Condensed isoindoline as inhibitors of protein kinase c

 

(57) Abstract:

The invention relates to novel condensed isoindoline formula I

< / BR>
where ring b and F are independently from each other represent a 6-membered carbocyclic ring; R1means H, C1-C4alkyl; AND1and2pairs are selected from the group including =O, N, -OR", where R" denotes N;1and2pairwise mean =O; X in all positions independently from each other selected from the group: a) unsubstituted WITH1-C3alkylen, b) -S-; R3, R4, R5, R6independently from each other selected from the group comprising H1-C4alkyl; compounds 1 can be used for inhibiting protein kinase C (PKC) and inhibit the activity of tyrosine kinase (trk). Identified properties allow the use of compound 1 to inhibit proliferation of cancer cells. 2 C.p. f-crystals, 15 ill., 3 table.

The technical field

This invention relates to a condensed aryl or hetero-aryl-substituted the isoindole-2 - and-2,4-diones, which are hereinafter referred to as the "condensed isoindoline". The objects of this invention are also methods of obtaining and using these compounds.

Background image

Emitted from microorganisms substance, known as "K-a", in recent years attracted much attention for its multilateral functional activity, which it manifests. K-a is an alkaloid indolocarbazole, which was originally isolated from the culture Nocordiosis sp. (Kase, H. et al., 39 J. Antibiotics 1059, 1986). K-a is an inhibitor of several enzymes, including protein kinase C ("RCC") and tyrosine kinase (trk). The functional activity of the K-a are many and varied, for example, inhibition of tumor growth (U.S. patent 4877776 and 5063330; publication of the European patent 238011, issued Nomato), protivojinfectionnaya activity (U.S. patent 4735939), inhibition of inflammatory processes (U.S. patent 4816450), the treatment of diseases associated with nerve cells (WIPO publication WO 94/02488 issued by Cephalon, Inc. and Kyowa Hakko Kogyo Co., Ltd., published February 3, 1994).

These indolocarbazole have several common properties. In particular, they all have the bis-indole heterocyclic part. Staurosporin (allocated from Streptomyces sp.) and To-a (allocated from Nocordiosis sp.) contain part of the sugar molecule, which is attached two N-glycosidic bonds (to the nitrogen atoms of the indole). As the CTV. Indolocarbazole, as a rule, are lipophilic substances, so they are relatively easy to penetrate through biological membranes, and unlike proteins are characterized by a longer half-life in vivo.

The only drawback To-a with diverse and useful effect, is that the source of its receipt are microorganisms, so this stuff has to be distinguished from culture medium by fermentation. In the scientific literature describes the synthesis of K-a, but it is not suitable for commercial applications (Wood, J. et al., J. Am.Chem. Soc., 1995, 117, 10413). Therefore, compounds having a functional effect inherent To a, but produced by chemical synthesis, would have a big advantage compared to indolocarbazole compounds known at the present time.

A brief statement of the substance of the invention

This invention relates to compounds, which are referred to as the "condensed isoindoline". These compounds are biologically active. Condensed isoindoline are not containing indole molecules that can be synthesized de novo by chemical means.

Ctoma nitrogen in a 12 - or 13-positions (alphabetical designations of the rings, see the article Porter et al., 57, J. Org. Chem. 2105, 1992, used for reference purposes). In addition, condensed isoindoline not contain part of a sugar molecule attached two N-glycosidic bonds. Due to the fact that these compounds is missing a specified part of a sugar molecule, they can be easily synthesized. Useful and attractive is that these do not contain indole compounds that are not microorganisms, easy to synthesize, and that they have a biological activity, which make them suitable for a number of applications where previously only used by certain indolocarbazole.

Condensed isoindoline of the present invention have the following General formula (formula I):

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The preferred condensed isoindoline expressed by formula II

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Below details the structural elements of these compounds. In the compounds of formulas I and II element "X" in the rings C and E is nitrogen.

In this description of the invention are also considered preferred synthesis methods, including methods of obtaining lactam isomers.

Condensed isoindoline are designed for different judgments and/or indolocarbazole to enhance the function and/or viability of nerve cells, for inhibition of protein kinase C (CSWs) and inhibit the activity of tyrosine kinase (trk). Last specified action is directly related to the inhibition of proliferation of cancer cells, including cancer of the prostate. Due to the diverse action of the compounds according to the present invention can find application in a number of areas, including research and therapy.

Detailed description of the invention

I. Brief description of the drawings

In Fig. 1 depicts a graph showing the effect of derivative I-1 and I-2 condensed isoindoline activity h spinal cord.

In Fig. 2 depicts a graph showing the stimulatory effect of condensed isoindoline activity h at the base of the forebrain.

In Fig.3 shows a diagram of the synthesis of derivatives of bisingen.

In Fig.4 shows a diagram of the synthesis of condensed isoindoline.

In Fig.5 shows a diagram of the synthesis of condensed isoindolines, in which X is-C(=0)-.

In Fig.6 depicts the synthesis of condensed isoindoline (X = carbonyl) from 1-indanone.

In Fig.7 shows a diagram of the synthesis kondensirowannyh of isoindoline using Michael's reaction.

In Fig. 9 shows a diagram of the synthesis of selected condensed isoindoline using the Wittig reaction.

In Fig.10 shows a diagram of the synthesis of X-bis-condensed alkyl of isoindolines.

In Fig.11 shows a diagram of the synthesis of heterocyclic isoindoline with condensed ring Century.

In Fig.12 shows a diagram of the synthesis of bis-heterocyclic of isoindoline with condensed rings and F.

In Fig.13 shows a diagram of the synthesis of derivatives of bis-benzothiophene.

In Fig.14 shows a diagram of the synthesis of derivatives of interevention.

In Fig. 15 shows a diagram of the synthesis of condensed isoindoline using the Diels-alder reaction using acetylenedicarboxylate.

II. Condensed isoindoline

The object of the present invention are condensed isoindoline formula I

< / BR>
where ring b and F are independently from each other selected from the group including

(a) a 6-membered carbocyclic aromatic ring, in which up to 3 carbon atoms substituted by nitrogen atoms;

(b) 5-membered carbocyclic aromatic ring; and

(c) a 5-membered carbocyclic aromatic ring, in which
Ohm nitrogen and sulfur or nitrogen atom and oxygen;

R1selected from the group comprising H; alkyl with 1-4 carbon atoms; aryl; arylalkyl; heteroaryl; heteroaromatic; COR9where R9selected from the group comprising alkyl with 1-4 carbon atoms, aryl and heteroaryl; -OR10where R10selected from the group comprising h, alkyl with 1-4 carbon atoms; -CONH2, -NR7R8, -(CH2)nNR7R8and (CH2)nNR7R8where n is 1-4, and

(a) R7and R8independently from each other selected from the group comprising h, alkyl with 1-4 carbon atoms; or

(b) R7and R8together form a bridging group of the formula - (CH2)2-X1-(CH2)2- where X1selected from the group comprising-O-, -S -, and-CH2-;

AND1and2pairs are selected from the group comprising N, N; N,

-OR11where R11denotes H, alkyl with 1-4 carbon atoms, aryl with 6-10 carbon atoms or heteroaryl; N, -SR11; N, -N(R11)2; =0; =S and =NR11where AND1and2together may denote an atom with a double bond;

B1and2pairs are selected from the group including H, H; H, -OR11; N, -SR11; H, -N(R11); = 0; =S, and-NR11where IN1and22and1and2means =0;

X in all positions independently from each other selected from the group including:

(a) an unsubstituted alkylene with 1-3 carbon atoms;

(b) alkylen with 1-3 carbon atoms, substituted R2where R2choose from a group including:

