was supplied reflux, the reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 72 hours. The reaction mixture was allowed to cool to room temperature and the solvent evaporatedin the vacuum. The residue was distributed between EtOAc (50 ml) and water (50 ml), the aqueous phase was additionally extracted with EtOAc (2×25 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. Ostate the purified gradient column flash chromatography on silica gel with elution CHCl 3:MeOH(100:0→95:5), obtaining specified in the title compound as a colourless oil (30 mg, 12%).

δH(CDCl3, 500 MHz) of 0.85 (3H, t, J=7,5, CHCH2CH3), of 1.12 (3H, d, J=5, CHCH3OH), of 1.52 (6H, d, J=7,5, CH[CH3]2), 1,61-of 1.41 (2H, m, CHCH2CH3), 3,82 (1H, m, CHMe2), 4,75-of 4.57 (1H, m, CHCH3OH), 4,82 with 4.64 (2H, m, NHCH2Ar), and 5.8 (1H, s, of user., NH), 6,85 (1H, s, of user., NH), 7 (1H, d, J=10, ArH), 7,31-of 7.23 (2H, m, ArH), to 8.45 (1H, s, of user., ArH); δC(CDCl3, 250 MHz) 158,61 (C), 157,35 (C), 156,97 (C), 147,62 (CH), 137,43 (CH), 129,22 (C), 128,72 (C), 128,92 (CH), 123,14 (CH), 118,32 (C), 69,64 (CH), 60,91 (CH2), 57,68 (CH), 23,55 (CH2), 22,53 (2×CH3), 12,12 (CH3); m/z 438,3 (M+H).

Example 12

2-Fluoro-N-((6-methylpyridin-2-yl)methyl)-N-purine-6-amine

To a stirred solution of 6-chloro-2-porporino (0.4 g, 2.3 mmol) in n-BuOH (50 ml) in an argon atmosphere at 0°C was added DIEA (2.5 ml, 14.7 mmol), followed by addition of (6-methylpyridin-2-yl)methanamine (0.36 g, 2,95 mmol). The reaction mixture was stirred at this temperature for 1 hour, then gave it to return to room temperature and was stirred for 4 hours, it was observed that the reaction is not yet completed, and therefore the reaction mixture was heated to 100°C and left at this temperature for 8 hours. The solvent is evaporatedin the vacuumand the residue was purified gradient column flash chromatography on forces is the Kagel when the elution CHCl 3:MeOH(100:0→90:10), obtaining the product as a white solid; yield: 0.35 g (60%).

δH(CDCl3, 500 MHz) of 2.44 (3H, s, CH3), a 4.83-of 4.54 (3H, m, NH and NHCH2), 7,1-7,05 (2H, m, ArH), to 7.61-of 7.55 (1H, m, ArH), 8,13-of 8.09 (1H, m, ArH), 8,61(1H, s, of user., NH); δC(CDCl3, 500 MHz) 159,41 (C), 158,78 (C), 158,53 (C), 157,19 (C), 155,38 (C), 147,69 (CH), 137,03 (C), 136,93 (CH), 121,34 (CH), 117,76 (CH), 117,31 (C), 46,47 (CH2), 23,9 (CH3); m/z 259,2 (M+H).

2-Fluoro-9-isopropyl-N-((6-methylpyridin-2-yl)methyl)-N-purine-6-amine

To a stirred solution of 2-fluoro-N-[(6-methylpyridin-2 - yl)methyl-N-purine-6-amine (0.3 g, at 1.17 mmol) in dimethylformamide (10 ml) at room temperature in an argon atmosphere was added powdered anhydrous K2CO3(0.8 g, 5 EQ., to 5.85 mmol), followed by the addition of 2-bromopropane (1,15 ml, 11.7 mmol). The reaction mixture was stirred at room temperature for 24 hours until TLC with a mixture of DCM:ether:Meon (55:40:5) showed that the reaction went to completion. The solvent is evaporatedin the vacuumand the residue was distributed between EtOAc (100 ml) and water (100 ml). The aqueous phase was additionally extracted with EtOAc (2×50 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified column chromatography on silica gel with elution CHCl3:MeOH (98:2), with the specified header connection is in the form of a slightly yellow film (180 mg, 51%).

δH(CDCl3, 500 MHz) of 1.6 (6H, d, J=7,5, CH[CH3]2), to 2.54 (3H, s, CH3), a 4.86-to 4.73 (3H, m, NHCH2and CHMe2), 7,06 (1H, d, J=10, ArH), 7,14 (1H, d, J=10, ArH), 7,53 (1H, t, J=10, ArH), and 7.8 (1H, s, ArH); δC(CDCl3, 500 MHz) 158,42 (C), 158,04 (C), 156,16 (C)156 (C), 155,49 (C), 150,12 (C), 137,67 (CH), 136,93 (CH), 121,89 (CH), 118,1 (C), 118,53 (CH), 47,15 (CH), 45,52 (CH2), 24,38 (CH3), 22,58 (2×CH3); m/z 301,2 (M+H);19F NMR δ -50,22.

2R,3S-3-(9-Isopropyl-6-((6-methylpyridin-2-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol [12]

To a stirred solution of 2-fluoro-9-isopropyl-N-(6-methylpyridin-2-ylmethyl)-N-purine-6-amine (180 mg, 0.59 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (1 ml, 10 EQ., 5,6 mmol), followed by addition of (2R,3S)-3-aminoindan-2-ol (0.34 g, 6 mmol). The flask was supplied reflux, the reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 72 hours. The reaction mixture was allowed to cool to room temperature and the solvent evaporatedin the vacuum. The residue was distributed between EtOAc (50 ml) and water (50 ml), the aqueous phase was additionally extracted with EtOAc (2×25 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified gradient column flash chromatography n the silica gel with elution CHCl 3:MeOH(100:0→95:5), obtaining specified in the title compound as a slightly yellow oil (28 mg, 13%).

δH(CDCl3, 500 MHz) of 1.05 (3H, t, J=7,5, CHCH2CH3), of 1.13 (3H, d, J=5, CHCH3OH), was 1.43 (6H, d, J=7,5, CH[CH3]2), 1,6-1,4 (2H, m, CHCH2CH3), a 1.75 (1H, s, of user., NH), 2,32 (3H, s, ArCH3), 3,95 (1H, s, of user., CHMe2), 4,57-a 4.53 (1H, m, CHCH3OH), 4,8-4,6 (2H, m, NHCH2Ar), and 5.8 (1H, s, of user., NH), PC 6.82 (1H, s, of user., NH), to 6.95 (1H, d, J=5, ArH), 7,05 (1H, d, J=5, ArH), 7,45 to 7.4 (2H, m, ArH); δC(CDCl3, 500 MHz) 160,17 (C), 157,9 (C), 156,72 (C), 154,8 (C), 150,12 (C), 136,8 (CH), 134,55 (CH), 121,64 (CH), 118,58 (CH), 114,87(C), 71,59 (CH), 59,66 (CH), 46,41 (CH), 44,79 (CH2), 24,12 (CH2), 23,41 (CH3), 21,59 (CH3), 16,18 (CH3), 10,6 (CH3); m/z 384,3 (M+H).

Example 13

2-Fluoro-N-((3-methylpyridin-2-yl)methyl)-N-purine-6-amine

To a stirred solution of 6-chloro-2-porporino (0.4 g, 2.3 mmol) in n-BuOH (50 ml) in an argon atmosphere at 0°C was added DIEA (2.5 ml, 14.7 mmol), followed by addition of (3-methylpyridin-2-yl)methanamine (0.36 g, 2,95 mmol). The reaction mixture was stirred at this temperature for 1 hour, then was allowed to return to room temperature and was stirred for 4 hours, it was observed that the reaction is not yet completed, and therefore the reaction mixture was heated to 100°C and left at this temperature for 8 hours. The solvent is evaporatedin the vacuumand the residue was purified the gradient column chromatography on silica gel with elution CHCl 3:MeOH(100:0→90:10), obtaining the product as a white solid; yield: 0.3 g (51%).

δH(CDCl3, 500 MHz) to 2.35 (3H, s, CH3), 4,71 (3H, m, NH and NHCH2), from 7.24 (1H, s, of user., ArH), 7,63 (1H, s, of user., ArH), of 8.28 (1H, s, of user., ArH), scored 8.38 (1H, s, of user., ArH); δC(CDCl3, 500 MHz) 161,03 (C), 158,49 (C), 154,77 (C), 152,16 (C), 144,86(CH), 137,53 (CH), 137,14 (CH), 130,67 (C), 121,58 (CH), 118,73 (C), 42,87 (CH2), 18,16 (CH3); m/z 259,3 (M+H).

2-Fluoro-9-isopropyl-N-((3-methylpyridin-2-yl)methyl)-N-purine-6-amine

To a stirred solution of 2-fluoro-N-(3-methylpyridin-2-yl)methyl-N-purine-6-amine (0.3 g, at 1.17 mmol) in dimethylformamide (10 ml) at room temperature in an argon atmosphere was added powdered anhydrous K2CO3(0.8 g, 5 EQ., to 5.85 mmol), followed by the addition of 2-bromopropane (1,15 ml, 11.7 mmol). The reaction mixture was stirred at room temperature for 24 hours until TLC with a mixture of DCM:ether:Meon (55:40:5) showed that the reaction went to completion. The solvent is evaporatedin the vacuumand the residue was distributed between EtOAc (50 ml) and water (50 ml). The aqueous phase was additionally extracted with EtOAc (2×50 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified column chromatography on silica gel with elution CHCl3:MeOH (98:2), obtaining specified in the header of the is placed in the form of a slightly yellow film (170 mg, 48%).

