WO2010070365A1

WO2010070365A1 - 1,3-heterocycles condensed with monoterpene skeleton, their use and pharmaceutical compositions comprising such compounds

Info

Publication number: WO2010070365A1
Application number: PCT/HU2009/000102
Authority: WO
Inventors: Ferenc FÜLÖP; Zsolt Szakonyi; V. Peter Pallai
Original assignee: Bioblocks Magyarország Gyógyszerkémiai És Fejlesztö Kft.
Priority date: 2008-12-18
Filing date: 2009-12-10
Publication date: 2010-06-24
Also published as: HU0800768D0; HUP0800768A2

Abstract

The invention relates to chiral compounds with monoterpene skeleton of general formula (I) - where in the formula X stands for O or H2; W stands for O, S, N-R2 or Ph-R3; Y stands for O or N-R4; R1 stands for H, C1-4Alk or (C2)1-4-Ph; R2 stands for C1-4Alk or Ph-R3; R3 stands for H, C1-4Alk, C1-4Alk-O or Hlg; R4 stands for H or Ph; and one of the signs --- means the presence of a double bond and the other means the absense of a double bond, with the proviso that only one of W and Y may simultaneously stand for oxygen - as well as to their prodrugs and salts formed with pharmaceutically acceptable acids. Furthermore, the invention relates to cytostatic pharmaceutical compositions comprising one or more compounds of general formula (I) and usual inert pharmaceutical carriers and/or auxiliary agents, to the use of the compounds of general formula (I) for preparing cytostatic pharmaceutical compositions as well as to the treatment and/or curing of cancerous illnesses.

Description

1,3-HETEROCYCLES CONDENSED WITH MONOTERPENE SKELETON, THEIR USE AND PHARMACEUTICAL COMPOSITIONS COMPRISING SUCH COMPOUNDS

FIELD OF THE INVENTION The invention relates to 1,3-heterocycles condensed with monoterpene skeleton, their use and pharmaceutical compositions comprising such compounds.

BACKGROUND OF THE INVENTION

It is known that the number of cancerous illnesses is increasing from year to year all over the world, and no drug is known which could be successfully used against all cancer types. Besides, the reaction of patients' organism to various drugs is very different. Therefore, the widening of drug choice is very important for overcoming cancerous diseases.

In the fight against malignant tumorous diseases several methods are already known (surgical intervention, irradiation, hormone treatment, use of cytostatics) which, supplemented with results in diagnostics, have together resulted in considerable progress in the recent years. In spite of the results, the methods used today show several disadvantages, the primary reason of which is that the molecular mechanism of cell division is not yet known. Therefore, one cannot successfully interfere in the development of the disease. Consequently, the way to recovery or the delay in the course of the disease goes often together with the ablation of individual organ pieces and in case of use of cytostatics even with troubles in blood-formation.

The most frequently used treatments in cancer therapy are surgical and irradiation treatment as well as chemotherapy. In irradion therapy, beside the widely used ionizing irradiation, in special cases, e.g. with skin cancers, phototherapy and, combined with irradiation and chemotherapy, local hyperthermy are used. In the accessories of chemotherapy, when assorting according to their effect, origin or structure, alkylating agents, plant-alkaloids, antibiotics, antimetabolites, other agents (e.g. asparaginase), as well as the similarly often used various hormones are equally found. Recent strategies used in chemotherapy are: combined chemotherapy; venous or arterial infusion in small doses and for long time of chemotherapeutical agents for decreasing toxicity; chemotherapy with high doses for overcoming drug resistency; the same combined with autologous marrow-transplantation; combination of chemo- therapeutical drugs and agents modifying biological response; enhanced use of adjuvant and neoadjuvant chemotherapy. The most frequently used agents modifying biological response are interferons, tumor necrosis factor, limphokines, e.g. interleukin-2, and monoclonal antibodies. Various dietetic methods and the use of still not cleared up serum preparations belong to methods that are used even today but do not show proved results in treating tumours. WO 2007130404 describes condensed pyrrol derivatives that can be used for treating lung cancer.

European patent application No. 1,557,415 describes condensed pyrimidine derivatives showing anticancerous effect. European patent application No. 1,631,225 discloses condensed heterocyclic compounds useable for treating skin diseases and tumours.

US 2005/0080096 Al describes condensed pyrimidinone, quinazolinone and phthalazinone derivatives that can be used for treating or preventing tissue damage originating from cell damage or cell decay due to necrosis or apoptosis.

US 7,071,189 describes pyrimidine derivatives having antitumorous effect. US 7,071,218 describes pyrrole derivatives that can be used for treating metabolism disturbances and cancer.

US 7,129,351 describes condensed pyrimido compounds that can be used for treating breast, colon, liver and pancreas tumours.

The most characteristic disadvantages of the methods as used at present are toxicity, harmful side-effects of considerable degree, tumor-specificity of small degree, development of resistency and limited effect spectrum. Most cytotoxic drugs used in cancer therapy cannot distinguish between neoplastic and normal proliferating cells. Therefore, in order to avoid irreversible damages of organs containing such cells (e.g. marrow, bowel) the drugs should be used in doses that are not able to kill all present neoplastic cells [Pharmac. Ther., 49, 43-54 (1991)]. Irradiation therapy may give rise to acute and chronic irradiation damages and, at the same time, it is, as a matter of course, inefficient in case of hypoxic cells and certain tumor types.