(1) OR10; -SR10; R15where R15is an alkyl with 1-4 carbon atoms;

phenyl; naphthyl; arylalkyl with 7-15 carbon atoms; N; -SO2R9; -CO2R9; -COR9; alkyl, alkenyl and quinil with 1-8 carbon atoms, where

(i) each alkyl, alkenyl or quinil with 1-8 carbon atoms not substituted; or

(ii) each alkyl, alkenyl or quinil with 1-8 carbon atoms substituted by one of the substituents selected from the group comprising 1-3 aryl with 6-10 carbon atoms; heteroaryl; F; Cl; Br; I; -CN; -NO2; IT; OR9;- (CH2)nNR7R8where n is 1-4; -OCOR9, -OCONHR9; 0-tetrahydropyranyl; NH2; -N R7R8, -NR10COR9; -NR10CO2R9; -NR10CONR7R8; -NHC(=NH)NH2; -NR10SO2R9; -S(O)yR11where y is 1 or 2; -SR11; -CO2R9; -CONR7R8; -CHO; -COR9; -CH2OR7; -CH= NNR11R12where R< -CH=NOR11; -CH=NR9; -CH= NNHCH(N= NH)NH2; -SO2NR12R13where R13selected from the group comprising H, alkyl with 1-4 carbon atoms, aryl with 6-10 carbon atoms and heteroaryl, or R12and R13together form a bridging group; -PO(OR11)2, -OR14where R14is an amino acid residue, obtained after removal of the hydroxyl group of the carboxyl group; or

(2) a monosaccharide of 5-7 carbon atoms, in which each hydroxyl group independently may be substituted or is substituted by H, alkyl with 1-4 carbon atoms, alkylcarboxylic with 2-5 carbon atoms or CNS group with 1-4 carbon atoms; and

(C) a functional group selected from the group including

-CH= CH-; -NON-NON-; -O-; -S-; -S(= O)-; -S(S=O)2-; -C(R10)2-; -C= C(R2)2; -C(= O)-; -C(=NOR11); (OR11) (R11)-; -C(=O)CH(R15)-; -CH(R15)C(= O)-; -(=NOR11)CH(R15); -CH(R15)C(=NOR11)-; CONR15; NR15CO; -CH2Z-; -ZCH2- and-CH2ZCH2- where Z is-CR11; -O-; -S-; -C(=O)OR11; -C(=NOR11and-NR11;

R3, R4, R5and R6independently from each other selected from the group comprising H; aryl; geter>; NH2; -CH2HE; -CH2OR14; -NR7R8; -NR10OR9; -NR10CONR7R8; -SR11; -S(O)yR11where y is 1 or 2; -CO2R9; -COR9; -CONR7R8; -CHO; -CH= NOR11; -CH= NR9; -CH=NNR11R12; -(CH2)nSR9where n equals 1 to 4; -(CH2)nS(O)yR9; -CH2SR15where R15is an alkyl with 1-4 carbon atoms; -CH2S(O)yR14;

- (CH2)n, NR7R8; - (CH2)nOTHER14; alkyl, alkenyl, quinil with 1-8 carbon atoms, where

(a) each alkyl, alkenyl or quinil with 1-8 carbon atoms not substituted; or

(b) each alkyl, alkenyl or quinil with 1-8 carbon atoms substituted by one of the substituents selected from the group comprising 1-3 aryl with 6-10 carbon atoms; heteroaryl; F; Cl; Br; I; -CN; NO2; IT; OR9; -O(CH2)nNR7R8; -OCOR9; -OCONHR9; 0-tetrahydropyranyl; NH2; -NR7R8; -NR10COR9; -NR10CO2R9; -NR10ONR7R8; -NHC (=NH) NH2; -NR10SO2R9; -S(O)yR11where y is 1 or 2; -SR11; -CO2R9; -CONR7R8; -CHO; COR9; -CH2OR7; -CH=NNR11/SUP> or a monosaccharide of 5-7 carbon atoms, in which each hydroxyl group independently may be substituted or is substituted by H, alkyl with 1-4 carbon atoms, alkylcarboxylic with 2-5 carbon atoms or CNS group with 1-4 carbon atoms.

Preferred embodiments of this invention relate to the condensed isoindoline formula II

< / BR>
Preferably AND1and2pairs are selected from the group including H, H; H, HE; and =0; and In and In pairs chosen from the group comprising H, H; H, HE; and =0; provided that A1and2or1and2denote = 0.

R1preferably is N. When R1denotes COR9and R9denotes aryl, R9preferably is phenyl or naphthyl.

X in one of the positions or in both positions is preferably unsubstituted alkylene with 1-3 carbon atoms, -O - or-S-. When X is substituted by R2preferred group R2is OR10. When R2denotes arylalkyl with 7-14 carbon atoms, such group is preferably benzyl. When R2denotes alkyl, alkenyl or quinil, this group predpochtitel, alkenyl or quinil where the Deputy is aryl, the aryl is preferably phenyl or naphthyl. When Deputy group R2is-S(O)yR11where R11denotes aryl, it is preferably phenyl or naphthyl. When Deputy group R2is-CH= NNR11R12or-SO2NR12R13where R12or R13denotes aryl, it is preferably phenyl or naphthyl. When R12and R13taken together , represent a connecting group, such a connecting group is preferably - (CH2)2-X1- (CH2)2- where X1selected from the group comprising-O-, -S -, and-CH2-.

R3, R4, R5and R6preferred are N. When at least one of R3, R4, R5and R6denotes aryl, it is preferably an aryl with 6-10 carbon atoms, more preferably it is phenyl or naphthyl, provided that R3or R4denotes N, and R5or R6denotes N. When at least one of R3, R4, R5and R6denotes alkyl, alkenyl or quinil with 1-8 carbon atoms, it preferably t is N, and R5or R6denotes H.

Here the numerical ranges include all these numbers. For example, interval, defined as "1-4 carbon atoms" includes the values 1, 2, 3 and 4. Except where otherwise noted, the terms "aryl" or "heteroaryl" guess aryl or heteroaryl group can be substituted or unsubstituted.

Used in the definition of R14the term "amino acid" means a molecule that contains both amino group and carboxyl group. The definition of this term is "a-amino acid, which is a carboxylic acid to the carbon atom which is adjacent to the carboxyl group attached to the functional amino group. a-Amino acids may be natural or synthetic. Amino acids can also include "dipeptides", which is defined here as two amino acids connected by peptide bond. Thus, the structural elements of the dipeptides are not limited to a-amino acids and can be any molecule that contains both amino group and carboxyl group. The preferred a-amino acids, dipeptides, such as lysyl-alanine, and aminoalkanoic acids with 2-8 atoms and the pharmaceutically acceptable salts of the derivatives of condensed isoindoline. Used herein, the term "pharmaceutically acceptable salts" means salts of the accession of inorganic acids such as hydrochloride, sulfate and phosphate, or salts accession of organic acids, such as acetate, maleate, fumarate, tartrate and citrate. Examples of the pharmaceutically acceptable metal salts are alkali metal salts such as sodium salt and potassium, salts of alkaline earth metals, such as salt, magnesium salt and calcium salt, aluminum salt and zinc. Examples of the pharmaceutically acceptable ammonium salts are ammonium salt and a salt of Tetramethylammonium. Examples of pharmaceutically acceptable salts of the accession of organic amine salts are the research and piperidine. Examples of pharmaceutically acceptable salts of joining amino acids are salts of lysine, glycine and phenyl-alanine.

Consider here the connection can be included in the pharmaceutical compositions obtained by mixing these compounds with pharmaceutically acceptable nontoxic excipients and diluents. As mentioned above, such compositions can be used for parenteral administration, for example, in the form of liquid solutions or suspensions;

for oral introduction the aerosol;

or for dermal application, for example, in the form of adhesive tape.