δH(CDCl3, 500 MHz) is 1.51 (6H, d, J=7,5, CH[CH3]2), of 2.28 (3H, s, CH3), 4,68 with 4.65 (3H, m, NHCH2and CHMe2), 7,08-7,05 (1H, m, ArH), and 7.4 (1H, d, J=5, ArH), 7,72 (1H, s, ArH), to 7.93 (1H, s, of user., NH), a 8.34 (1H, d, J=5, ArH); δC(CDCl3, 500 MHz) 160,09 (C), 158,43 (C), 155,97 (C), 154,06 (C), 145,85 (CH), 137,73 (CH), 137,54 (CH), 130,67 (C), 122,28 (CH), 118,73 (C), 47,28 (CH), 42,87 (CH2), 22,43 (2×CH3), 17,66 (CH3);19F NMR δ -50,20; m/z 301,2 (M+H).

2R,3S-3-(9-Isopropyl-6-((3-methylpyridin-2-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol [13]

To a stirred solution of 2-fluoro-9-isopropyl-N-(3-methylpyridin-2-ylmethyl)-N-purine-6-amine (170 mg, of 0.56 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (1 ml, 10 EQ., 5,6 mmol), followed by addition of (2R,3S)-3-aminoindan-2-ol (0.34 g, 6 mmol). The flask was supplied reflux, the reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 72 hours. The reaction mixture was allowed to cool to room temperature and the solvent evaporatedin the vacuum. The residue was distributed between EtOAc (50 ml) and water (50 ml), the aqueous phase was additionally extracted with EtOAc (2×25 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified gradient column flash chromate what graphy on silica gel with elution CHCl 3:MeOH(100:0→95:5), obtaining specified in the title compound as a slightly yellow oil (28 mg, 13%).

δH(CDCl3, 500 MHz) 1 (3H, t, J=7,5, CHCH2CH3), 1,1 (3H, d, J=5, CHCH3OH), the 1.44 (6H, d, J=7,5, CH[CH3]2), 1,6-1,4 (2H, m, CHCH2CH3), a 1.75 (1H, s, of user., NH), and 2.3 (3H, s, ArCH3), 3,92 (1H, s, of user., CHMe2), 4,56-to 4.52 (1H, m, CHCH3OH), 4,7 (2H, s, of user., NHCH2Ar), or 6.1 (1H, s, of user., NH), 7,06 (1H, DD, J=2,5, 5, ArH), 7,41 (1H, DD, J=2,5, 5, ArH), 7,47 (1H, s, ArH), of 8.37 (1H, d, J=5, ArH); δC(CDCl3, 500 MHz) 160,21 (C), 154,68 (C), 153,65 (C), 146,07 (CH), 137,53 (CH), 134,53 (CH), 130,59 (C), 122,31 (C), 122,13 (CH), 115,14 (C), 71,73 (CH), 59,93 (CH), 46,43 (CH), 43,11 (CH2), 25,33 (CH2), 22,63 (CH3), 17,61 (CH3), 17,25 (CH3), 11,67 (CH3); m/z 384,3 (M+H).

Example 14

(R)-1-(9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)propan-2-ol [14]

To a stirred solution of 2-fluoro-9-isopropyl-6-[(pyridine-3-ylmethyl)amino]purine (300 mg, 1.05 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (2 ml, 10 EQ., 10.5 mmol), followed by addition of (R)-1-aminopropan-2-ol (0,395 g's, 5.25 mmol). The flask was supplied reflux, the reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 72 hours. The reaction mixture was allowed to cool to room temperature and the solvent evaporatedin the Vake is IU . The residue was distributed between ethyl acetate (50 ml) and water (50 ml), the aqueous phase was additionally extracted with ethyl acetate (2×25 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified gradient column flash chromatography on silica gel with elution CHCl3:MeOH(100:0→95:5), to obtain the pure product as a colourless oil (38 mg, 10.6 per cent).

δH(250 MHz, CDCl3) to 1.15 (3H, d, J=7,5, CHCH3OH), a 1.45 (6H, d, J=7,5, CH[CH3]2), 3,3-3,2 (1H, m, NHCHHCHMeOH)), 3,45-to 3.36 (1H, m, NHCHHCHMeOH), 3,96 to-3.9 (1H, m, CHMe2), 4,58-4,47 (1H, m, CHMeOH), 4,69 (2H, d, J=5, NHCH2Ar), 5,23 (1H, t, J=5, NHCH2CHMeOH), 6,32 (1H, s, of user., NHCH2Ar), 7,16-7,11 (1H, m, ArH), and 7.4 (1H, s, ArH), and 7.6 (1H, d, J=7,5, ArH), and 8.4 (1H, d, J=2,5, ArH), 8,55 (1H, s, ArH); δWith(250 MHz, CDCl3160,07 (C), 154,95 (C), 154,7 (C), 151,1 (C), 149,27 (CH), 148,54 (CH), 135,3 (CH), 134,73 (CH), 123,39 (CH), 114,61 (C), 68,83 (CH), 50,09 (CH2), 46,49 (CH), 41,95 (CH2), 22,49 (2×CH3), 20,85 (CH3); m/z 342,2 (M+H).

Example 15

1,1,1-Cryptor-3-(9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)propan-2-ol [15]

To a stirred solution of (2-fluoro-9-isopropyl-N-purine-6-yl)pyridine-3-ylmethylamino (300 mg, 1.05 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (1.4 ml, 10 EQ., 8 mmol), followed added the eat 3-amino-1,1,1-tryptophan-2-ol (1 g, 7.8 mmol) [obtained by the reaction of ammonia with 2-(trifluoromethyl)oxirane]. The flask was supplied reflux, the reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 72 hours. The reaction mixture was allowed to cool to room temperature and the solvent evaporatedin the vacuum. The residue was distributed between EtOAc (50 ml) and water (50 ml), the aqueous phase was additionally extracted with EtOAc (2×25 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified gradient column flash chromatography on silica gel with elution CHCl3:MeOH(100:0→95:5), obtaining specified in the title compounds as colorless oil (62 mg, 15%).

δH(CDCl3, 500 MHz) of 1.53 (6H, d, J=7,5, CH[CH3]2), 3,64 of 3.6 (1H, m, NHCHHSON[CF3][CH3]), 3,76-to 3.73 (1H, m, NHCHHCOH[CF3][CH3]), 4,18-4,16 (1H, m, CHMe2), 4,62-4,56 (1H, m, CHOHCF3), 4,72 (3H, s, of user., NHCH2Ar and NHCH2Ar), 7,22-of 7.2 (1H, m, ArH), 7,52 (1H, s, ArH), 7,66 (1H, d, J=10, ArH), of 8.47 (1H, d, J=5, ArH), 8,58 (1H, s, ArH); δC(CDCl3, 500 MHz) 159,77 (C), 154,68 (C), 149,04 (CH), 148,45 (CH), 135,44 (CH), 135,03 (CH), 134,44 (C), 125,91 (C), 123,49 (CH), 114,78 (C), 71,57 (CH), 46,76 (CH), 42,94 (CH2), 32,2 (CH2), 22,47 (2×CH3);19F δ -78,48; m/z 396,2 (M+H).

Example 16

1,1,1-Cryptor-3-nitropentane-2-ol

1-Nitropropane (3.25 g, 36.6 mmol), cryptanalytically (5,85 g, 36.6 mmol, 90% purity) and powdered To2CO3(0.34 g, 2.5 mmol) were mixed and stirred at 60°C for 3 hours, then at room temperature for 3 days. Was added a saturated solution of salt (10 ml) and 1N. a solution of HCl (10 ml), and the lower organic layer was separated. The aqueous layer was extracted with ether (2×30 ml) and the combined organic layers were washed with saturated salt solution, dried over Na2SO4, filtered and concentrated in vacuum. The residue was purified flash chromatography on silica gel with a gradient elution with a mixture of CH2Cl2:hexane (50:50), CH2Cl2:hexane (75:25), CH2Cl2(100%) and Meon:CH2Cl2(5:95), to obtain the product as a waxy white solid (4.4 g, 64%).

δH(CDCl3, 500 MHz) was 1.04 to 1.00 (3H, m, CHCH2CH3), 2,16 of 1.99 (2H, m, CHCH2CH3), 4,37 (1H, s, of user., OH), 4,71-4,59 (2H, m, 2×CH); δC(CDCl3, 500 MHz) DQC. (126,79, 126,70; 124,55, 124,46; 122,30, 122,21; 120,06, 119,96, CF3), two peaks (88,34, 87,60 CH NO2), eight peaks(71,13, 70,87, 70,81, 70,62, 70,56, 70,36, 70,30, 70,05 CHOH), two peaks (23,57 and 21,98 CH2), two peaks (9,77 and 9,66 CH3); for 71.5 (CH), 54,03 (CH), 25,5 (CH2), 11,05 (CH3).

19F NMR δ -76,2 and -77,5.