The chemotherapeutic agents also show many kinds of toxic side-effects. They may damage the central nervous system, the blood-forming organs, the mucous membrane of bowel and intestines and every cells that are able to multiply. Besides, they may cause damage to liver-, kidney-, lung- and heart- muscles. All cytostatics damage the immunoreactivity of the organism [Proc. Roy. Soc. Med., 63, 1063- 1066 (1970)]. Many of them show teratogenic or carcinogenic effect, may be they cause infertility or increase the frequency of secondary tumour treatments. 60-70% of the tumours automatically cannot be or can be only difficultly influenced by chemotherapy, or resistency or cross-resistency is developed in them during treatment.

Similar disadvantages show the processes that modify biological responses and use own mechanism of the organism. Namely, they are effective only in case of some tumour types and, on the other hand, they also cause toxic side-effects [Harrison's Principles of Internal Medicine, 12^th ed., International Edition, McGraw-Hill, Inc., New York, Vol. 2, 1587-1599 (1991)]. The interferons also show many and grave side-effects, among others they are e.g. cardiotoxic [Chest, 99, 557-561 (1991)]. The monoclonal antibodies have not fulfilled expectations, either [Eur. J. Cancer, 27, 936-939 (1991)].

Recently Zhou et al. have rendered account of use of compounds of 2-arylimino-substituted-l,3- thiazolidinone type exerting selective cytotoxicity on cancerous cells of drug-resistant lung carcinoma [Zhou H. Y., Wu S. H., Zhai S. M., Liu A. F., Sun Y., Li R. S., Zhang Y., Ekins S., Swaan P. W., Fang B. L., Zhang B., Yan B., J. Med. Chem., 51(5), 1242-1251 (2008)].

The inventors of the present application also have recently rendered account of the synthesis of chiral compounds of 2-arylimino-spirooxazolidine structure with monoterpenic skeleton, showing structural relationship with the above compounds [Szakonyi Z., Hetenyi A., Fϋlδp F., ARKIVOC part (iii), 33-42 (2008)]. During their work, starting from α-pinene, a natural monoterpene, they obtained chiral 2-arylimino-l,3-oxazolidines by ring-closure of thiocarbamide adducts obtained from aminodiols. Considerable disadvantage of the compounds and synthesis described in this publication is that exclusively five-membered heterocycles of spirooxazolidine structure could be prepared from aminodiols; the authors state that six-membered heterocyclic rings condensed with monoterpenic skeleton such as 1,3- oxazines could be revealed not even in traces. On the other hand, according to our investigations an important condition of the anticancerous effect is the hydrophilic 1,3-oxazine ring system condensed with hydrophobic monoterpene skeleton. It follows that the chiral compounds prepared according to the above publication do now show anticancerous effect.

1,3-Oxazines that are analogous to 1,3-oxazolidines form a compound family that is interesting both from chemical and pharmacological points of view [Lazar L., Fϋlόp F.: 1,3-Oxazines and their Benzo Derivatives, Comprehensive Heterocyclic Chemistry III (Eds.: Katritzky A. R., Ramsden C. A., Scriven E. F. V., Taylor R. J. K.), Elsevier, Vol. 8 (Volume Ed.: Aitken A. R.), pp. 373-459 (2008)]. These compounds do not have a hydrophilic 1,3-oxazine ring system condensed with hydrophobic monoterpenic skeleton, either, and thus they do not exert anticancerous effect. Their further disadvantage is that their chirality does not result from the natural source used during their preparation; therefore, an expensive chiral catalyst of complicated structure is needed for their preparation.

SUMMARY OF THE INVENTION

The object of the present invention is to develop novel compounds, the chirality of which derives from the natural source [(-)-mirtenale and (+)-α-pinene] as used during their preparation; besides, the compounds are readily available and need inexpensive starting materials, they show beside favourable therapeutic effect toxic effect not at all or only in a minimal degree and eliminate disadvantages of known tumour therapeutical compositions and processes, namely toxicity, specificity of small degree and small effect spectrum, and render possible prevention of tumorous deseases and, resp., stopping of division of abnormally growing cells and thereby curing of cancerous diseases.

The invention is based on the recognition that the novel terpene derivatives of general formula (I) render possible to attain the above object since they are non-toxic compounds of anticancerous effect that are able to kill abnormal tumour cells even in very low concentration ranges.

Based on the above the invention relates to chiral compounds of monoterpenic skeleton of general formula (I) - where in the general formula (I) X stands for O or H₂; W stands for O, S, N-R² or Ph-R³; Y stands for O or N-R⁴; R¹ stands for H, C_1-4AIk or (CH₂)_1-4-Ph; R² stands for C_MAlk or Ph-R³;

R³ stands for H, C₁.₄Alk, C_1-4 AIk-O or HIg; R⁴ stands for H or Ph; and one of the signs — means the presence of a double bond and the other means the absence of a double bond, with the proviso that only one of W and Y may simultaneously stand for oxygen — , as well as to their prodrugs and salts formed with pharmaceutically acceptable acids.