These compositions can be used in the form of a dosage form and you can get well-known methods in pharmacology, for example, in accordance with the description given in the Handbook Remington's Pharmaceutical Sciences (Mack Pub. Co., Easton, PA, 1980). Preparations for parenteral administration may contain as diluents distilled water or saline, polyalkylene glycols such as polyethylene glycol, vegetable oils, hydrogenated naphthalenes and the like. Particularly useful fillers that regulate the release of active compounds that are biologically compatible and biologically destructible lactides, copolymers of lactide and glycolide or copolymers of polyoxyethylene and polyoxypropylene. Other potentially beneficial substances that contribute parenteral introduction of these active compounds are the copolymers of ethylene and vinyl acetate, osmotic pumps, implantable delivery systems, and liposomes. Preparations for inhalation contain as excipients lactose, or may be aqueous solutions containing, for example, polyoxyethylene-9-lauric ether, glycohol Locke nose. These preparations may also contain glycocholate for transbukkalno introduction, salicylate for rectal injection or citric acid for vaginal administration. Compositions for adhesive tapes are preferably lipophilic emulsion.

The substances according to the present invention can be used as the sole active component in the pharmaceutical composition or in combination with other active ingredients, such as other growth factors that enhance the viability of nerve cells or axonal regeneration in the treatment of cancer or HIV infection.

The content of compounds of the present invention in pharmaceutical compositions depends on a number of factors, including the dose of the drug, chemical properties (e.g., hydrophobicity) of the compounds used and the method of introduction. As a rule, the compounds of this invention can be used for parenteral administration in an aqueous physiological buffer solution containing about 0.1 to 10% connection ratio of weight to volume. Typical doses range from about 1 μg/kg to about 1 g/kg of body weight per day; a preferred dose is from about 0.01 mg/kg to 100 gnosti disease, General health needs of the subject, the relative biological efficacy of the selected compound, composition of fillers and method of drug administration.

III. The use of condensed isoindoline

Condensed isoindoline have important functional pharmacological activity, which may find application in a number of areas, including research and therapy. Condensed isoindoline, as a rule, have a positive impact on the functioning and/or viability of cells that respond to trophic factor, and inhibit the enzymatic activity, in particular, of enzymes such as trk and CSWs.

Influence on the function and/or viability of cells that respond to trophic factor, for example, nerve cells can be determined using any of the following tests: (1) analysis cholineacetyltransferase ("ChAT") in the culture of cells of the spinal cord; or (2) analyze the activity of ChAT in the culture of nerve cells in the base of the forebrain ("BFN"). Inhibition of the enzymatic activity can be determined using assays for inhibition of RCC and trk tyrosine kinase.

The term "impact", ispolzovanie their activities. A positive impact can be seen as a "strengthening" or "stimulation", and the negative impact is defined as "inhibiting" or "inhibition".

The terms "amplification" or "stimulation" used in conjunction with the terms "function" or "viability" means that the presence of condensed isoindoline has a positive effect on the function and/or viability of cells that respond to trophic factor, compared with cells not exposed to condensed isoindoline. In particular, for example, cholinergic neurons, not limiting the invention, it is possible to show that the condensed isoindole enhances the viability of cholinergic neurons with an increased risk of death (due to damage, disease, a degenerative condition or natural development) compared with cholinergic neurons, which are not exposed to such condensed isoindoline, if the period of functioning of the treated population of cells is increased compared with the untreated population of cells.

Used herein, the terms "inhibiting" and "inhibition" means that kondensirovannogo isoindoline.

Used herein, the term "neuron", "cell line neurons" and "nerve cell" means, without limiting this definition, heterogeneous population types of nerve cells, with one or more mediators and/or perform one or more functions; such cells preferably are cholinergic and sensory neurons. Used here, the phrase "cholinergic neurons" means the neurons of the Central and peripheral nervous system, neurotransmitter is acetylcholine; examples of such cells are the neurons of the base of the forebrain and the spinal cord. Used here, the phrase "sensory neurons" means the neurons that respond to stimuli (e.g., temperature, movement) of the skin, muscles and joints; examples of such cells are the neurons of the DRG.

Discussed here are "cells that respond to trophic factor" are cells with receptors that can specifically bind trophic factor; examples of such cells are neurons (e.g., cholinergic and sensory neurons) and cells are not neurons (e.g., monocytes and neoplastic cells).

The term "trk" refers to the family of high-affinity receptors it is to neurotrophin.

A. impact on the functioning and/or viability of cells that respond to trophic factor

Consider condensed isoindoline can be used to enhance the function and/or viability of the cell lines neurons. In this regard, condensed isoindoline can be applied separately or together with other condensed isoindoline, as well as in combination with other effective molecules, such as indolocarbazole, which also have the ability to affect the function and/or viability of these cells.

A number of neurological disorders associated with loss, damage, destruction of axons, increased risk of death of nerve cells and so on, These violations include, but are not limited to, Alzheimer's disease; diseases associated with motor neurons (for example, amyotrophic lateral sclerosis); Parkinson's disease; cerebrovascular disorders (e.g., stroke, ischaemia); Huntington's disease; dementia in AIDS; epilepsy; multiple sclerosis; peripheral neuropathy (for example, the defeat of DRG neurons with peripheral neuropathy due to chemotherapy), including diabetic neuropathy; narushennogo or spinal cord.

As shown in the examples in this section the description of the invention, the ability of condensed isoindoline to enhance the function and/or viability of the cell lines neurons, can be determined using (1) analysis of ChAT activity of the spinal cord or (2) analyze the activity of ChAT base of the forebrain.

ChAT catalyzes the synthesis of acetylcholine neurotransmitters and is considered an enzymatic marker for Funktsionalnyi Holi-magiceskih neurons. Functional neurons are also able to survive. The viability of neurons analyzed by quantitative determination of the specific absorption and enzymatic transformation of the dye (e.g., calcein AM) viable neurons.

Due to the diverse action of condensed isoindolines, they are effective in several analyses. These compounds can be used to create in vitro models of the viability of nerve cells, their functioning and identification or analysis of other synthetic compounds that have the same effect as condensed isoindoline. These compounds can be used in scientific research, which aims to study the. For example, if the condensed isoindoline that cause certain cell functions (for example, metagene), mark a radioactive isotope, it is possible to identify, isolate and purify the target cells, which are associated condensed isoindoline, with a view to their subsequent studies.

Degeneration, death or neoclerodane neurons are symptoms of many neurological disorders in humans, which include, without limitation, Alzheimer's disease; diseases associated with motor neurons (for example, amyotrophic lateral sclerosis);

Parkinson's disease; cerebrovascular disorders (e.g., stroke, ischaemia); Huntington's disease; dementia in AIDS; epilepsy; multiple sclerosis; disorders associated with shock or penetrating injuries of the brain or spinal cord; peripheral neuropathy; violations caused by the exciting amino acids. These compounds effectively increase the activity of ChAT, the scope of the present invention includes the use of these compounds for treating disorders associated with, for example, low activity or loss of DRG neurons.

Example III (A)(1). Analysis activity the national cholinergic neurons. Cholinergic neurons are directly related to the formation of the hippocampus, olfactory nuclei, nuclei miloslavov matter, cortical substance, amygdala and parts of the thalamus. Located in the spinal cord motor neurons are cholinergic neurons containing ChAT (Phelps et al., J. List. Neurol. 273:459-472 (1988)). The activity of ChAT used to study the effects of neurotrophins (e.g., NGF or NT-3) on the viability and/or function of cholinergic neurons. Analysis of ChAT is also used to determine changes in the levels of ChAT in cholinergic neurons.