3-Amino-1,1,1-triterpene-2-ol

1,1,1-rifter-3-nitropentane-2-ol (3 g, 16 mmol) was dissolved in methanol (40 ml). Added Nickel Raney catalyst and the reaction mixture is vigorously stirred in a hydrogen atmosphere for 24 hours. The catalyst was filtered and the filtrate was concentratedin the vacuumwith obtaining relatively pure amine (2.35 g, 94%).

δH(CDCl3, 500 MHz) of 1.05 (3H, t, J=5, CHCH2CH3), 1,55-of 1.45 (1H, m, CHCHHCH3), to 1.8-1.7 (1H, m, CHCHHCH3), 2,95 and 2.9 (1H, m, CHEt), 3,95-of 3.85 (1H, m, CHOHCF3); δc (CDCl3, 500 MHz) 124,5 (C), for 71.5 (CH), 54,03 (CH), 25,5 (CH2), 11,05 (CH3).

1,1,1-Cryptor-3-(9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol [16]

To a stirred solution of (2-fluoro-9-isopropyl-N-purine-6 - yl)pyridine-3-ylmethylamino (300 mg, 1.05 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (1.4 ml, 10 EQ., 8 mmol), followed by addition of 3-amino-1,1,1-triterpene-2-ol (1 g, 6.4 mmol) [obtained by the reaction of ammonia with 2-(trifluoromethyl)oxirane]. The flask was supplied reflux, the reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 72 hours. The reaction mixture was allowed to cool to room temperature and the solvent evaporatedin the vacuum. The residue was distributed between EtOAc (50 ml) and water (50 ml), the aqueous phase was dopolnitelnaya EtOAc (2×25 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified gradient column flash chromatography on silica gel with elution CHCl3:MeOH(100:0→95:5), obtaining specified in the title compounds as a colorless film (42 mg, 9.4 per cent).

δH(CDCl3, 500 MHz) was 1.04 (3H, t, J=7,5, CHCH2CH3). of 1.56 (6H, d, J=7,5, CH[CH3]2), 1,8-of 1.73 (2H, m, CHCH2CH3), 4,13-4,07 (1H, m, CHMe2), 4,22-4,2 (1H, m, CHEt) 4,57-4,55 (1H, m, CHOHCF3), 4,71 (2H, s, of user., NHCH2Ar), to 5.08 (1H, s, of user., NH), 7,2 (1H, DD, J=5, 10 ArH), 7,51 (1H, s, ArH), to 7.64 (1H, d, J=10, ArH), 8,46 (1H, d, J=5, ArH), 8,55 (1H, s, ArH); δC(CDCl3, 500 MHz) 159,28 (C), 154,72 (C), 149,07 (CH), 148,47 (CH), 135,44 (CH), 134,83 (C), 134,48 (CH), 124,12 (C), 123,44 (CH), 114,62 (C)73,1 (CH), 55,88 (CH), 46,70 (CH), 41,78 (CH2), 23,68 (CH2) 22,36 (2×CH3), 11,55 (CH3);

19F δ -74,83; m/z 424,2 (M+H).

Example 17

1,1,1-Cryptor-3-(9-isopropyl-6-((6-(trifluoromethyl)pyridin-3-yl)methylamino)-N-2-ylamino)pentane-2-ol [17]

To a stirred solution of 2-fluoro-9-isopropyl-N-(6-(triptorelin-3-ylmethyl)-N-purine-6-amine (200 mg, of 0.56 mmol) in n-BuOH/DMSO (5 ml, 4:1) at room temperature in an argon atmosphere was added DIEA (1.4 ml, 10 EQ., 8 mmol), followed by addition of 3-amino-1,1,1-triterpene-2-ol (0,377 g, 2.4 mmol). The flask was supplied reflux, the reaction mixture was placed in a preheated oil BA is th at 140°C and stirred at this temperature for 72 hours. After the specified time, the reaction took place only at 30%. Spent adding amerosport within 4 days to complete the conversion. The reaction mixture was allowed to cool to room temperature and the solvent evaporatedin the vacuum. The residue was distributed between EtOAc (50 ml) and water (50 ml), the aqueous phase was additionally extracted with EtOAc (2×25 ml) and the combined organic phase was washed with saturated salt solution (50 ml), dried (MgSO4) and evaporatedin the vacuum. The residue was purified gradient column flash chromatography on silica gel with elution CHCl3:MeOH(100:0→95:5), obtaining specified in the title compound as a brown powder (30 mg, 11%).

δH(CDCl3, 500 MHz) of 0.95 (3H, t, J=7,5, CHCH2CH3), and 1.56 (6H, d, J=7,5, CH[CH3]2), 1,721-to 1.61 (2H, m, CHCH2CH3), 3,22 was 3.05 (1H, m, CHEt), 3,88 (1H, m, CHMe2), 4,75-of 4.57 (1H, m, CHCF3OH), 4,92 was 4.76 (2H, m, NHCH2Ar), a 5.3 (1H, s, of user., NH), 6,83 (1H, s, of user., NH), 7,55 (1H, d, J=10, ArH), to 7.61-7,53 (2H, m, ArH), cent to 8.85 (1H, s, ArH); δC(CDCl3, 250 MHz) 157,67 (C), 156,65 (C), 156,9 (C), 146,62 (CH), 139,55 (CH), 128,22 (C), 126,84 (C), 125,97 (CH), 123,14 (CH), 120,32 (C), 71,12 (CH), 61,56 (CH2), 56,43 (CH), 25,74 (CH2), are 22.42 (2×CH3), to 11.56 (CH3);19F δ -67,87, -74,30; ESMS 492 (M+1).

Example 18

1,1,1,3,3,3-Hexamer-2-((9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)methyl)propane-2-ol [18]

To a stirred solution of 2-fluoro-9-isopropyl-6-[(pyridine-3-ylmethyl)amino]purine (50 mg, 0,175 mmol) in n-BuOH/DMSO (2.5 ml, 4:1) at room temperature and in an atmosphere of argon was added diisopropylethylamine (135 mg, 1.05 mmol), followed by the addition of 2-(aminomethyl)-1,1,1,3,3,3-hexaferrite-2-ol obtained by the reaction of 30% ammonium hydroxide solution with 2,2-bis[(trifluoromethyl)oxirane]. The reaction mixture was placed in a preheated oil bath at 140°C and stirred at this temperature for 3 days. The reaction course was monitored using IHMS, and it was only 30%. Added newly obtained 2-(aminomethyl)-1,1,1,3,3,3-hexaferrite-2-ol, and the reaction continued. This addition was performed for three consecutive days to achieve full completion. After removal of solvent the residue was purified column flash chromatography, obtaining specified in the title compounds as a pale brown powder (19 mg, 23%).

δH(CDCl3, 500 MHz) of 1.57 (6H, d, J=7,5, CH[CH3]2), 3,48-3,44 (1H, m, NHCHHSON[CF3]2, 3,86-3,81 (1H, m, NHCHHCOH[CF3]2, 4,62-4,555 (1H, m, CHMe2), 4,78-4,74 (2H, m, NHCH2ArH), to 5.35 (1H, s, of user., OH), 6,36 (1H, s, of user., NH), 7,27-7,22 (2H, m, ArH), 7,7-to 7.67 (1H, m, ArH), and 8.5 (1H, d, J=5, ArH), 8,66 (1H, s, ArH); δC(CDCl3, 500 MHz) 159,99 (C), 154,63 (C), 149,53 (C), 149,05 (C), of 148.6 (CH), 137,79 (C), 135,41 (C), 135,38 (CH), 135 (CH), 123,44 (H), 75,03 (C), 47,62 (CH2), 46,84 (CH), 42,05 (CH2), 22,1 (2×CH3); m/z 464,2 (M+H).

Analysis kinases

Total estimated 21 connection in vitro recombinant kinases and tumor cells and compared with seliciclib. Most of these cdk inhibitors, as found, are more powerful than seliciclib.

To assess the activity of these compounds against protein kinases in vitro, they were skanirovali against CDK2 and CDK9. Analysis kinases were performed in 96-well tablets using recombinant CDK-tsiklonov obtained in Cyclacel Ltd. Dundee, UK. Analyses of CDK2 and CDK9 were performed in a total volume of 25 ál in buffer for analysis (25 mm b-glycerol, 20 mm MOPS, 5 mm EGTA, 1 mm DTT and 1 mm NaVO3, pH 7.4), which was added 2-4 μg of the active enzyme with the appropriate substrates (purified histone H1 for CDK2/cyclina E and CDK2/cyclina And, biotinyl-Ahx-(YSPTSPS)4for CDK9/cyclina T1). The reaction was initiated by adding a mixture of Mg/ATP (15 mm MgCl2+100 μm ATP with 30-50 kBq per well of [-32P]-ATP)and mixtures were incubated for 15 min (CDK2/cyclin E), 30 min (CDK2/cyclin A) or 45 min (CDK9/cyclin T1), as necessary, at 30°C). Reaction with CDK was stopped by adding 25 μl of 75 mm phosphoric acid, followed by filtration through filter plate D81 (Whatman Polyfiltronics, Kent, UK). Against CDK9 reaction was stopped by adding 25 μl of 75 mm phosphoric acid, and then 5 µl is astora avidin 10 mg/ml in each well and additionally incubated for 2 min followed by filtration, as in the analysis of CDK2. After washing 3 times with 75 mm phosphoric acid tablets were dried, was added to scintillation fluid and incorporated radioactivity was assessed by scintillation counter (TopCount, Packard Instruments, Pangbourne, Berks, UK). Connections for kinase assays were prepared as a 10 mm source of standardized solutions in DMSO and diluted to 10% solution of DMSO in buffer for analysis. Data were analyzed using software to adjust the curves (Xlfit, Version 4.00, ID Business Solutions Ltd, Guildford, Surrey, UK) to determine the IC50(concentration of a compound that inhibits the kinase activity by 50%). Average of two indicators can be found in tables 2 and 4. The results show that many of the compounds are more active inhibitors of CDK2 and CDK9 than a kindred connection seliciclib.