In the formulae AIk stands for alkyl, HIg stands for halogen, Ph stands for phenyl, Me stands for methyl, and Et stands for ethyl.

The compounds according to the invention can be used without toxic effect for preventing tumour development, for avoiding tumour-development in case of transplantation, to prevent metastases as well as to treat and cure tumorous patients in direct, adjuvant or combined way.

DETAILED DESCRIPTION OF THE INVENTION

From the compounds according to the invention the following have been found to have particularly advantageous effect: - (lR,2R,7S,9R)-( 10, 1 O-dimethyl-S-oxa^-aza-tricyclo^.1.1.O²⁷]undec-4-yilidene)-(3-chloro- phenyl)-amine,

- ( lR,2R,7S,9R)-{ 10, 1 O-dimethyl-S-oxa^-aza-tricyclolT.1.1.O²⁷]undec-4-yilidene)-(3-methoxy- phenyl)-amine,

- (lΛ,2Λ,7S,9Λ)-(10,10-dmeώyl-5-oxa-3-aza-tricyclo[7.1.1.0^{2 7}]undec-4-ylidene)-(4-methyl- phenyl)-amine,

- ( \R,2R,7S,9R)-(l 0, 1 O-dimethyl-S-oxaO-aza-tricyclof?.1.1.0²⁷]undec-4-ylidene)-(4-fluoro- phenyl)-amine,

- (yΛ^Λ. /^PΛXS-methyl-lO.lO-dimethyl-S-oxa-S-aza-tricyclop.l.l.O^^-undec^-ylidene)^- chlorophenyl-amine hydrochloride, - (lR,2R,7R,9R)-( 10, 10-dimethyl-S-oxa-S-aza-tricyclo^.1.1.0^{2 7}]ιindec-4-ylidene)-phenylamine,

- (15,25,7/?,95)-(10,10-dimethyl-5-oxa-3-aza-tricyclo[7.1.1.0²-⁷]undec-4-ylidene)-phenylamine, and

- (lS,2S,7S,9S)-( 10, 10-dimethyl-5-oxa-3-aza-tricyclo[7.1.1.0^{2 7}]undec-4-yilidene)-phenylamine. The invention also relates to the pharmaceutically accceptable salts of the above compounds. The term .pharmaceutically acceptable salts" relates to salts that are not toxic for living organisms. Such salts can be prepared by reacting the compounds of general formula (I) with an inorganic or organic acid. These salts often show more advantageous dissolution properties than the compounds as used for their preparation and thus they can be often used more advantageously e.g. for preparing liquid or emulsified compositions. The term ,,acid addition salt" relates to a salt of the compound of general formula (I) that is prepared by reacting a compound of general formula (I) with a mineral or organic acid. The pharmaceutically acceptable acid-addition salts are described in detail for example in the article by S. M. Berge, L. D. Bighley and D. C. Monkhouse [J. Pharm. Sci. 66:1 (1977)]. The compounds according to the invention have a basic character; accordingly, they react with several inorganic or organic acid forming pharmaceutically acceptable acid-addition salts.

The pharmaceutically acceptable acid-addition salts according to the invention can be prepared by reacting a compound of general formula (I) with equimolar or excess amount of an acid. The reagents are generally reacted in a suitable solvent such as diethyl ether, tetrahydrofuran, methanol, ethanol, isopropanol, benzene and the like. The salts are usually precipitated from the solution within one-two hours but the time necessary for this precipitation can reach even 10 days. The salts can be separated by filtration or any other usual method.

From the inorganic acids generally used for salt-formation we mention here the hydrochloric acid, hydrobromic acid, hydrogen iodide, sulphuric acid, phosphoric acid and the like while from the organic acids useable for this purpose we mention the p-toluenesulphonic acid, methanesulphonic acid, oxalic acid, p-bromophenolsulphonic acid, tartaric acid, citric acid, benzoic acid and the like. As examples of such pharmaceutically useable salts we mention the following salts of the compounds of formula (I): sulphate, pyrosulphate, hydrogen-sulphate, sulphite, phosphate, monohydrogen phosphate, dihydrogen phosphate, metaphosphate, pyrophosphate, chloride, bromide, iodide, acetate, propionate, decanoate, caprylate, acrylate, formiate, isobutyrate, caproate, heptanoate, oxalate, malonate, succinate, suberate, sebacate, fumarate, maleate, benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, hydroxybenzoate, metoxybenzoate, phthalate, sulphonate, phenyl acetate, phenyl propionate, phenyl butyrate, citrate, lactate, /3-hydroxy butyrate, tartarate, methanesulphonate, propanesulphonate, 1,5- naphthalinedisulphonate, naphthaline- 1 -sulphonate, naphthaline-2-sulphonate, mandelate and the like. The term ,,prodrug" relates to derivatives of compounds of general formula (I) according to the invention which contain a chemically or metabolicly decomposing group and which, after biotransformation, become pharmaceutically effective.

The compounds of formula (I) contain one or more asymmetry centre and thus they exist as enantiomers or diastereoisomers. The invention comprises both the mixtures and the separate isomers. The compounds of general formula (I) also exist in tautomeric forms, and the invention comprises both the mixtures and the separate tautomers.