Derivatives of condensed isoindoline increase ChAT activity when performing analysis in the culture of dissociated spinal cord of the rat embryo (Fig. 1). Connection 1-2 increases the activity of ChAT 150% compared to control cultures (which are not processed condensed isoindoline) after cultivation of the cells on the tablet for 2-3 hours for their attachment to the control wells with tissue culture. When performing these analyses condensed isoindole added directly to the culture of dissociated spinal cord. Compounds of the present invention, the zoom is ina least 120% compared with the activity of the control culture. The increase in ChAT activity was observed after a single injection of condensed isoindoline. The specified connection was introduced on the same day when received culture of dissociated cells of the spinal cord. The increase in the activity of ChAT was detected after 48 hours.

Methods.

Cells of the spinal cord of the embryo rats dissociatively and used to perform analyses in accordance with the described methods (Smith et al., J. Cell Biology 101:1608-1621 (1985); Glicksman et al., J. Neurochem. 61:210-221 (1993)). Dissociatively cells were obtained from the spinal cord of rats (14-15 days embryo development) using standard methods of dissociation by trypsin (Smith et al., J. Cell Biology 10:1608-1621 (1985)). The cells were placed in 6 x 105cells/cm2in wells with tissue culture plastic tablet coated with poly-1-ornithine, in the environment of N2without serum, supplemented with 0.05% of bovine serum albumin (Bottenstein et al., PNAS USA 76:514-517 (1979)). Cultures were incubated for 48 hours at a temperature of 37oWith in a humid atmosphere consisting of 5% CO2and 95% air. The ChAT activity was determined after 2 days in vitro no method Fonnula (Fonnum, J. Neurochem. 24: 407-409 (1975)), modified by Macmanaman etc. and Gliksman and others (McManaman et al. . Developmental Biology 125:311-320 (1988); Glicksman et al., e condensed isoindoline analyzed in relation to their ability to increase the activity of ChAT in the cultures of the base of the forebrain. It is established that the condensed isoindoline increase ChAT activity in the cultures of the base of the forebrain (Fig.2). Control cultures not treated condensed isoindoline.

When performing preliminary analyses of ChAT activity base forebrain compounds 1-3 and 1-4 did not increase the activity of ChAT.

Methods.

In embryos of rats was dissected to the base of the forebrain (17 or 18 day embryo development) and dissociatively cells neutral protease (DispaseTM, Collaborative Research). Neurons were placed with a density of 5 x 104cells/well (1.5 x 105cells/cm2) plates coated with poly-1-ornithine and laminin. Cells were cultured at 37 ° oWith in the environment of N2without serum, containing 0.05% bovine serum albumin, in a humid atmosphere consisting of 5% CO2and 95% air. The ChAT activity was determined after 5 days of cultivation with the help of ChAT analysis, described in example III(A) (1).

C. Inhibition of enzymatic activity

The ability of indolocarbazole To-a to inhibit the enzymatic activity of the RCC is well known and described in the scientific literature. The inhibition activity of the RCC is considered one of sporitelny diseases, allergic and cancerous conditions, as described in the following reference materials, namely U.S. patent 4877776 and 4923986; in the published description of the invention the European patent 558962 (issued September 8, 1993, the company E. R. Squibb &Sons, Inc.); Tadka, T. et al., 170(3) Biochem. Biophys. Res. Comm. 1151, 1980). Tyrosine kinase, which include trk, are enzymes that catalyze the transfer of phosphate from adenosine triphosphate (ATP) to the hydroxyl group of tyrosine many key proteins. It is established that activated tyrosine kinase proteins are products of approximately half of the known oncogenes (see, Chang, C-J & Geahlen, R. L. 55(11) J. Nat. Prods. 1529, 1992), therefore, to suppress, mediation, weakening and/or prevention of some cancers it is necessary to inhibit the activity of protein kinases (see above, Chang, C-J).

Due to the relationship between the activity of protein kinases and certain diseases and disorders (e.g. cancer), condensed isoindoline can be used effectively in scientific research and medical practice. In the research work of these compounds can be used, for example, to create analyses and models that enable us to understand the role of inhibition In medical practice connections inhibiting such enzymatic activity, can be used to remedy the harmful effects of these enzymes in terms of the occurrence of such disorders as cancer.

The following data indicate that inhibition of the enzymatic activity under consideration condensed isoindoline as a result of performing the following analyses: (1) analysis of the inhibition activity of the RCC; (2) analysis of the inhibition of the activity of tyrosine kinase trkA.

Example III(B) (1). Analysis of the inhibition activity of the RCC Condensed isoindoline inhibit the activity of protein kinase C (table VI). Described in the literature analysis of protein kinase C (Murakata et al., U.S. patent 4923986; Kikkawa et al., J. Biol. Chem. 257:13341-13348 (1982)) was performed at several concentrations of condensed isoindolines and determined concentration, allowing to inhibit 50% of protein kinase C (IC50table I and III.

Example III(C) (2). Analysis of the inhibition of the activity of tyrosine kinase trkA

Condensed isoindoline inhibit the tyrosine kinase activity when performing enzyme-linked immunosorbent assay (ELISA). trkA is the high affinity receptor for neurotrophins. Condensed isoindoline were placed on 96-groove C- (PLC) /protein pGEX) (see Rotin et al., 11 EMBO J. 559, 1992). These compounds are then analyzed for the ability to inhibit the phosphorylation of the substrate receptor trkA, PL. II and III

Methods

96-well plates to perform enzyme-linked immunosorbent assay (Nunc) were coated with 100 µl/well fosforiliruyusciye substrate (40 µg/ml PLC / fused protein pGEX) in 20 mmol of tributary, pH of 7.6, 137 mmol NaCl and 0.02% NaN3and left over night at a temperature of 4oC. Then the tablets three times washed with TBST (20 mmol Tris-buffer, pH of 7.6, 137 mmol NaCl, 0.2% tween-20) and blocked with 3% bovine serum albumin in TBST for 1 hour at a temperature of 37oC. the Tablets three times washed with TBST, and then twice washed with TBS (TBST without tween-20). Then to the reaction mixture (50 mmol N-2-hydroxyethylpiperazine-N'-2-econsultancy acid (HEPES), pH 7.4, 5 mmol MnCl2, 5 ml MgCl2, 140 mmol NaCl, 16 umol ATP and 15 ng trkA in a total volume of 100 μl) was added condensed paracervical in different quantities. For a negative control sample in the reaction solution was injected into a 100 mmol ethylenediaminetetraacetic acid (EDTA). Then the plates were incubated at temperature 37oC for 15 minutes. Added identifying antibodies, cha is Uchenie 1 hour at a temperature of 37oC. Then the tablets three times washed with TBST and incubated for 1 hour at a temperature of 37oWith goat protivolednym immunoglobulin, labeled alkaline phosphatase (1:2000 in TBST (Bio-Rad)). Culture thrice washed with TBST, and then twice washed with TBS, resulting in the use of reconstituted adenine dinucleotide phosphate (NADPH) as a substrate for alkaline phosphatase and implementation of reactions combination diaphorase and alcohol dehydrogenase (amplification system GIBCO-BRL method ELISA) obtained dyed product. Painted product identified at 490 nm in a spectrophotometer to read the tablets (Bio-tek).

IV. General description of methods for the synthesis of

Compounds of the present invention obtained in accordance with the following common ways of obtaining. Compounds in which X = CH2CH2shown on Fig.3. Known 2-2'-beenden (3, R1, R2, R3, R4, R5, R6= N; example IV(2)) obtained by using the improved method of combining 2-(tributylstannyl)of indene (2) 2-bromoindene (1), catalyzed by palladium. 2-(Tributylstannyl)inden (2) (example IV(1)) (Fig.3) obtained from 2-bromoindene (1) according to the method described in the scientific literature (J.-known in the field of organic synthesis.