The definition of IC50on line tumor cells H460 NSCLC using analysis of cytotoxicity with Alamar Blue

To determine the activity of compounds in the cells, the cytotoxicity of each compound was determined in relation to the cell line non-small cell lung cancer (NSCLC) N. Standard methods for the determination of cytotoxicity was performed as follows: cells N were seeded in 96-well plates in accordance with their doubling time (3000 cells per well) in RPMI medium containing 10% FCS, and incu is Aravali over night at 37°C and 5% CO 2. The medium was removed and added to 100 μl of fresh medium containing concentrations of the respective compounds, and cells were incubated for 72 hours at 37°C and 5% CO2. 10% standardized solution of alamar blue (Roche, Lewes, United Kingdom) were prepared in medium and 100 μl was added to the cells, which were incubated for 2 hours. Absorption was measured on polyatresia (multi-label counter Wallac Victor 2 1420 when 544-595 nm. Average of three independent experiments are presented in table 3.

Two compounds have values IC50less than 1 µm (connection[1]and the connection[11]).

Comparison of the mode of action of the compounds of the present invention and seliciclib

Previously it was shown that seliciclib triggers apoptosis through its effects on transcription. Thus, inhibition of CDK7 and CDK9, which are responsible for the phosphorylation of RNA polymerase II, which is required for the initiation and continuation of transcription. As a result of inhibition of transcription levels of several proteins with a short half-life of decline, such as Mcl-1, starting, thereby apoptosis.

To confirm that these compounds also cause lower regulation of Mcl-1, cells N were seeded at 5×105cells on 10 cm2Cup with 10 ml of RPMI medium containing 10% FCS, and incubated overnight at 37°C and 5% CO2. Then to the cells was added 1 m is 11× concentrated compound, and incubated for 5 or 24 hours. The medium was removed and the attached cells were washed in 5 ml PBS. Cells were literally on cups by adding 100 μl of buffer for lysis (50 mm HEPES) pH 7,20 mm NaCl, 1 mm DTT and a cocktail of protease inhibitors (1:1000) and phosphatase inhibitors (10 mm sodium pyrophosphate, 10 mm sodium fluoride and 1 mm orthovanadate sodium). Lysates were rapidly frozen in liquid nitrogen and stored at -70°C. Frozen lysates were thawed and processed by the ultrasound pulses 2×10 seconds on ice. The protein concentration in each lysate was determined using a kit for determining protein BCA (Pierce) according to manufacturer's instructions. Lysate (30 µg) were mixed with 1× buffer for loading on the gel containing 10% β-mercaptoethanol, and were separated in 4-12% bis-Tris polyacrylamide gel using denaturing electrophoretic conditions (Invitrogen, Glasgow, United Kingdom). Proteins were transferred to nitrocellulose membranes (Shleicher &Schuell, Dassel, Germany) using wet electrophoretic transfer. Membranes were stained crimson S to confirm equal loading prior to blocking in 5% solution of nonfat milk in PBS with 0.05% tween-20 (PBSTM) for 2 hours. Membranes were incubated overnight at 4°C with the first antibody (rabbit polyclonal antibodies against Mcl-1, Santa Cruz), diluted 1:1000 PBSTM. Membranes were washed 2×5 minutes followed by washing 2×10 minutes in PBS and 0.05% tween-20 (PBST), and incubated during 1 hour in PBSTM, containing the second antibody conjugated with horseradish peroxidase. Membranes were washed as above and incubated with enhance the chemiluminescence solution (Amersham) and exposed on x-ray film (Amersham).

As expected, seliciclib shows very modest changes, as the highest concentration used in this experiment is close to the value of the IC50for this connection. Connection[1]clearly superior seliciclib by its ability to reduce the levels of Mcl-1 with significant effects observed at 1.5 μm after 24 hours. This is in agreement with its increased activity.

In the second experiment investigated the effect of additional connections on the levels of Mcl-1. In this case, cells In processed 0,5; 1,5; 4.5 and 13.5 μm for 5 hours, at this time the cells were collected for Western blotting analysis. The results are presented in figure 2.

The results show that the compound [1], as can be seen, is the most effective compound of this group is to down-regulation of the levels of Mcl-1 and has an activity greater than the activity seliciclib. However, it is clear that all these compounds cause a decrease in Mcl-1 at concentrations equivalent to about 2-3 times the value of the IC50.

Inhibition of cytochrome P450 (definition IC505 isoforms)

A who

To install inhibited if the connection [1] activity five isoforms of CYP by analyzing the metabolism of specific substrates of CYP. Comparative analyses were performed using the compounds of the prior art [A1], [A2] and [A3], is presented below:

The experimental technique

Compounds [A1], [A2] and [A3] was synthesized in accordance with the methods presented in WO 2004/016612 (Cyclacel Limited).

Inhibition of CYP1A

Liver microsomes person (0.25 mg/ml) and NADPH (1 mm) were incubated with six concentrations of the test compounds(0,05; 0,25; 0,5; 2,5; 5; 25 μm in DMSO; final concentration of DMSO=0,35%) in the presence of the evaluated substrate ethoxyresorufin (0.5 µm) for 5 min at 37°C. Selective inhibitor of CYP1A, alpha naphthoflavone, skanirovali together with test compounds as positive control.

Inhibition of CYP2C9

Liver microsomes person (0.25 mg/ml) and NADPH (1 mm) were incubated with six concentrations of the test compounds(0,05; 0,25; 0,5; 2,5; 5; 25 μm in DMSO; final concentration of DMSO=0,25%) in the presence of the evaluated substrate tolbutamide (120 μm) for 60 min at 37°C. Selective inhibitor ofCYP2C9sulfaphenazole, skanirovali together with test compounds as positive control.

Inhibition of CYP2C19

Microsome assay pecen of man (0.25 mg/ml) and NADPH (1 mm) were incubated with six concentrations of the test compounds (0,05; 0,25; 0,5; 2,5; 5; 25 μm in DMSO; final concentration of DMSO=0,25%) in the presence of the evaluated substrate mephenytoin (25 μm) for 60 min at 37°C. Selective inhibitor ofCYP2C19, tranilcipromin, skanirovali together with test compounds as positive control.

Inhibition of CYP2D6

Liver microsomes person (0.25 mg/ml) and NADPH (1 mm) were incubated with six concentrations of the test compounds(0,05; 0,25; 0,5; 2,5; 5; 25 μm in DMSO; final concentration of DMSO=0,25%) in the presence of the evaluated substrate dextromethorphan (5 μm) for 30 min at 37°C. Selective CYP2D6 inhibitor, quinidine, skanirovali together with test compounds as positive control.

Inhibition of CYP3A4

Liver microsomes person (0.25 mg/ml) and NADPH (1 mm) were incubated with six concentrations of the test compounds(0,05; 0,25; 0,5; 2,5; 5; 25 μm in DMSO; final concentration of DMSO=0,26%) in the presence of the evaluated substrate midazolam (2.5 μm) for 5 min at 37°C. Selective inhibitor ofCYP3A4, ketoconazole, skanirovali together with test compounds as positive control.

For incubation with CYP1A reaction was stopped by adding methanol, and the formation of the metabolite, resorufin, was monitored by fluorescence (excitation wave=535 nm wave radiation=595 nm). For incubation withCYP2C9,CYP2C19, CYP2D6, andCYP3A4the reaction was interrupted by adding containing methanol internal standard. The samples are then centrifuged and the supernatant was combined for the simultaneous analysis of 4-hydroxytryptamine, 4-hydroxymephenytoin, dextromethorphan and 1 hydroxymidazolam plus internal standard using LC-MS/MS using typical conditions for LC-MS/MS. In the final sample before analysis was added formic acid in deionized water (final concentration=0.1%of). Reduced formation of metabolites in comparison with control media used to calculate the values of the IC50(concentration of test compound which gives 50% inhibition).

Results

IC50(μm) for each connection in respect of the five isoforms of CYP presented in table 5.

Data show that the three compounds are strong inhibitorsCYP3A4while the connection [1] is not. As the value of the IC50connections [1] is significantly higher than its IC50for cells (see, table 7), this shows that the cytotoxic concentration there is not expected to impact on activityCYP3A4. This is important, becauseCYP3A4involved in the metabolism of a large number of drugs. IfCYP3A4inhibited one drug, it can lead to unexpected toxicity due to reduced metabolism of substratesCYP3A4, leading, thus, to clearly p is increased levels of these funds.

Identification of the substrate of the cytochrome P450

Purpose

To determine which of the major cytochrome P450 isoforms involved in the metabolism of four of the tested compounds.