The compounds of general formula (I) and their salts may also exist in the form of solvates which also fall under the scope of the invention. The solvate preferably comprises a hydrate or an ethanolate. The invention also relates to radiolabelled derivatives of compounds of general formula (I) which can be used for biological studies.

The invention also relates to processes of preparation of compounds of general formula (I). The process according to the invention is illustrated by reaction scheme 1 where the meaning of symbols and, resp., abbreviations is as follows: AIk = alkyl; Me = methyl; Ph = phenyl. For preparing the compounds according to the invention one proceeds according to reaction scheme 1 in the following way. ai) The synthesis of 4-pyrimidinones is carried out by melting suitable azetidinones with aryl imidates without organic solvent, at higher temperatures. Detailed experimental description and the features of the prepared compounds are given in Examples 1 and 2. a₂) The compounds can be also prepared by reacting the suitable amino acid esters and arylimidates in ethanol at higher temperatures. b) The first step of synthesis of 2,4-pyrimidinediones and 2-thioxo-4-pyrimidinones is carried out by condensing amino acid esters and corresponding isocyanates and, resp., isothiocyanates in an organic solvent. Following evaporation the intermediate product is boiled with aqueous hydrochloric acid solution. The synthesis is carried out by starting from ethyl-(l/?,2/?,35,5Λ)-2-amino-6,6-dimethyl- bicyclo[3.1.1]heptane-3-carboxylate of formula 4. The detailed experimental description and the features of the prepared compounds are given in Examples 3 and 4. c) The first step of the synthesis of 2-alkyl- and, resp., 2-arylamino-l,3-oxazines can be carried out by condensing the suitable aminoalcohols and suitable alkyl- and, resp., arylisotbiocyanates in an organic solvent. After evaporation the intermediate product is stirred at room temperature in a methanolic methyl iodide solution and then in a methanolic potassium hydroxide solution. Detailed experimental description and features of the prepared compounds are given in Examples 5 to 17.

The invention also relates to the preparation of anticancerous pharmaceutical compositions comprising compounds of general formula (I) or their salts. According to the invention one proceeds by mixing one or more compounds or salts of general formula (I) with usual inert pharmaceutical carriers and/or auxiliary agents to anticancerous pharmaceutical compositions.

Furthremore, the invention relates to the use of one or more compounds of general formula (I) and/or the salts of such compounds for preparing anticancerous pharmaceutical compositions.

The invention also provides a method for treating and/or curing cancerous illnesses which comprises administering an effective amount of the pharmaceutical composition according to the invention to a patient in need of such treatment.

The term ,,treatment" is used here in the usual sense, that is it relates to prevent, hinder, improve, suppress or slow down the cancerous disease or to decrease the gravity of the illness.

In the practice the compounds of general formula (I) according to the invention are administered in solid, liquid or spray form alone or in combination with one or more usual pharmaceutical carrier or auxiliary agent. Accordingly the invention comprises also pharmaceutical compositions which comprise an effective amount of compounds of general formula (I) together with a pharmaceutically acceptable carrier. As a matter of course, the compounds according to the invention can be also administered together with other anticancerous agents, too. As solid carriers useable in the compositions according to the invention one or more agents can be used that simultaneously serve as sliding, solubilizing, suspending, binding, filling, pressing, disintegrating, flavouring or encapsulating agents.

In the case of powders the carrier can be a finely divided solid material that is mixed with finely divided particles of compounds of general formula (I). In the case of tablets, the compound of general formula (I) is mixed in suitable ratios with a carrier that has the required pressing properties, and the mixture is pressed to the desired form and size.

The above-mentioned powders and tablets contain the compound of general formula (I) in an amount ranging up to 99% by mass.

The compositions of spray form are generally used as aerosol compositions for absorption by the skin surface or through the lungs.

As examples of the solid carriers that can be used in the compositions according to the invention we mention the talc, calcium phosphate, magnesium stearate, lactose, dextrin, starch, gelatine, cellulose, methyl cellulose, sodium carboxymethylcellulose, polyvinylpyrrolidone and waxes of low melting point.

In the compositions according to the invention any pharmaceutically acceptable liquid carrier can be used which is suitable for preparing solutions, suspensions, emulsions, syrups and therapeutical drinks. The compounds of general formula (I) can be dissolved or suspended in a pharmaceutically acceptable liquid carrier such as water, organic solvents, or pharmaceutically acceptable oils or fats or a mixture thereof. The said liquid composition may contain other suitable pharmaceutical additives such as solubilizing agents, emulsifying agents, buffers, conserving agents, sweeteners, flavouring agents, suspending agents, thickeners, colouring agents, viscosity regulators, stabilizers, osmotic pressure regulators and the like.

For oral or parenteral administration the following examples of liquid carriers are mentioned: water solutions containing additives such as cellulose derivatives, preferably sodium carboxymethylcellulose, alcohols including primary and secondary alcohols such as glycols, alcohol derivatives or oils such as fractionated coconut oil and arachis oil. In the case of parenteral administration the carrier oil can be an ester such as ethyl oleate or isopropyl myristate.

The orally administrable compositions according to the invention can be liquid or solid. The compositions according to the invention that are sterile solutions or suspensions can be used as intramuscular, intraperitoneal or subcutaneous injections. The sterile solutions can be administered intravenously, too.