The cycloaddition reaction of compounds of General formula 3 to the imide of maleic acid (method 1) is carried out preferably at a temperature of 160-200oWith, gives the corresponding tetrahydroisoquinoline 4 (R1, R2, R3, R4, R5, R6= H; example IV(3)). Cycloaddition reaction of 2-(2-indenyl)indeno were not previously documented. Cycloaddition reaction dienes to the imides of maleic acid are well known (see, for example, J. hm. Soc., Perkin Trans. 1, 1990, 2475). Connection example IV(3) digitalout by known methods using, for example, 2,3-dichloro-5,6-dicyan-1,4-benzoquinone, Pd on charcoal, sulfur or sodium nitrite (U.S. patent 4912197 and therein references), to obtain the corresponding aromatic derivative 5 isoindoline (example IV (4)) (compound 1-1, R1, R2, R3, R4, R5, R6= H). The lactams of General formula 6 (example IV(5), the connection 1-2, R1, R2, R3, R4, R5, R6= H) can be obtained by restoring imide 5 reductants (e.g., zinc amalgam, with gaseous hydrogen chloride, zinc amalgam in acetic acid, zinc in glacial acetic acid or hydride reducing agents, such as alumalite lithium). In Tehama (General formulas 6 and 7). Isomers position of the lactam can be divided by standard methods such as recrystallization

or chromatography, for example, chromatography on columns or liquid chromatography high resolution. The imides can be restored to hydroxylation (8, Fig.3), in which AND1AND2or1IN2= H, HE, using hydride reducing agents, such as borohydride or alumoweld (U.S. patent 4192107 and 4923986 and the links). The obtained hydroxyl group can be easily converted to CNS or thioalkyl group (U.S. patent 4923986). Derivatives in which AND1AND2or1IN2taken together , represent S or N, receive in accordance with the description given in the application for the European patent 0508792 Al.

In accordance with method II (Fig.15) cycloaddition of the corresponding diene to acetylenedicarboxylate (R = lower alkyl) gives the corresponding aromatic compounds of General formula 39. Isobenzofuran (General formula 40) can be obtained by dealkylation of ester nucleophiles (e.g., Lil, NaCN, NaSCH3, NaSCN, and so on) and then get anhydride in acetic anhydride. Imides of General formula 41 can be obtained by inter the General formula 42 can be obtained by restoring imide 41 reducing agents (for example, amalgam of zinc, with gaseous hydrogen chloride, zinc amalgam in acetic acid, zinc in glacial acetic acid or hydride reducing agents, such as alumalite lithium). In those cases where R2, R3, R4, R5or R6are not H or group X have different values, get isomers position of the lactam (General formula 42 and 43). Isomers position Lantana can be divided by standard methods such as recrystallization or chromatography, for example, chromatography on columns or liquid chromatography high resolution. The imides can be restored to hydroxylation (44, Fig.15) as described above.

In particular, compounds in which X = S, shown in Fig.13. 2,2'-Dibenzothiophene (25, R1, R2, R3, R4, R5, R6= N; example IV(6)) obtained by the method described for compound 3. The cycloaddition reaction of compounds of General formula 25 to diethylazodicarboxylate preferably at a temperature of 180-200oWith an appropriate carboalkoxylation (26, R1, R2, R3, R4, R5, R6= N; example IV(7)). Cycloaddition reaction of 2,2'-besensitive were not previously documented. Carbomethoxybiphenyl can example IV(8)) by dealkylation of ester nucleophiles (for example, Lil, NaCN, NS3, NaSCN, and so on) and then get anhydride in acetic anhydride (method II). Imides of General formula 28 ((compound 1-3), R1, R2, R3, R4, R5, R6= N; example IV(9)) can be obtained as a result of interaction of isobenzofuran formula 27 with 1,1,1,3,3,3,-hexamethyldisilazane and methanol as described above (method II). The lactams of General formula 29 ((compound 1-4), R1, R2, R3, R4, R5, R6= N; example IV(10)) can be obtained by restoring imide 28 reducing agents as described in method I. Imides can be restored to hydroxylation (30, Fig.13) in accordance with the description above.

The synthesis of the asymmetric interevention (X = S, CH2) is shown in Fig. 14. 2-(2'-Indenyl)benzothiophen (31, R1, R2, R3, R4, R5, R6= N; example IV(11)) obtained by the method described above for the compounds of formula 3. The cycloaddition reaction of compounds of General formula 31 to diethylazodicarboxylate gives the corresponding carboalkoxylation (32, R1, R2, R3, R4, R5, R6= N; example IV(12)). Carbomethoxybiphenyl 32 is turned into the appropriate isobenzofuran (33, R1, R21, R2, R3, R4, R5, R6= N; example IV(14)) can be obtained in accordance with the methods shown in Fig.13. The lactams of General formula 35 ((compound I-6), R1, R2, R3, R4, R5, R6= N; example IV(10)) can be obtained by restoring imide 34 reductants (e.g., zinc amalgam, with gaseous hydrogen chloride, zinc amalgam in acetic acid, zinc in glacial acetic acid or hydride reducing agents, such as alumalite lithium). In those cases where R2, R3, R4, R5or R6are not N receive isomers position of the lactam. Isomers position of the lactam can be divided by standard methods such as recrystallization or chromatography, for example, chromatography on columns or liquid chromatography high resolution. The imides can be restored to hydroxylation (37, Fig.14) as described above.

Derivatives of condensed isoindoline General formula II in which X= CH2CH2or CH=CH, is obtained using the methods described for compounds of formula 6 or 7 (Fig.3) except that instead of 2-bromoindene and/or alsamixergui 2-bromo-3,4-dihydronaphthalene can be obtained from 1 - or 2-tetralone in accordance with the methods of organic synthesis. Replacing the derivatives of indene derivative 2-benzocycloheptene (J. Am. Chem. Soc. 13:1344, (1991); J. Org. Chem. 44:1342 (1979)) gives a condensed isoindoline formula II in which X = CH2CH2CH2. Derivatives of ketone, in which X represents C=0, can be obtained by oxidation of imida (5) or lactam (6 or 7) standard oxidants (e.g., SeO2, SGAs3, PA2SGAs7or Mno2) (Fig.5). Alternatively, the derivative (11) where X = (From=0, N), can be obtained by using as starting material 2-bromantan-1-he (9) (J. Org. Chem., 1994, 59, 3453), which you can use to obtain 2-(tributylstannyl)-inden-1-he (10) according to the method described in example IV(1) (Fig.6). Similarly, the derivative (12) where X = (C=0, C=0), can be obtained by the interaction of 2-bromantan-1-it (9) 2-(tributylstannyl)-inden-1-one (Fig.7).

Connections with other groups of X such as X = S, O or C=0 (formula II) can be obtained by cycloaddition of the corresponding diene or imide of maleic acid as described in method I (Fig.4), or in accordance with the methods described for method II (Fig.15). For example, 2-(2 -(1-oxoindole))inden (X = (C=0), CH2), 2,2'-(1-oxo)beenden (X=(C=0), (= 0)), 2-(2-indenyl)benzothiophen (X= CH2S), 2-(2-indene is Catania corresponding 2-three-butylstannyl derived from the corresponding 2-bromoalkyl or heteroaryl derivative. Required connections can also be obtained by treating, for example, 2-(2-benzothiazyl)benzofuran (X = S, O) or 2-(2-benzothiazyl)benzothiophene (X = S, S) - imide of maleic acid in the presence of an acid catalyst, such as triperoxonane acid or Lewis acid (SnCl4Et2AlCl), which gives a compound of General formula 13 (Fig.8). These compounds can be collisional the corresponding derivatives 14 of condensed isoindoline by treating the catalyst, for example Pd(OAc)2in glacial acetic acid or Pd(OAc)2, tetrachloro-1,4-benzoquinone in C2H4Cl2(Fig.8).