The experimental technique

Preparations of human CYP450 enzyme, expressed cDNA co-expressed with human NADPH-cytochrome-P450-reductase (Bactosomes™), were delivered Cypex Ltd. Bactosomes™ (the final concentration of CYP1A2 100 pmol/ml, CYP2C8 50 pmol/ml,CYP2C925 pmol/ml,CYP2C19100 pmol/ml, CYP2D6 50 pmol/ml andCYP3A425 pmol/ml)in 0.1m phosphate buffer pH 7.4 and the test compound (final substrate concentration=5 μm; final concentration of DMSO=0,25%) pre-incubated at 37°C before the addition of NADPH (final concentration=1 mm) to initiate the reaction. Incubation was performed using the control lactosome (in the absence of P450 enzymes)in order to identify any non-enzymatic decomposition. The final volume of incubation was 25 µl. Connection known that they specifically each metabolized by CYP450 isoform, was used as control compounds.

Each connection incubated separately for 0, 5, 15, 30 and 45 min with each of the CYP isoforms. Reactions were stopped by adding containing methanol internal standard at the appropriate time. Tablets for incubation was centrifuged at 2500 rpm is in for 20 min at 4°C to precipitate the protein. After deposition of protein supernatant was combined in a cassette, comprising up to 4 connections, and analyzed using typical conditions LC-MS/MS.

Data analysis

The relationship of the peak area ln (which was adjusted for any loss during incubation with control lactosome) were placed on the schedule according to the time, and determined the gradient of the line.

The rate constant of elimination (k)=(-gradient)

The half-life(t1/2)(min)=0,693/k

Results

The half-life (minutes) for each connection in the presence of each of the six CYP presented in table 6.

Data show that in this system Bactosomes™ connection [1] is not a substrate for the six tested CYP isoforms. There is a big difference with the other three compounds, since they are all substratesCYP3A4and the two also are substrates of CYP1A2. This difference agrees well with the difference of inhibition of CYP discussed in table 4. A common mechanism leading to inhibition of CYP exists if the connection is also a substrate for this CYP. As you can see, the compound [1] is neither a substrate nor an inhibitor ofCYP3A4while the other three compounds are them.

Analysis kinases

To assess the activity of these compounds against protein kinases in vitro, they were skanirovali agains the CDK2 and CDK9. Analysis kinases were performed in 96-well tablets using recombinant CDK-tsiklonov obtained in Cyclacel Ltd. Dundee, Uk. Analyses with CDK2 and CDK9 were performed in a total volume of 25 µl of buffer for analysis (25 mm b-glycerol, 20 mm MOPS, 5 mm EGTA, 1 mm DTT and 1 mm NaVO3, pH 7.4), which was added 2-4 μg of the active enzyme with the appropriate substrates (purified histone H1 for CDK2/cyclina E and CDK2/cyclina And, biotinyl-Ahx-(YSPTSPS)4for CDK9/cyclina T1). The reaction was initiated by adding a mixture of Mg/ATP (15 mm MgCl2+100 μm ATP with 30-50 kBq per well of [-32P]-ATP)and mixtures were incubated for 15 min (CDK2/cyclin E), 30 min (CDK2/cyclin A) or 45 min (CDK9/cyclin T1), as necessary, at 30°C). Reaction with CDK was stopped by adding 25 μl of 75 mm phosphoric acid, followed by filtration through filter plate D81 (Whatman Polyfiltronics, Kent, UK). Against CDK9 reaction was stopped by adding 25 μl of 75 mm phosphoric acid, and then 5 μl of the solution avidin 10 mg/ml in each well and additionally incubated for 2 min followed by filtration, as in the analysis with CDK2. After washing 3 times with 75 mm phosphoric acid tablets were dried, was added to scintillation fluid and incorporated radioactivity was assessed by scintillation counter (TopCount, Packard Instrument, Panbourne, Berks, UK). Connections for analyses kinases were prepared as a 10 mm initial standardized R. the cross-sections in DMSO and diluted to 10% solution of DMSO in buffer for analysis. Data were analyzed using software to adjust the curves (Xlfit, Version 4.00, ID Business Solutions Ltd, Guildford, Surrey, UK) to determine the IC50(concentration of a compound that inhibits the kinase activity by 50%). Average of two indicators can be found in tables 6 and 8.

Results

The results in table 7 show that the connection of the prior art [A3] and the compound [1] are the most active inhibitors of kinases.

The definition of IC50in the lines of tumor cells using analysis of cytotoxicity with Alamar Blue

To determine the activity of these compounds in the cells, the cytotoxicity of each compound is determined in respect of a number of cell lines. Standard methods for determination of cytotoxicity was performed as follows: cells were seeded in 96-well plates in accordance with their time of doubling (2-5000 cells per well) in medium RPMI or DMEM containing 10% FCS, and incubated overnight at 37°C and 5% CO2. The medium was removed and added to 100 μl of fresh medium containing increasing concentrations of the respective compounds, and cells were incubated for 72 hours at 37°C and 5% CO2. 10% standardized solution of alamar blue (Roche, Lewes, United Kingdom) were prepared in medium and 100 μl was added to the cells, which were incubated for 2 hours is. Absorption was measured on polyatresia (multi-label counter Wallac Victor 2 1420 when 544-595 nm.

Results

The results of the analysis at the cellular cytotoxicity against 21 cell line are presented in table 8. Connection [1] is significantly more active than compounds [A1], [A2] and [A3] prior art.

Various modifications and variations of the present invention, without departing from the essence and scope of the invention will be obvious to experts in this field. Although the present invention has been described in connection with specific preferred variant implementation, it will be clear that the invention claimed should not be unduly limited to such specific choices of implementation. Of course, various modifications of the described methods of implementation of the present invention which are obvious to experts in this field, as implied, are covered by the invention.

Table 1
Selected compounds of the present invention
Connection.StructureName
[1](2R,3S-3-(6-(4,6-dimethylpyridin-3-ylmethylamino)-9-isopropyl-n-purine-2-ylamino)pentane-2-ol
[2]2R,3S-3-(9-isopropyl-6-((6-methylpyridin-3-yl)methylamino)-n-purine-2-ylamino)pentane-2-ol

[3](2R,3S-3-(6-(3-chlorobenzylamino)-9-isopropyl-n-purine-2-ylamino)pentane-2-ol
[4](2R,3S-3-(6-(3-forbindelsen)-9-isopropyl-n-purine-2-ylamino)pentane-2-ol
[5]2R,3S-3-(9-(cyclopropylmethyl)-6-(pyridine-3-ylmethylamino)-n-purine-2-ylamino)pentane-2-ol
[6]2R,3S-3-(6-cyclopropylamino)-9-isopropyl-n-purine-2-ylamino)pentane-2-ol
[7]2R,3S-3-(6-(cyclopropylamino)-9-isopropyl-n-purine-2-ylamino)pentane-2-ol

[8] 2R,3S-3-(6-(cyclobutylamine)-9-isopropyl-n-purine-2-ylamino)pentane-2-ol
[9]2R,3S-3-(9-isopropyl-6-(pyridine-4-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol
[10]2R,3S-3-(9-isopropyl-6-(2,6-dimethylpyridin-4-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol
[11]2R,3S-3-(9-isopropyl-6-((6-trifluoromethyl)pyridin-3-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol
[12]2R,3S-3-(9-isopropyl-6-((6-methylpyridin-2-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol
[13]2R,3S-3-(9-isopropyl-6-((3-methylpyridin-2-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol

[14](R)-1-(9-isopropyl-6-(feast of the Dean-3-ylmethylamino)-N-purine-2-ylamino)propan-2-ol
[15]1,1,1-Cryptor-3-(9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)propan-2-ol
[16]1,1,1-Cryptor-3-(9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)pentane-2-ol
[17]1,1,1-Cryptor-3-(9-isopropyl-6-((6-(trifluoromethyl)pyridin-3-yl)methylamino)-N-2-ylamino)pentane-2-ol
[18]1,1,1,3,3,3-hexamer-2-((9-isopropyl-6-(pyridine-3-ylmethylamino)-N-purine-2-ylamino)methyl)propane-2-ol

Table 2
Summary table analyses kinase in vitro in choosing compounds of the second generation and seliciclib. The values of the IC50expressed in microns. Compounds were analyzed in two games, and in both cases seliciclib took the test as a control, representing seliciclib 1 and 2, which gave similar results
Connection CDK2/Cyclin ECDK2/cyclin aCDK9/Cyclin T1
SDSDSD
Seliciclib 10,420,081500,112 030,35
[1]0,020,0010,090,000,100,02
[2]0,520,153,251,033,630,76
[3]0,010,0040,100,0040,060,00
[4]0,0 0,010,120,010,080,02
Seliciclib 20,280,03NDND2,350,61
[5]0,830,22NDND789NA
[6]0,050,003NDND0,270,03
[7]0,040,01NDND0,380,01
[8]0,160,05NDND0,500,25
[9]0,040,01NDND0,610,01
[10]0,050,02NDND1,040,30
[11]0,040,01NDND0,250,10
[12]0,050,01NDND2,040,44
[13]0,770,06NDNDof 5.841,21
[14]0,290,14NDND6,532,74
[15]0,570,04NDND7,402,81
[16]0,090,01NDND2,140,50
ND - not determined
NA - not analyzed