The compounds according to the invention can be formulated together with pharmaceutically active other substances. The choice of such materials depends on the aim of using the composition.

In the compositions according to the invention the concentration of compounds of general formula (I) depends on several factors such as method of administration, chemical nature of the compounds, state and clinical indication of the patient. Therefore, the concentration can vary in a wide range. Generally the concentration of the active agent can be 0.002-99% by mass, for example 0.01-70% by mass, preferably 0.05-40% by mass, more preferably 0.1-20% by mass.

A composition according to the invention can be administered enterally, parenterally, locally, rectally or systemicly, depending on the prescription and the active agent used, e.g. in the form of tablets, capsules, powders, granules, syrup, spray, or injection solution.

The compositions for enteral administration can be for example powders, simple or coated tablets, tablets with protracted effects, soft capsules, hard capsules, rectal suppositories, suspensions and solutions which, if desired, may contain the active agents together with one or more usual carriers.

The compositions for parenteral administration can be e.g. preparations for intradermal, subcutaneous, intraperitoneal or intravenous injections or infusions. Other parenteral compositions can be applied not only on the skin but also on the mucous membrane. Such local compositions can be e.g. gels, creams, ointments, shampoos, soaps, sprays, rinsing agents, smears, aerosols, and other pharmaceutical preparations suitable for local administration.

The compounds according to the invention preserve their effect when used in any form, e.g. when used in the usual oral, rectal or injection form.

In the case of oral administration the daily dose of the compounds according to the invention amounts for adults to 200-1000 mg, preferably 50-500 mg, more preferably 20-100 mg. These doses can be increased or decreased depending on the state of the patients.

The acute oral toxicity test of the compounds according to the invention in cornseed-oil on female mice proved that the compounds are not toxic in a daily dose of 2000 mg/kg. The (1S,5S)- and (lR,5R)-apopinene (b and e) serving as starting compounds for preparing the compounds according to the invention were synthesised according to literature methods, in the way as shown in the enclosed reaction scheme 2 [Shibuya K., Synth. Commiin., 24, 2923-2941 (1994); Lightner,

D.A. and Crist, B.V., Tetrahedron, 41, 3021-3028 (1985)]. The (-)-(lR,5S)-myrtenal (a) and the (+)- (lR,5R)-α-pinene (c) are commercial products (Sigma-Aldrich Kft).

When describing the syntheses of the compounds according to the invention we refer to reaction schemes 3-8.

In reaction scheme 3 the preparation of 4-pyrimidinones is shown, starting from chiral azetidinones of formula 1. In reaction scheme 4 the preparation of 2,4-pyrimidinediones and 2-thioxo-4-pyrimidinones is shown, starting from aminocarboxylic esters of formula 4.

In reaction schemes 5-8 the preparation of 1,3-oxazines condensed with monoterpenes is shown, starting from aminoalcohols of formulae 7a-c, 14, 16 and 18.

Schemes 1 to 8 are illustrated in Fig. 1 in the enclosed drawing. The reactions are followed by thin-layer chromatography. The working up can be carried out by evaporation or extraction. The product can be purified by crystallization or chromatography. The structure and purity of compounds is proved and, resp., controlled by NMR spectroscopy, melting-point measurement and/or CHN microanalyses.

Experimental General Procedures. ¹H NMR spectra were recorded by Bruker Avance DRX 400 spectrometer at 400.13 MHz (¹H) and 100.61 MHz (¹³C) [D=O (TMS)] in CDCl₃ or in D₂O in a 5-mm tube. Chemical shifts are expressed in ppm (D) relative to TMS as internal reference. J values are given in Hz. Microanalyses were performed on a Perkin-Elmer 2400 elemental analyser. Optical rotations were obtained with a Perkin-Elmer 341 polarimeter. Melting points were determined on a Kofler apparatus and are uncorrected. Chromatographic separations were carried out on Merck Kieselgel 60 (230-400 mesh ASTM). Reactions were monitored with Merck Kieselgel 60 F,₅₄-precoated tic plates (0.25 mm thickness).

The enantiomeric purities of the prepared compounds were determined by means of GC measurements involving direct separation of the enantiomers on a CHIRASIL-DEX CB column (2500x0.25 mm I.D.) at 160 ⁰C and 80 kPa for azetidinones. IR spectra were measured with a FT-IR spectrometer.

Example 1 (lΛ^ΛJ^^^^-ChlorophenyO-lOjlO-dimethyl-S^-diazatricyclo^.l.l.O^lundec-S-eπ-β- one (compound 3) (Scheme 3)

A mixture of 1.80 g (10.9 mmol) of (lΛ,2Λ,5S,7Λ)-8,8-dimethyl-3-azatricycIo[5.1.1.0²'⁵;|nonan- 4-one (1) and 2.60 g (14.2 mmol) of ethyl 4-chlorobenzimidate was heated in neat at 130 ⁰C for 24 h. The mixture was then rubbed with 30 ml of «-hexane and, after 1 h standing at room temperature, the resulting crude product was filtered off and purified by column chromatography (silica gel, chloroformethyl acetate = 15:1, R_f = 0.30).