The method of cross-linking catalyzed by palladium, which is well known in the field of organic synthesis, is used to produce other derivatives, such that X, shown in Fig.8, indicates 1-3 carbons (inclusive), which carry out the reaction mix vinyl-2-(triftormetilfosfinov) derived the corresponding cyclic ketone or 2-triflate derived the corresponding aryl or heteroaryl part with acceptable derivative of tin, described above.

Derivatives in which the nitrogen, having a group R1attached hydrogen atom Derivatives of the formula I, in which the substituents R3, R4, R5or R6are not H are in accordance with the above procedures, using as starting compound appropriately substituted intermediate compound, or by using the standard methods used in the field of organic synthesis for the interconversion of functional groups.

Derivatives with a substituent R2in which the group X is an olefin double bond (formula II) can be obtained by referirovanija derivatives in which X=(C=0, C=0 or C=0, N) (Fig.9), as a result of implementation of Wittig reactions known in the field of organic synthesis. The resulting alkenes (15) can be restored to the compounds of General formula (16) (Fig.9). Derivatives in which X = CH2you can easily alkilirovanii as a result of interaction with a strong base, such as BuLi, NaNH2or LiN(iPr)2and subsequent processing of the appropriate electrophile (J. Med. Chem 1992, 35, 3919; J. Org. Chem, 1991, 56, 4499) (Fig.10).

Condensed isoindoline formula I in which ring b and/or F are independently from each other to contain the atoms of nitrogen, oxygen, or sulfur, as indicated above for E1and E2you can get just as carbocyclic heterocyclic ring (the nitrogen is in any of 6 positions) shown in Fig.11. The cycloaddition reaction of compounds of General formula 17 to the amide of maleic acid gives the compounds of General formula 18. The dehydrogenation of the intermediate 18 is similar to the method of obtaining the compounds according to example IV(4) (Fig. 3) gives the derivative imide General formula 19 (Fig.11). Isomers of the lactam of General formula 20 can be obtained by the interaction of derivatives imide General formula 19 by reducing agents, such as amalgam of zinc and Hcl, zinc amalgam in acetic acid or hydride reducing agents, for example, alumoweld lithium. Isomers provisions can be divided by standard methods such as recrystallization or chromatography, for example, chromatography on columns or liquid chromatography high resolution. The imides restore to hydroxylation in which AND1AND2or1IN2= H, HE, hydride reducing agents such as borohydride or alumoweld.

Compounds in which ring b or F is a 5-membered ring containing oxygen or sulfur, can be obtained by using as starting substances, respectively periperal or tieneral instead indeno in accordance with the scheme of the synthesis shown in Fig.3 and 4. Periperal condensed with the ommun. 53:1770 (1988); Can. J. Chem. 56:1429 (1978); C. R. Hebd. Seances Acad. Sci., Ser. With 281: 793 (1975)). Tienerporno with condensed rings can also be obtained as described in the literature methods and their modifications (patent application Belgium BE 899925; Ind. J. Chem. 20B:271 (1981); Can J. Chem. 56: 1429 (1978); Bull. Soc. Chim. Fr. 11-12 pt2:2511 (1975); C. R. Hebd. Seances Acad. Sci., Ser. 277:1149 (1973)).

Alternatively, compounds in which ring b or F contains nitrogen atoms, can be obtained by using as starting compounds derived cycloalkylcarbonyl pyridine (X = C1-C3alkylen). Methods for the synthesis of derivatives of cycloalkanones described in the literature (J. Med. Chem. 36:3381 (1993), Chem Ber., 1970, 103, 2403), and these compounds can be used to obtain the intermediate compounds of General formula 21 or 22 (Fig.12). Derivative cycloalkyl or cycloalkanones with condensed ring can be converted into cyclic vinylboronic using known methods of organic synthesis. Intermediate compounds, such as vinylboronic are acceptable substrates for the method of stitching the tin shown in Fig.3 and 4.

Compounds in which the ring F contains an oxygen atom, can be obtained by using as starting compounds fullcontent the x ring F contains sulfur atoms, can be obtained by using as starting substances condensed cycloalkylation. Thienyl and foreline derivatives condensed with cycloalkyl ring can be obtained using the steps described in the literature methods (Acta Chem. Scand. 25:1287 (1971); J. Am. Chem. Soc. 103:2760 (1981)) or their modifications. These intermediate compounds can be converted into furyl - or tiercicannoca or furyl - or teenycinema. Source materials can alternatively be converted into the corresponding cyclic vinylboronic using known methods of organic synthesis. Intermediate compounds, such as vinylboronic, can be used to obtain the desired intermediate compounds according to the methods of stitching tin shown in Fig.3 and 4.

Rings b and F can be replaced by heteroatoms, as shown in Fig.12. Intermediate compound 23 can be obtained from intermediate compounds of tin 22 is replaced with a heteroatom in the ring, and cyclic vinylboronic 21, substituted heteroatom in the ring F, (or the corresponding triflate intermediate compounds of cyclic ketone). Derivative imide and lactam containing heteroatoms in the rings b and F, can be obtained by the methods shown nesoedinimye Michael and subsequent reaction ring closure, catalyzed by palladium, which is shown in Fig.8. Recovery of imides in accordance with the previously described methods gives the lactam isomers.

Legend

1H NMR spectrum1H NMR; (s) - singlet, (d) - doublet, (m) - multiplet, (bs) - broad singlet, (q) Quartet, (t)- triplet, (dd) doublet of doublets;

MS - mass spectrum; ES - electrostatic ionization; APCI - chemical ionization at atmospheric pressure; DMF is dimethylformamide; DMSO is dimethyl sulfoxide; THF - tetrahydrofuran; etc. - melting point.

Example IV(1). 2-(Tributylstannyl)inden

In a round bottom flask containing 2-brainden (1.64 g, 8.4 mmol) in 75 ml of E t3N, add palladium (II) acetate (304 mg, of 11.4 mmol), tetrakis (triphenylphosphine)palladium (0) (775 mg, 0.7 mmol) and hexabutylditin (6.4 ml, 12.7 mmol). The reaction mixture is heated to boiling point under reflux and follow the progress of the reaction by thin-layer chromatography (silica gel, tO: hexane, 1:5). After 1 hour, the starting material is completely consumed. The reaction mixture is allowed to cool to room temperature, diluted with CH2Cl2and filtered through celite. The solvent is removed under reduced pressure and the obtained with ozracing oil (containing trace amounts hexabutylditin). This connection is used in the next stage without further purification. 1H NMR (300 MHz, CDC13): _0.9 (m, 15H), 1.2 (m, 6N), 1.6 (m, 6N), 3.5 (s, 2H), 7.10 (s, 1H), 7.5-7.2 (m, 4H).

Example IV(2). 2,2'-Beenden

In a 100 ml round bottom flask, equipped with reflux condenser, load 1.2 g (6.3 mmol) of 2-bromoindene, 3.4 g (8.4 mmol) of 2-tributylstannyl (example IV(1)) and 70 ml of ethanol. To this mixture add 442 mg (0,63 mmol) chloride bis(triphenylphosphine)palladium(II). The reaction mixture is stirred at the boiling temperature under reflux for 16 hours, after which it is allowed to cool to room temperature, diluted with diethyl ether (50 ml) and filtered through a layer of aluminum oxide. The solvent is concentrated under reduced pressure and the obtained product is recrystallized from toluene, giving 870 mg (60%) 2,2'-bientina, etc.= 238o(In reference literature indicated, etc.= 238oC;

Chem. Ber., 1988, 121, 2195).