Table 3
A summary table of the values of the IC50for cells presents connections, which was determined on cells N. The last column shows how many of these compounds are more active than seliciclib
ConnectionAverage IC50(µm)SDThe rate of increase of activity
[1]0,50,024,0
[2]35 5,50,3
[3]1,50,18,0
[4]1,50,18,0
[5]34,43,20,3
[6]1,70,27,1
[7]2,40,75,0
[8]2,90,64,1
[9]2,30,35,2
[10]2,40,15,0
[11]0,90,213,3
[12] 7,20,11,7
[13]19,45,30,6
[14]16,72,30,7
[15]20,35,20,6
[16]4,10,62,9
Seliciclibto 12.02,2NA

Table 4
Summary table analyses kinase screening compounds [17] and [18] (µm)
CDK2ECDK2ACDK1B
Connection.Series 1Series 2AVG. SDSeries 1Series 2AVG.SDSeries 1Series 2AVG.SD
[17]0,3180,3070,3130,0080,3050,4310,3680,0896,12111,2358,6783,616
[18]0,3360,2790,3080,0400,7041,1940,9490,3475,8826,7256,3030,596
CDK4D1 CDK7HCDK9T1
Connection.Series 1Series 2AVG.SDSeries 1Series 2AVG.SDSeries 1Series 2AVG.SD
[17]>10>10>101,8511,7771,8140,0532,0465,4263,7362,390
[18]>10>10>105,0456,0095,5270,682 3,9543,2103,5820,526

Table 5
IC50(M) for compounds [A1], [A2] and [A3] prior art and compounds [1] of the present invention in respect of the five isoforms of CYP
CYP1ACYP2C19CYP2C9CYP2D6CYP3A4
[Al]>25to 19.9>25>251,63
[A2]>25>25>25>250,48
[A3]>25>25>25>251,88
[1]>25>25the 3.8 >2522,5

Table 6
The half-life (min) for compounds [A1], [A2] and [A3] prior art and compounds [1] of the present invention in the presence of each of the six CYP
CYP1A2CYP2C19CYP2C8CYP2C9CYP2D6CYP3A4
[Al]29,258,3>45>45>4528,3
[A2]49,5>45>45>45>4532,6
[A3]>45>45>45>45>4518,0
[1]>45>45>45>45>45>45

Table 7
Activity against inhibition of kinases (IC50, M) compounds [A1], [A2] and [A3] prior art and compounds [1] of the present invention
Kinase[A1][A2][A3][1]
CDK2A0,370,200,110,04
CDK2E0,130,070,010,02
CDK9T10,340,280,090,10

Table 8
The results of the analysis at the cellular cytotoxicity of compounds [A1], [A2] and [what 3] prior art and compounds [1] of the present invention against various cell lines
[A1][A2][A3][1]
H1650of 6.49the 3.651,220,46
B16to 7.772,590,66
HeLa6,643,30
MDA-MB-4366,63,380,970,44
H20525,162,360,940,29
LoVo4,502,200,820,70
Saos-25,312,481,40
CT26.WT6,69 4,881,63
H2926,562,340,910,37
Colo2054,482,310,820,31
HT-294,151,701,17
NCI-H4602,80of 2.210,700,50
LP-11,640,47
A5492,951,600,470,16
MESSA3,621,590,500,16
MESSA-Dx514,69charged 8.52a 3.87HCT 116of 1.570,44
MCF7the 3.651,640,450,26
NCI-H9295,162,350,790,35
A27802,601,100,380,41
H3582,690,840,330,18
Average4,88to 2.290,790,35

1. The compound of formula (I) or its pharmaceutically acceptable salt

where R1and R2, each independently, represents H, C1-6-alkyl or C1-6-halogenated;
R3and R4, each independently, represents H, C1-6-alkyl or C1-6-halogenated;
R5represents a C1-6-alkyl or the 3-12-cycloalkyl or3-12-cycloalkyl-C1-6-alkyl, each of which may be optionally substituted by one or more Oh groups;
R6represents a

where Y is N, X and Z are CR9;
R7, R8and each R9independently represent H, C1-6-alkyl or C1-6-halogenated; where at least one of R7, R8and R9is not N.

2. The compound according to claim 1, where one of R1and R2is N, and the other is With1-6-alkyl.

3. The compound according to claim 1, where one of R1and R2is N, and the other is stands, ethyl or isopropyl.

4. The compound according to claim 1, where R1is ethyl, and R2is N.

5. The compound according to claim 1, where R3and R4, each independently, represents H, C1-6-alkyl or C1-6-halogenated, and where at least one of R3and R4is not N.

6. The compound according to claim 1, where one of R3and R4is N, and the other is With1-6-alkyl or C1-6-halogenation.

7. The compound according to claim 1, where R3is N, and R4is C1-6-alkyl or C1-6-halogenation.

8. The compound according to claim 1, where R3is N, and R4is stands.

9. The compound according to claim 1, where X is CH, Z is one the-IU, and R7is N, and R8is Me.

10. The compound according to claim 1, where X is CH, Z is C-IU, and R7and R8both are N.

11. The compound according to claim 1, where X is CH, Z is C-CF3and R7and R8both are N.

12. The compound according to claim 1, where R5is isopropyl.

13. The compound according to claim 1, which is chosen from the following:

[1](2R,3S-3-(6-((4,6-dimethylpyridin-3-ylmethylamino)-9-isopropyl-9H-purine-2-ylamino)pentane-2-ol
[2]2R,3S-3-(9-isopropyl-6-((6-methylpyridin-3-yl)methylamino)-9H-purine-2-ylamino)pentane-2-ol
[11]2R,3S-3-(9-isopropyl-6-((6-trifluoromethyl)pyridin-3-yl)methylamino)-N-purine-2-ylamino)pentane-2-ol
[17]1,1,1-Cryptor-3-(9-isopropyl-6-((6-(trifluoromethyl)pyridin-3-yl)methylamino)-N-2-ylamino)pentane-2-ol

14. The compound (2R,3S-3-(6-((4,6-dimethylpyridin-3-ylmethylamino)-9-isopropyl-N-purine-2-ylamino)pentane-2-ol or its pharmaceutically acceptable salt or ester.

15. Pharmaceutical composition having the properties of an inhibitor of CDK enzyme selected from CDK1, CDK2, CDK3, CDK4, CDK6, CDK7, CDK8 and CDK9, containing a therapeutically effective amount is about connection according to claim 1 in a mixture with a pharmaceutically acceptable diluent, excipient or carrier, or mixtures thereof.

16. The use of compounds according to claim 1 for obtaining a medicinal product for the treatment of proliferative diseases, mediated by the activity of the CDK enzyme selected from CDK1, CDK2, CDK3, CDK4, CDK6, CDK7, CDK8 and CDK9.

17. The application of article 16, where the specified proliferative disease is cancer or leukemia.

18. The application of article 16, where the proliferative disease is glomerulonephritis, rheumatoid arthritis or psoriasis.

19. The use of compounds according to claim 1 for obtaining a medicinal product for the treatment of viral diseases, mediated by the activity of the CDK enzyme selected from CDK1, CDK2, CDK3, CDK4, CDK6, CDK7, CDK8 and CDK9, where a viral disease selected from diseases caused by human cytomegalovirus (HCMV), herpes simplex virus type 1 (HSV-1), human immunodeficiency virus type 1 (HIV-1) and varicella zoster virus and herpes zoster (VZV).

20. The use of compounds according to claim 1 for obtaining a medicinal product for the treatment of diseases of the Central nervous system, mediated by the activity of the CDK enzyme selected from CDK1, CDK2, CDK3, CDK4, CDK6, CDK7, CDK8 and CDK9, where disease of the Central nervous system is Alzheimer's disease.

21. The use of compounds according to claim 1 for obtaining a medicinal product for the treatment of hair loss.

22. The use of compounds according to claim 1 for obtaining a medicinal product for the treatment of shock.

<> 23. The application of article 16, where this compound is used in an amount sufficient to inhibit at least one CDK enzyme.

24. The use of compounds according to claim 1 for the treatment of neurodegenerative disease mediated by the activity of the CDK enzyme selected from CDK1, CDK2, CDK3, CDK4, CDK6, CDK7, CDK8 and CDK9.

25. The application of paragraph 24 for the treatment of neuronal apoptosis.

26. The use of compounds according to claim 1 for the inhibition of protein kinase, which is a cyclin-dependent kinase selected from CDK1, CDK2, CDK3, CDK4, CDK6, CDK7, CDK8 and CDK9.

27. A method of treating a proliferative disease mediated by the activity of the CDK enzyme selected from CDK1, CDK2, CDK3, CDK4, CDK6, CDK7, CDK8 and CDK9, including introduction to the mammal a therapeutically effective amount of a compound according to claim 1.

28. The method according to item 27, where the specified connection is used in a quantity sufficient to inhibit at least one CDK enzyme.

29. The compound according to claim 1 for use in medicine for the treatment of a disease mediated by the activity of the CDK enzyme selected from CDK1, CDK2, CDK3, CDK4, CDK6, CDK7, CDK8 and CDK9.

30. The compound according to claim 1 for the treatment of a disease mediated by the activity of the CDK enzyme selected from CDK1, CDK2, CDK3, CDK4, CDK6, CDK7, CDK8 and CDK9, where the disease is selected from proliferative diseases, viral diseases, neurodegenerative diseases, disease is Central nervous system, hair loss and shock.