Isolated compound: 1.75 g (53%); mp: 225-226 ⁰C; [a]™ = +85.8 (c = 1, MeOH); IR = 3211, 2904, 1660, 1277, 839 cm^"1. Anal. Calcd. for C_nH₁₉ClN₂O (302.80): C, 67.43; H, 6.32; N, 9.25. Found: C, 67.25; H, 6.64; N, 9.17. ¹H NMR (CDCl₃) δ (ppm): 0.98 (3H, s), 1.04 (IH, d, J = 10.6 Hz), 1.27 (3H, s), 1.97-2.03 (IH, m), 2.22-2.32 (3H, m), 2.64-2.71 (IH, m), 3.01 (IH, dt, J = 2.0; 10.6 Hz), 4.58 (IH, d, J = 10.6 Hz), 7.42 (2H, d, J = 8.6 Hz), 7.70 (2H, d, J = 8.6 Hz). ¹³C NMR (CDCl₃) δ (ppm): 21.1 (Me), 26.9 (CH₂), 27.0 (Me), 28.2 (CH₂), 33.0 (CH), 40.2 (C_q), 41.1 (CH), 48.0 (CH), 59.3 (CH), 128.3 (CH₂₁), 129.6 (CHJ, 132.8 (C,), 137.8 (C,), 148.1 (C_q), 175.0 (C,, C=O).

The starting material (lΛ,2Λ,55,7Λ)-8,8-dimethyl-3-azatricyclo[5.1.1.0^2>5]nonan-4-one was prepared as below:

A mixture of 12.21 g (100.0 mmol) of (-)-(15,55)-apopinene (compound b), prepared via literature method [Lightner, D.A. and Crist, B.V. Tetrahedron, 41, 3021-3028 (1985)], and 14.30 g (101.2 mmol) of chlorosulfonyl isocyanate (CSI) was stirred in 300 ml of dry ethyl ether for 48 h at room temperature. 20.4 g (162 mmol) of dry sodium sulfite in 140 ml of water were then cautiously added dropwise to the solution while the pH was held at 7-8 by the addition of 20% aqueous potassium hydroxide. After 2 h stirring at the appropriate pH, the organic phase was separated and the aqueous layer was extracted with diethyl ether (2x100 ml). The combined organic layer was dried (Na₂SO^ and evaporated, and the white crystalline product obtained was recrystallized from isopropyl ether.

Isolated compound: 13.54 g (82%); mp: 68-72 ⁰C; [α]_p = -80.0 (c = 0.5, MeOH; ee >99%); IR = 3247, 2914, 1710, 1380, 1256, 1189 cm^"1.

Example 2 (l/?,2Λ,75,9Λ)-4-(3-ChIorophenyl)-10,10-dimethyl-3,5-diazatricyclo[7.1.1.0²'⁷]undec-3-en-6- one (compound 2) (Scheme 3)

The synthesis of compound 2 was accomplished by analogy with Example 1, starting from 1.40 g

(8.5 mmol) of (l/^Λ.SSJφ-S.S-trimemyl-S-azatricyclo^.l.l.O^nonan^-one (1), prepared according to Example 1, and 2.02 g (11.0 mmol) of ethyl 3-chlorobenzimidate. Isolated compound: 1.12 g (43%); mp: 144-147 ⁰C; [α]*⁰ = +70.5 (c = 0.5, MeOH); ER = 3245,

2918, 1661, 1575, 1256, 737 cm^'1. Anal. Calcd. for C_nH₁₉ClN₂O (302.80): C, 67.43; H, 6.32; N, 9.25.

Found: C, 67.57; H, 6.19; N, 9.47. ¹H NMR (CDCl₃) δ (ppm): 0.98 (3H, s), 1.06 (IH, d, J = 10.6 Hz),

1.27 (3H, s), 1.96-2.01 (IH, m), 2.22-2.32 (3H, m), 2.67-2.73 (IH, m), 3.01 (IH, dt, J = 2.5; 10.6 Hz),

4.58 (IH, d, J = 11.1 Hz), 7.36 (IH, t, J= 7.8 Hz), 7.44 (IH, d, J= 8.6 Hz), 7.68 (IH, d, J= 7.6 Hz), 7.86 (IH, s), 9.15 (IH, br s). ¹³C NMR (CDCl₃) δ (ppm): 21.1 (Me), 26.9 (CH₂), 27.0 (Me), 28.1 (CH₂), 33.0

(CH), 40.2 (C_q), 41.1 (CH), 48.0 (CH), 59.2 (CH), 125.0 (CH₃,), 127.5 (CH₃,), 130.6 (CH₃₁), 131.6 (CHJ,

135.6 (C_q), 136.1 (C,), 148.2 (C,), 175.4 (C,, C=O). Example 3 (lΛ^Λ^S^^-lO^O-Dimethyl-S-phenyl-SjS-diazatri-cycloIT.l.l.O^'lundecan^^dione

(compound 5) (Scheme 4)

1.15 g (5.4 mmol) of ethyl (lΛ,2Λ,35,5Λ)-2-amino-6,6-dimethylbicyclo[3.1.1]heptane-3- carboxylate (4) and 0.68 g (5.7 mmol) of phenyl isocyanate were dissolved in 40 ml of toluene. The reaction mixture was stirred at room temperature for 2 h, then evaporated to dryness. The resulting product was washed with w-hexane and filtered off. The obtained crystalline crude product was refluxed with 60 ml of 5N hydrochloric acid for 3 h. After cooling the mixture was extracted with chloroform (3x30 ml). The combined organic layer was dried (Na₂SO₄), filtered and evaporated resulting in white crystalline product which was purified by column chromatography (silica gel, «-hexane:ethyl acetate = 2:1, R_f= 0.3).