1H NMR (300 MHz, D13): 3.73 (s, 4H), 6.93 (s, 2H), 7.30 (m, SH). MS (ES+) m/z = 231 (M +1)

Example IV(3). 1a, 3A,4,7-Tetrahydroindene[2,3-C]indenyl[2,3-e]isoindole-1,3-dione

In a tightly we cork the test tube of borosilicate download 98 mg (0.4 mmol) of 2,2'-biondina (example IV(2)), 43 mg (0.44 mmol) of imide maleic acid the mixture to 130oC for 24 hours, after which it is allowed to cool to room temperature and concentrating the solvent under reduced pressure. The crude solid is purified by chromatography on columns (silica gel, mixture of 10-75% EtOAc - hexane) to give 50 mg (38%) of a white solid substance, etc., 244-247oC.

1H NMR (300 MHz, D13): 3.68 (s, 4H), 3.80 (m, 2H), 4.00 (bs, 2H), 7.24 (m, 7H), 7.53 (d, J = 7 Hz, 2H), MS (ES+) m/z = 328 (M +1)

Example IV(4). 1 N-Indenyl[2,3-C] -1H-indenol[2,3-e]isoindole-1,3-dione (compound I-1)

To a mixture of compound from example IV(3) (50 mg, 0.15 mmol) in toluene (4 ml) in one portion added solid 2,3-dichloro-5,6-dicyan-1,4-benzoquinone (79 mg, 0.35 mmol). The reaction mixture is heated in nitrogen atmosphere at a temperature of 65-70oC for 4 hours. The solution is cooled in an ice bath and the solid is collected by filtration. The crude residue was washed with cold methanol, giving a pale yellow solid (28 mg, 63%), etc., 244-247oC.

1H NMR (300 MHz, Dl3): 9.17 (d, 4 Hz, 2H), 7.55 (m, 7H), 4.15 (s, 4H); MS (ES): m/z 346 (M+1).

Example IV(5). 1 N-Indenyl[2,3-C] -1 H-indenyl[2,3-e]-3H-isoindole-1-he (compound I-2)

Get the amalgam of zinc, which 122 mg (1.9 mmol) of zinc dust suspended in 1 ml of water and add 35 mg (0.08 mmol) mu washed with water and once with ethanol.

The above is an amalgam of zinc suspended in 5 ml of ethanol and add compound I-1 (example IV(4)) (10 mg, 0.03 mmol). Add a few drops of conc. HCl, after which the reaction mixture is heated under reflux for 3 hours. When heated mixture within the first hour of the yellow color disappears. The reaction mixture is allowed to cool to room temperature, and the solution was concentrated under reduced pressure. The residue is dissolved in 10 ml of a mixture of THF - EtOAc (1:1), washed with saturated solutions Panso3and NaCl and dried (MgSO4). The desiccant is removed by filtration and the solvent concentrated under reduced pressure to obtain 8 mg (88%) of lactam as a white solid, T. p. 256oC.

1H NMR (300 MHz, D13): 9.20 (d, 8 Hz, 1H), 7.50 (m, 6N), 6.24 (s, 1H), 4.83 (s, 2H), 4.05 (s, 2H), 3.95 (s, 2H); MS (ES): m/z 310 (M+1).

Example IV(6). 2,2'-Dibenzothiophene

In a two-neck round-bottom flask equipped with a reflux condenser, a load of 3.3 g (15.6 mmol) of 2-bromobenzoate, 7,3 g (17 mmol) of 2-(tri-n-botillo)benzothiophene and 40 ml of toluene. To this mixture is added 360 mg (0.3 mmol) of tetrakis(triphenylphosphine)palladium(0) and 5 mg of 2,6-di-tert-butyl-p-cresol (BHT). The reaction mixture is heated under reflux in techenie dissolved in DMF and filtered through celite. The solvent is removed in vacuum and the solid is ground to powder with hexane that give 3.58 g (a 13.4 mmol, yield 85%) of 2,2'-dibenzothiophene in silver-black solids, etc.= 260-262oC.

1H NMR (300 MHz, DMSO-d6) 7.98 (m, 2H), 7.62 (s, 1H), 7.36 (m, 2H), 7.25 (s, 1H).

Example IV(7). 3,4-Carbomethoxybiphenyl[1,2-a]dibenzothiophene

In a tightly we cork glass tube load of 1.02 g (3.8 mmol) of 2,2'-dibenzothiophene, 3,1 ml (19 mmol) of diethylazodicarboxylate and 5 mg of 2,6-di-tert-butyl-p-cresol (BHT). The reaction vessel sealed in an atmosphere of N2and heated at a temperature of 190oC. the Reaction mixture is left to interact for 24 hours. Then the reaction vessel is cooled and the contents transferred into a round bottom flask, adding l3and remove the solvent in vacuo. Solid absorb diethyl ether and filtered to obtain 468 mg (1.07 mmol, 28%) of 3,4-carbomethoxybiphenyl[1,2-a]dibenzothiophene in the form of a pale yellow solid, etc., 206-207oC. a Second crystallization gives another 280 mg of the substance, resulting in a total yield of up to 45%.

1H NMR (300 MHz, DMSO6) 8.27 (d, 7.4 Hz, 2H), 8.05 (d, 7.9 Hz, 2H), 7.65 (m, 4H), 4.57 (q, 7.1 Hz, 4H), 1.38 (t, 7.1 Hz, 6H).

Note the ol) 3,4-carbomethoxybiphenyl[1,2-a] dibenzothiophene and 50 ml of DMF. To this mixture is added 124 mg (2.5 mmol) of sodium cyanide and 476 mg (2.5 mmol) of solid three-hydrate lithium iodide. The reaction mixture is heated to a temperature of 150oWith and analyzed by thin-layer chromatography. Then enter the additional number of NaCN and LiI. The reaction lasts for 36 hours, and during this time add a total of 4 equivalent NaCN and LiI, after which the starting material is completely consumed. The reaction mixture is cooled to room temperature and poured into cold (0o(C) an aqueous solution of Hcl. The mixture is filtered and washed with water. The obtained solid is dried in vacuum.

The above crude solid is loaded into a round bottom flask and add 50 ml of acetic anhydride. Then the reaction mixture is heated to boiling point under reflux. The mixture is heated under reflux for 4 hours, after which the results of thin-layer chromatography shows that the reaction is complete (a new spot at Rf of 0.65 in a mixture of EtOAc; hexane, 1:1). The solvent is removed and the crude oil purified flash chromatography to obtain a bright yellow-orange solid. This solid is ground to powder with diethyl ether, giving 16 the society, etc. >300oC.

1H NMR (300 MHz, DMSO-d6) 9.54 (dd, 5.2 Hz, 2.5 Hz, 2H), 8.34 (dd, 4.0 Hz, 3.3 Hz, 2H), 7.73 (m, 4H).

Example IV(9). Benzothieno[2,3-C] benzothieno[2,3-e] isoindole-1,3-dione (compound I-3)

Benzothieno[4,5-a] benzothieno[6,7-a] isobenzofuran-1,3-dione (75 mg, 0.2 mmol) dissolved in 3 ml of DMF. To this mixture is added to 4.4 ml (to 20.8 mmol) of 1,1,1,3,3,3-hexamethyldisilazane and 30 μl (1 mmol) of methanol. After about 15 minutes, the suspension becomes transparent. After 1 hour, perform thin-layer chromatography, which shows that starting material consumed almost completely. The reaction mixture was stirred over night, the total reaction time is 18 hours. The solvent is removed to obtain 65 mg (0.18 mmol, yield 80%) benzothieno[2,3-C] benzothieno[2,3-e]isoindole-1,3-dione as a yellow solid, etc. >300oC.