 

Same patents:

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to new purine derivatives of formula (I) and to their pharmaceutically acceptable salts exhibiting the properties of adenosine receptor A2A agonists. The compounds can find application for preparing a drug for treating an inflammatory or obstructive respiratory disease. In formula

,

R1, R2 and R3 are those as specified in the patent claim.

EFFECT: preparing new purine derivatives of formula (I) or their pharmaceutically acceptable salts showing the properties of adenosine receptor A2A agonists.

8 cl, 2 tbl, 264 ex

FIELD: chemistry.

SUBSTANCE: invention relates to an improved method of producing abacavir of formula (I) or salts or solvates thereof. Abacavir has strong HIV-1 and HIV-2 selective inhibitor activity and can be used in treating patients infected with HIV. The method involves i) closing the ring in a compound of formula (IV) by first reacting said compound (IV) with an anhydrous hydrochloric acid solution in (C1-C6)-alcohol, preferably isopropanol, and then with tri(C1-C4)-alkylorthoformate in anhydrous conditions to obtain a compound of formula (III), ii) reaction of the compound of formula (III) with cyclopropylamine to obtain a compound of formula (II) and iii) hydrolysis of the compound of formula (II) to obtain abacavir (I) or salt thereof. R1 denotes a (C1-C4)-alkyl radical, preferably isopropyl.

EFFECT: obtaining an end product with higher output and higher quality.

12 cl, 6 ex

FIELD: chemistry.

SUBSTANCE: nucleic base (e.g. uracil, cytosine, adenine, guanine, hypoxanthine, xanthine or similar) reacts with perfluoroalkyl halide in the presence of sulphoxide, peroxide and an iron compound to obtain a perfluoroalkyl-substituted nucleic base.

EFFECT: high cost effectiveness as an intermediate compound for producing medicinal agents.

15 cl, 6 tbl

FIELD: chemistry.

SUBSTANCE: invention relates to an improved method of producing abacavir of formula (I) or pharmaceutically acceptable salt thereof, involving reaction of a compound of formula (II), (I) (II), where R denotes H or (C1-C4)-alkyl radical with an inorganic base such as an alkali metal hydroxide in a mixture of (C1-C6)-alcohol and water; and extraction of abacavir of formula (I) in form of a free base or in form of a pharmaceutically acceptable salt by treating said free base with a corresponding acid.

EFFECT: method ensures high degree of conversion without racemation, enables to minimise formation of impurities and considerably shortens reaction time.

19 cl, 10 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention relates to novel compounds of formula I in free form or in form of pharmaceutically acceptable salt, which possess properties of adenosine receptor A2A agonists. In formula I , R1 represents (C1-C8)alkylcarbonyl, (C3-C8)cycloalkylcarbonyl, -SO2(C1-C8)alkyl, phenyl(C1-C4)alkylcarbonyl or -(C=O)-C(=O)-NH(C1-C8)alkyl, optionally substituted with R4; R2 represents H or (C1-C8)alkyl, optionally substituted with (C6-C10)aryl; R3 represents halogen or(C2-C8)alkinyl, or R3 stands for aminogroup, optionally substituted with (C3-C8)cycloalkyl, optionally substituted with amino, or R3 represents (C1-C8)alkylaminogroup, optionally substituted with hydroxy, phenyl or R5, or R3 stands for R6, optionally substituted with amino or -NH-C(=O)-NH-R7, or R3 stands for -NH-R6, optionally substituted with -NH-C(=O)-NH-R7, or R3 stands for (C1-C8)alkylaminocarbonyl, optionally substituted with. -NH-C(=O)-NH-R8; R4, R5 and R6 represent independently 5- or 6-member heterocyclic ring, which contains one-two N ring heteroatoms, optionally substituted with amino or (C1-C8)alkyl; and R7 and R8 represent independently 5- or 6-member heterocyclic ring, which contains one-two ring heteroatoms selected from N and S, and is optionally substitutedf with halogen, (C1-C8)alkylsulfonyl or 5- or 6-member aromatic heterocyclic ring, which contains one N ring heteroatom. Invention also relates to pharmaceutical composition and to application of said compounds for treatment of states, mediated by activation of adenosine receptor A2A.

EFFECT: obtaining composition, which possesses properties of adenosine receptor A2A agonists.

10 cl, 3 tbl, 80 ex

FIELD: chemistry.

SUBSTANCE: invention relates to novel purine derivatives of general formula I in free form or in form of a pharmaceutically acceptable salt which have A2A agonist properties. In formula I , R1 denotes a N-bonded 5-6-member heterocyclic group containing 1-4 nitrogen atoms in the ring, which can be optionally substituted with oxo, phenyl or C1-8-alkyl, optionally substituted with hydroxy; R2 is hydrogen or C1-C8-alkyl, optionally substituted with hydroxy or 1-2 phenyls possibly substituted with hydroxy or C1-C8-alkoxy; R3 is C2-C8-alkynyl or C1-C8-alkoxycarbonyl, or R3 is amino substituted with C3-C8-cycloalkyl, optionally substituted with amino, hydroxy, benzyloxy or NH-C(=O)-NH-R6, or R3 is amino substituted with R4, -R4-benzyl or C5-C10-mono- or bicarbocyclic group, optionally substituted with hydroxy or C1-C8-alkoxycarbonyl, or R3 is aminocarbonyl optionally substituted with R5, or R3 is C1-C8-alkylamino optionally substituted with hydroxy, R5, NH-C(=O)-C1-C8-alkyl, -MH-SO2-C1-C8-alkyl, -NH-C(=O)-NH-R6 or phenyl, optionally substituted with phenyloxy, or R3 is a N-bonded 5-member heterocyclic group containing 1 nitrogen atom in the ring which may optionally be substituted with amino, C1-C8-alkylamino, di(C1-C8-alkyl)amino and other groups.

EFFECT: compounds can be useful in treating conditions mediated by activation of the adenosine A2A receptor, especially inflammatory or obstructive respiratory tract diseases.

9 cl, 5 tbl, 161 ex

FIELD: organic chemistry, heterocyclic compounds, biochemistry.

SUBSTANCE: invention relates to new compounds - purine derivatives of the general formula (I): in free form or salt wherein X means oxygen or sulfur atom or group NR5; R1 means alkyl, alkenyl, cycloalkyl, benzocycloalkyl, cycloalkylalkyl or aralkyl group that can be substituted optionally with hydroxy-, carboxy-group or alkoxycarbonyl; or if X means NR5 then R1 can mean alternatively heterocyclic group taken among benzylpiperidyl or the formula: ; or group of the formula (II): ; R2 means hydrogen atom, alkyl or alkoxy-group; R3 means hydrogen atom, alkoxy-, carboxy-group, carboxyalkyl, alkoxycarbonyl, -N(R9)R10, (C1-C4)-alkylene-SO2N(R11)R12 or -CON(R13)R14; or if two substitutes R2 and R3 are joined to adjacent carbon atoms in indicated benzene ring then in common with carbon atoms to which they are joined they mean heterocyclic group comprising 5-10 ring atoms among them one or two atoms mean heteroatoms taken among nitrogen, oxygen and sulfur atom; R4 means hydrogen atom, alkoxy-, carboxy-group, carboxyalkyl, -SO2N(R11)R12, -N(R9)R10 or -CON(R13)R14; or if two substitutes R3 and R4 are joined to adjacent carbon atoms in indicated benzene ring then in common with carbon atoms to which they are joined they mean heterocyclic group comprising 5-6 ring atoms among them one or two atoms mean heteroatoms taken among nitrogen, oxygen or sulfur atom; R5 means hydrogen atom or alkyl; R6, R7 and R8 mean hydrogen atom, or one of these radicals means -SO2NH2, -N(CH3)COCH3, -CONH2 and two others mean hydrogen atom; R9 means hydrogen atom or alkyl; R10 means hydrogen atom, -COR15 wherein R15 means alkyl, alkoxy-group; or R9 and R10 in common with nitrogen atom to which they are joined mean heterocyclic group comprising 5 or 6 ring atoms among them one or two atoms mean heteroatoms taken among nitrogen and oxygen atom; R11 means hydrogen atom or alkyl; R12 means hydrogen atom, alkyl, hydroxyalkyl, carboxyalkyl or alkoxycarbonylalkyl; or R11 and R12 in common with nitrogen atom to which they are joined mean heterocyclic group comprising 5 or 6 ring atoms among them one or two atoms mean heteroatoms taken among nitrogen and oxygen atom; R13 and R14 each and independently of one another means hydrogen atom or alkyl with exception of 2-(para-n-butylanilino)-6-methoxypurine, 2-(para-n-butylanilino)-6-(methylthio)purine, 2,6-di-(phenylamino)-purine, 2,6-di-(para-tolylamino)-purine and 2-(para-tolylamino)-6-(phenylamino)-purine.

EFFECT: valuable biochemical properties of compounds.