Isolated compound: 1.05 g (68%); mp: 210-211 ⁰C; [α£° = +57.2 (c = 1, MeOH); IR= 3230, 2909, 1722, 1681, 1441, 1215, 695 cm^"1. Anal. Calcd. for C₁₇H₂₀N₂O₂ (284.35): C, 71.81; H, 7.09; N, 9.85. Found: C, 72.15; H, 7.38; N, 9.56. ¹H NMR (CDCl₃) δ (ppm): 0.93 (3H, s), 1.27 (3H, s), 1.32 (IH, d, J = 11.8 Hz), 1.97-2.05 (2H, m), 2.24-2.35 (2H, m), 2.77 (IH, dd, J = 14.6; 4.5 Hz), 3.17 (IH, dt, J = 9.6, 1.5 Hz), 4.14 (IH, d, J = 9.6 Hz), 5.60 (IH, br s), 7.15 (2H, d, J = 1.6 Hz), 7.36-7.47 (m, 3H). ¹³C NMR (CDCl₃) δ (ppm): 20.5 (Me), 25.6 (CH₂), 26.8 (Me), 28.4 (CH₂), 33.6, 40.4 (C_q), 40.8 (CH), 48.6 (CH), 49.9 (CH), 128.9 (CHJ, 129.2 (CH₃₁), 129.7 (CH₃₁), 136.2 (C_q), 153.8 (C=O), 174.1 (C=O).

The starting material ethyl(l/?,2Λ,35,5Λ)-2-amino-6,6-dimethylbicyclo[3.1.1]heptane-3- carboxylate (4) was prepared as below:

1.16 g (7.0 mmol) of (lΛ,2/?,55,7Λ)-8,8-dimethyl-3-azatricyclo[5.1.1.0^2>5]nonan-4-one (1), prepared according to Example 1, was stirred in solution of 10% hydrochloric acid in dry ethanol (20 ml) at room temperature. After 1.5 h stirring, the solution was evaporated to dryness and the crystalline product obtained was recrystallized from diisopropyl ether/ethyl acetate mixture.

Isolated compound: 1.54 g (89%); mp: 138-139 ⁰C; [af° = +23.0 (c = 0.5, MeOH); IR = 2918, 1729, 1373, 1189 cm^"1.

Example 4 (lΛ,2Λ,75,9Λ)-10,10-Dimethyl-5-phenyl-4-thioxo-3,5-diazatricyclo[7.1.1.0^{2 7}lundecan-6-one

(compound 6) (Scheme 4)

0.21 g (1.55 mmol) of phenyl isothiocyanate and 0.31 g (3.0 mmol) of ethyl (lR,2R,3S,5R)-2- amino-ό.ό-dimethylbicyclop.l.lJheptan-S-carboxylate (4) prepared according to Example 3 were dissolved in 20 ml of toluene. The reaction mixture was stirred at room temperature for 2 h, then evaporated to dryness. The resulting crude product was refluxed with 20 ml of 3N hydrochloric acid for 3 h. After cooling the mixture was extracted with chloroform (3x30 ml). The combined organic layer was dried (Na₂SO₄), filtered and evaporated resulting in white crystalline product which was purified by recrystallization from diisopropyl ether/ethyl acetate mixture. Isolated confound: 0.67 g (75%); mp: 269-272 ⁰C; [α£° = +19.0 (c = 0.3, MeOH); IR= 2972, 1755, 1681, 1363, 699 cm ¹. Anal. Calcd. for C_nH₂₀N₂OS (300.42): C, 67.97; H, 6.71; N, 9.32. Found: C, 68.19; H, 6.47; N, 9.11. ¹H NMR (CDCl₃) δ (ppm): 0.91 (3H, s, Me-6), 1.26 (3H, s, Me-6), 1.33 (IH, d, H-4, J = 11.6 Hz), 2.00-2.13 (2H, m), 2.28 (IH, dd, J = 14.3; 9.8 Hz), 2.35-2.41 (IH, m), 2.78 (IH, dd, J 5 = 14.6; 4.0 Hz), 3.26 (IH, t, J= 9.6 Hz), 4.20 (IH, d, J= 10.6 Hz), 7.15-7.47 (m, 5H). ¹³C NMR (CDCl₃) δ (ppm): 20.6 (Me), 26.2 (CH₂), 26.8 (Me), 28.4 (CH₂), 33.3, 40.4 (C_q), 40.7 (CH), 48.1 (CH), 53.8 (CH), 129.2 (CH₃₁), 129.4 (CH₃₁), 129.6 (CH₃₁), 139.8 (C_q), 170.6 (C=O), 181.3 (C=S).