1H NMR (300 MHz, DMSO-d6) 9.8 (dd, 5.0 Hz, 4.1 Hz, 2H), 8.25 (dd, 4.9 Hz. 4.1 Hz, 2H), 7.70 (m, 4H):

Example IV (10). Benzothieno[2,3-C] benzothieno[2,3-e]isoindole-1-he (compound I-4)

To 10 ml of a suspension of zinc amalgam (3 equivalent) in ethanol added 68 mg (0.18 mmol) benzothieno[2,3-C]benzothieno[2,3-e]isoindole-1,3-dione in solution in 10 ml of acetic acid. To the reaction mixture add 5 ml of concentrated Hcl and manicom during the night, then it becomes transparent and becomes slightly yellowish-brown color. The reaction mixture is cooled and poured a layer of mercury. Remove most of the solvent, after which the mixture is diluted with ethyl acetate and washed twice with a saturated solution Panso3. The organic layer is dried over SO4filter and remove the solvent. The crude reaction mixture is purified by chromatography on columns that give 65 mg (0.18 mmol, yield 100%) benzothieno[2,3-C]benzothieno[2,3-e]isoindole-1-it is in the form of a yellowish-brown solid, etc., 225-226oC.

1H NMR (300 MHz, DMSO-d6) 10.17 (dd, 6.4 Hz, 2.7 Hz, 1H), 8.22 (s, 1H), 8.21 (dd, 4.1 Hz, 3.7 Hz, 2H), 8.11 (dd, 6.5 Hz, 2.2 Hz, 1H), 7.63 (c, 3.7 Hz, 2H), 7.55 (t, 3.7 Hz, 2H), 5.19 (s, 2H).

Example IV(11): 2-(2'-Indenyl)benzothiophen

In a two-neck round-bottom flask equipped with a reflux condenser, a load of 2 g (13.3 mmol) of 2-bromoindene, 5,1 g (11.9 mmol) of 2-(tri-n-botillo)benzothiophene and 50 ml of toluene. To this mixture was added 1 g (1.5 mmol) dichloride bis(triphenylphosphine)palladium(II) and 5 mg of 2,6-di-tert-butyl-p-cresol (BHT). The reaction mixture is heated under reflux for 16 hours and cooled. The solvent is removed, the reaction mixture absorb a mixture of DMF - THF and filtered through celite. The solvent UD the Jena as an orange solid, etc.= 260-265oC.

1H NMR (DMSO d6) 7.82 (m, 4H), 7.61 (s, 1H), 7.35 (m, 5H), 3.97 (s, 2H).

Example IV(12). 3,4-Carbomethoxybiphenyl[1,2 - a]dibenzothiophene

In a tightly we cork glass tube load 480 mg (3.95 mmol) of 2-(2'-indenyl)benzothiophene, 3.2 ml (19 mmol) of diethylazodicarboxylate and 10 mg of 2,6-di-tert-butyl-p-cresol (BHT). The reaction vessel sealed in an atmosphere of N2and heated at a temperature of 190oC. the Reaction mixture is left to interact for 24 hours. The reaction vessel is cooled, its contents are transferred into a round bottom flask, adding l3and remove the solvent. The crude substance is passed through a column of silica gel and collect the top faction. The obtained solid substance absorb diethyl ether and filtered to obtain 538 mg (1,29 mmol, 23%) of 3,4-carbomethoxybiphenyl[1,2-a] dibenzothiophene in the form of a pale orange solid, etc., 186oC.

1H NMR (300 MHz, DMSO-d6) 8.15 (d, 7.4 Hz, 1H), 7.95 (d, 7.4 Hz, 1H), 7.71 (m. 2H), 7.56 (m, 2H), 7.41 (m, 2H), 4.50 (q, 7.0 Hz, 4H), 4.22 (s, 2H), 1.33 (t, 7.0 Hz, 6H).

Example IV (13). Indenyl[4, 5-a] benzothieno[6, 7-a]isobenzofuran-1,3-dione

In a round bottom flask is charged with 250 mg (0.6 mmol) of 3,4-carbomethoxybiphenyl[1,2-a] dibenzothiophene and 10 ml of pyridine. analyzed by thin-layer chromatography. Then add an additional amount of LiI. The reaction lasts for 36 hours, and during this time add a total of 5 equivalents LiI, after which the starting material is completely consumed. The reaction mixture is cooled to room temperature and poured into cold (0o(C) an aqueous solution of Hcl. The mixture is filtered and washed with water. The obtained solid is dried in vacuum.

The above crude solid is loaded into a round bottom flask and add 50 ml of acetic anhydride. The reaction mixture is heated to boiling point under reflux. The mixture is heated under reflux for 4 hours, after which perform thin-layer chromatography, which indicates the completion of the reaction (a new spot at Rf of 0.6 in a mixture of EtOAc hexane, 1:1). The solvent is removed and the crude oil purified flash chromatography to obtain a bright yellow-orange solid. This solid is ground to powder with diethyl ether, giving 160 mg (0.44 mmol, yield 40%) indenyl[4,5-a]benzothieno [6,7-a]isobenzofuran-1,3-dione as a bright yellow solid, etc. >300oC.

1H NMR (300 MHz, DMSO-d6) _9.65 (d, J = 7.2 Hz, 1H), 8.76 (d, J = 7.6 Hz, 1H), 8.15 (s, J = 7.4 Hz, 1H), 7.78-7.38 (m, 5H), 4.] benzothieno[6,7-a] isobenzofuran-1,3-dione (120 mg, 0.35 mmol) dissolved in 3 ml of DMF. To this mixture is added to 7.4 ml (35 mmol) of 1,1,1,3,3,3-hexamethyldisilazane and 50 μl (1 mmol) of methanol. After about 15 minutes, the suspension becomes transparent. Thin-layer chromatography, which is done after 1 hour, show that the original substance is almost completely consumed. The reaction mixture was stirred over night, the total reaction time is 18 hours. The solvent is removed to obtain 35 mg (0.18 mmol, 30% yield) indenyl[2,3-C] benzothieno[2,3-e]isoindole-1,3-dione as an orange solid.

1H NMR (300 MHz, DMSO-d6) _12.72 (s, 1H), 9.84 (d, J = 8.3 Hz, 1H), 8.14 (d, J = 6.4 Hz, 1H), 8.07 (d, J = 7.5 Hz, 1H), 7.82-7.28 (m, 5H), 5.03 (s, 2H), MS (APcI) 342 (M + H)

Example IV (15). Indenyl[2,3-C] benzothieno[2,3-e]isoindole-1 - and-3-one (compound I-6)

To 2 ml of a suspension of Zn amalgam (3 equivalent) in ethanol is added 10 mg (0.3 mmol) indenyl[2,3-C]tianfan[2,3-e]isoindole-1,3-dione in solution in 10 ml of acetic acid. To the reaction mixture was added 1 ml of concentrated Hcl and heat it to the boiling temperature under reflux. The reaction mixture is heated under reflux for 3 hours, causing it to become transparent and becomes slightly yellowish-brown color. R is the Etat and washed twice its saturated solution Panso3. The organic layer is dried over MgSO4filter and remove the solvent gives compound in the form of a mixture indenyl[2,3-C]tianfan[2,3-e] isoindole-1 - and-3-one, which is a yellowish-brown solid.

Mass spectrum (chemical ionization at atmospheric pressure) 329 (M + H).

Despite the detailed description of the present invention, it is not limited to this description, and its scope is defined by the appended claims and all equivalent modifications.

1. Condensed isoindoline formula

< / BR>
where ring b and F are independently from each other represent a 6-cichlinae carbocyclic aromatic ring;

R1selected from the group comprising h, alkyl with 1-4 carbons;

AND1and2pairs are selected from the group including =O, N, OR11where R11is N;

IN1and2pairs represent =O;

X in all positions independently from each other selected from the group comprising (a) an unsubstituted alkylene with 1-3 carbon atoms and (b) -S-;

R3, R4, R5and R6independently from each other selected from the group comprising h, alkyl with 1-4 carbon.

2. Connection on p. 1, where X in lyubovnaya by p. 1, where X in any position or at both positions is-S-.

 

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