11 cl, 4 tbl, 221 ex

The invention relates to novel 2,6,9-triple-substituted purine derivative of General formula I, having the effect of selective inhibitors of kinases of the cell cycle, which can be used, for example, for the treatment of, for example, autoimmune diseases such as rheumatoid arthritis, systemic lupus erythematosus, diabetes type I, multiple sclerosis, and for the treatment of cancer, cardiovascular diseases such as restenosis, etc

The invention relates to new compounds of General formula I

< / BR>
where R is chosen from the group comprising R2, R2NH - or R3R4N-R5-, where R2selected from the group including9-C12-alkyl,

< / BR>
and

< / BR>
where each R6independently selected from the group including hydrogen, C3-C8-cycloalkyl,1-C4-alkyl and (CH2)m-phenyl, where m = integer 0-8; x = 1-8 integer; n = 0-8 integer; z is chosen from the group comprising phenyl, heterocycle, cycloalkyl and naphthalene, and M is chosen from the group comprising hydrogen, C1-C4-alkyl,

< / BR>
and

< / BR>
where each R6' are independently selected from the group including hydrogen, C3-C8-cycloalkyl,1-C4-alkyl and (CH2)m-phenyl, where m' = integer 0-8; n' = integer 0-8; x' = 1-8 integer; Q is hydrogen or C1-C4-alkyl, and Z' is chosen from the group comprising phenyl, heterozygote selected from the group including D, E,

< / BR>
and

< / BR>
where each D is independently selected from the group comprising trifluoromethyl, triptoreline and C1-C4-alkoxy; each E is independently selected from the group including Hal, HE and1-C8-alkyl; Z is chosen from the group comprising phenyl, cycloalkyl and naphthalene; each R6"is hydrogen, n = integer 0-8; x" = 1-8 integer, and M' is hydrogen, Z' may be optionally substituted by groups D', E', each D' is independently selected from the group comprising trifluoromethyl, triptoreline and C1-C4-alkoxy; each E' is independently selected from the group including Hal, HE and1-C8-alkyl; R3and R4selected from the group including hydrogen, C1-C4-alkyl and (CH2)y-phenyl, where y = 0-8 integer, provided that R3and R4both denote hydrogen; R5- C1-C8-alkylene and R1selected from the group including cyclopentyl, cyclopentenyl and isopropyl, and their pharmaceutically acceptable salts, optical isomers and hydrates, provided that when R2refers to a group

< / BR>
< / BR>
< / BR>
and

< / BR>
where D, b, R6", x", n", M' and Z" accept above values

the method of treatment of a patient with proliferative disorders by assigning the compounds I, the method of preventing apoptosis of nerve cells, ways of protecting nerve cells from apoptosis and destruction caused by antitumor agents, and pharmaceutical composition

The invention relates to new derivatives of purine of formula I, II, III and IV, pharmaceutical compositions and method of treatment of a pathological state characterized by thrombotic activity

FIELD: medicine.

SUBSTANCE: invention refers to medicine, namely obstetrics and gynaecology, and may be used for treating early miscarriage in females with primary hyperprolactinemia. That is ensured by prescribing a sparing regimen combined with the administration of dufaston. It is added with prescribing the preparation dostinex 0.125 mg - 0.250 mg at bedtime to a pregnant woman 1-2 times a week to the 10-12th week of pregnancy.

EFFECT: method provides higher clinical effectiveness, reduced side effects and complications.

3 ex

FIELD: chemistry.

SUBSTANCE: invention relates to peptidyl analogues of ghrelin having greater stability which are active with respect to the GHS receptor, having the formula given below: (R2)-A2-A3-A4-A5-A6-A7-A8-A9-A10-A11-A12-A13-A14-A15-A16-A17-A18-A19-A20-A21-A22-A23-A24-A25-A26-A27-A28-Rl, where values of A1-A28, R1 and R2 are given in the description, pharmaceutically acceptable salts thereof and pharmaceutical compositions containing an effective amount of said compound, as well as therapeutic and non-therapeutic applications thereof.

EFFECT: high stability.

22 cl, 3 tbl, 11 ex

FIELD: medicine.

SUBSTANCE: invention relates to medicine, namely to endocrinology and cardiology and deals with treatment of metabolic syndrome. For this purpose complex treatment, which includes antihypertensive diet, individually selected aerobic physical exercise, introduction of metformin in dose 500 mg two days per day and melaxen, is carried out. Melaxen is introduced before going to bed 30 minutes before sleeping in dose 1-2 mg during first two weeks, and then in dose 3 mg during the following ten weeks. Course of treatment constitutes 12 weeks.

EFFECT: complex of non-pharmacological and pharmacological therapy as well as empirically selected mode of melaxen introduction ensure efficient treatment due to essential reduction of body weight and reduction of insulin-resistance in said group of patients.

2 ex

FIELD: medicine.

SUBSTANCE: endothelial dysfunction is diagnosed if observing the twofold increase of a rat's blood glucose level. The technique involves determining antioxidant system values, a concentration of total NO metabolites, eNOS activity, a micro- and macrohaemodynamic status, a MDA concentration and Na+ K+ ATP-ase activity in myocardial and hepatic cell membranes. The values variations enables stating the presence of endothelial dysfunction in alloxan diabetes. Such dysfunction is corrected by the subcutaneous introduction of the preparation Ubiquinone (Coenzyme Q10) 0.11 mcl/100 g of animal's weight once a day for 30 days.

EFFECT: more accurate and reliable diagnosis of the vascular disorders accompanying said type of diabetes, and effective correction of such disorders also provides reproducibility, ease, availability, safety and low cost of performing the experiment.

2 cl, 2 tbl, 6 dwg, 1 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to a pharmaceutical composition showing stress-protective action which contains peptide R1-Lys1-Arg2-Pro3-R2 [SEQ ID NO:1] or R1-Lys1-Arg2-Arg3-Pro4-R2 [SEQ ID NO:2] wherein R1=NH2 or CH3CO and R2=OH or NH2 and a method for prevention and/or treatment functional or stress-induced disorders caused by extreme factors.

EFFECT: producing the pharmaceutical composition exhibiting stress-protective action.

3 cl, 9 tbl, 7 ex

FIELD: chemistry.

SUBSTANCE: invention relates to stable metastin derivatives, having excellent biological activity. The disclosed metastin derivatives improve stability, gelation tendency is reduced, pharmacokinetics are also improved, and an excellent cancer metastasis suppressing activity or a cancer growth suppressing activity is exhibited.

EFFECT: metastin derivatives of the present invention also have a gonadotropic hormone secretion suppressing activity and sex hormone secretion suppressing activity.

5 cl, 4 tbl, 18 ex

FIELD: chemistry.

SUBSTANCE: invention relates to substituted extratriene derivatives of general formula (values of radicals are given in the claim), useful in therapy, especially for treating and/or preventing steroid hormone-dependent disorders which require inhibition of 17β-hydroxysteroid dehydrogenase (17-HSD) enzyme type 1, type 2 and/or type 3, as well as salts thereof, pharmaceutical preparations containing said compounds, as well as methods of producing said compounds.

EFFECT: improved method.

41 cl, 98 ex

FIELD: medicine.

SUBSTANCE: invention refers to medicine and concerns a method of treating insulin resistance. A peripherally effective amount of melanocortin receptor-4 agonist is introduced in a patient. The melanocortin receptor-4 agonist represents cyclic peptides containing the core sequence His-D-Phe-Arg-Trp-Cys.

EFFECT: invention provides effective treatment of insulin resistance by the peptide melanocortin receptor-4 agonist in the peripheral introduction.

34 cl, 11 dwg, 13 tbl

FIELD: medicine, pharmaceutics.

SUBSTANCE: group of invention relates to medicine, namely - to oncology and can be used for treatment of a neoplasm associated with excessive expression or amplification of .epidermal growth factor receptor 2 (HER2). According to the invention, the method includes administration of an effective quantity of the active components combination containing herceptin and CCI-779 (temsirolimus). Additionally, the group of inventions deals with versions of the herceptin and CCI-779 combination application for production of an anti-tumour medication.

EFFECT: use of the inventions allows higher effectiveness of treatment of cancer associated with excessive expression or amplification of HER2.

30 cl, 3 dwg, 4 ex

FIELD: medicine.

SUBSTANCE: newborn rabbits' pancreases are placed in a salt solution with an antibiotic at temperature 4-10°C and grounded microfragments. Further they are incubated in a free medium from serum at first incubation temperature 36.6 to 37°C, during a first incubation period with periodic replacement of a serum-free medium and removal of spontaneously collapsed undesired cells until at least 80% of the remained cells represent beta cells. It is followed by incubation in the serum-free medium periodically replaced at second incubation temperature 22 to 29°C, during a second incubation period until at least 78-90 % of the remained cells represent beta cells.

EFFECT: invention allows producing beta cells applicable for transplantation to patients with diabetes for stimulating natural insulin development.

4 cl, 40 tbl, 14 ex

FIELD: medicine.

SUBSTANCE: invention refers to medicine, namely oncology and urology and may be used in treating the patients with oncourological diseases the clinical course of which is complicated by a pyoinflammatory pathology. That is ensured by a session of leukocytapheresis to produce suspended leukocytes in plasma blood components to 3 mil. cells. Suspended leukocytes are added with the preparation of recombinant interleukin-2 (Roncoleukin ®) 300 thous. units. The prepared mixture is incubated in a thermostat at temperature 37°C for 90 minutes that is followed by the intravenous infusion to the patient. The daily therapeutic course makes 2 to 7 procedures depending on severity of the disease, the patient's state and the clinical laboratory evidence.

EFFECT: method enables considerably reducing a risk of pyoinflammatory complications in the given category of patients due to activating immune competent cells and normalising the body immune system.

1 ex

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