Example 5 10 {\R,1R,1S,9R)-(10,1 O-Dimethyl-S-oxa-S-azatricyclo [7.1.1.0²'⁷] undec-4-ylidene)phenylamine

(compound 8a) (Scheme 5)

0.46 g (2.7 mmol) of (lΛ,2Λ,35,5Λ)-(2-amino-6,6-dimethylbicyclo[3.1.1]hept-3-yl)methanol (7a) and 0.39 g (2.8 mmol) of phenyl isothiocyanate were dissolved in 40 ml of toluene. The reaction mixture was stirred at room temperature for 3 h, then evaporated to dryness. The resulting product was 15 washed with n-hexane and filtered off. The resulting 0.52 g (1.7 mmol) of thiourea intermediate compound was dissolved in 20 ml of dry methanol, and 0.58 ml (9.3 mmol) of iodomethane was added to the solution in one portion. The reaction mixture was stirred at room temperature for 3 h, then evaporated to dryness. The resulting semisolid material was dissolved in 20 ml of 2,5N methanolic solution of potassium hydroxide. The mixture was stirred for further 4 h at room temperature, then evaporated to 20 dryness. The remaining crude product was dissolved in 30 ml of water and extracted with chloroform (3x30 ml). The combined organic layer was dried (Na₂SO₄), then filtered and evaporated resulting in white crystalline product which was purified by recrystallization from n-hexane.

Isolated compound: 0.38 g (78%); mp: 164-166 ⁰C; [α£° = +32.7 (c = 0.5, MeOH); IR = 2915, 1667, 1587, 1223, 763 cm^"1. Anal. Calcd. for C₁₇H₂₂N₂O (270.37): C, 75.52; H, 8.20; N, 10.36. Found: C, 25 75.87; H, 8.25; N, 10.11. ¹H NMR (CDCl₃) δ (ppm): 0.91 (3H, s, Me-6), 1.24 (3H, s, Me-6), 1.34 (IH, d, H-4, J = 10.7 Hz), 1.37-1.41 (m, IH), 1.92-2.17 (4H, m), 2.45-2.65 (IH, m), 3.86-4.12 (3H, m), 6.95-7.25 (5H, m). ¹³C NMR (CDCl₃) δ (ppm): 20.8 (Me), 25.9 (CH₂), 26.9 (Me), 28.6 (CH₂), 28.7 (CH), 39.4 (C_q), 40.9 (CH), 47.2 (CH), 52.2 (CH), 71.5 (CH₂), 122.6 (CH₃₁), 123.5 (CH₈₁), 129.1 (CH₃₁), 131.6 (C_q), 151.6 (C=N). 30

The starting material (lΛ,2Λ,35,5/?)-(2-amino-6,6-dimethyl-bicyclo[3.1.1]hept-3-yl)methanol (7a) was prepared as below:

To a slurry of 0.93 g (24.5 mmol) of LiAlH₄ in 150 ml of dry THF, 2.00 g (9.5 mmol) of (IR,2R,3S,5R) amino ester liberated from compound 4 (prepared according to Example 3) were added 35 dropwise at 0 ⁰C. After stirring at room temperature for 1.5 h (the reduction was monitored by means of TLC), the mixture was decomposed with the mixture of 10 ml of THF and 2.0 ml of water under ice cooling. The inorganic material was filtered off and washed with THF. After drying (Na₂SO₄) and evaporation, pale-yellow oil was obtained. The amino alcohol obtained was purified as the hydrochloride with recrystallizing from ethyl ether/ethanol mixture. Isolated compound: 1.45 g (70%); mp: 179-183 ⁰C; [α]²,⁰ = -16.4 (c = 0.5, MeOH); IR = 3123, 2917, 1529, 1457, 1051 cm^"1.

Example 6

5 (15,25,7Λ,95)-(10,10-Dimethyl-5-oxa-3-azatricyclo[7.1.1.0²'⁷]undec-4-ylideπe)phenyIamine

(compound 15) (Scheme 5) a) (15,25,5i?,75)-8,8-Dimethyl-3-azatricyclo[5.1.1.0²'^s]nonan-4-one

A mixture of 12.21 g (100.0 mmol) of (+)-(lΛ,5Λ)-apopinene (compound e), prepared via literature method [Lightner, D.A. and Crist, B.V. Tetrahedron, 41, 3021-3028 (1985)], and 14.30 g (101.2

10 mmol) of chlorosulfonyl isocyanate (CSI) was stirred in 300 ml of dry diethyl ether for 48 h at room temperature. 20.4 g (162 mmol) of dry sodium sulfite in 140 ml water was then cautiously added dropwise to the solution while the pH was held at 7-8 by the addition of 20% aqueous potassium hydroxide. After 2 h stirring at the appropriate pH, the organic phase was separated and the aqueous layer was extracted with diethyl ether (2x100 ml). The combined organic layer was dried (Na₂SO₄) and

15 evaporated, and the white crystalline product obtained was recrystallized from n-hexane.

Isolated compound: 13.54 g (82%); mp: 68-72 ⁰C; [αβ° = +61.5 (c = 0.5, MeOH; ee = 90%); IR = 3247, 2914, 1710, 1380, 1256, 1189 cm^"1. Anal. Calcd. for C₁₀H₁₅NO (165.23): C, 72.69; H, 9.15; N, 8.48. Found: C, 72.52; H, 9.27; N, 8.31. ¹H-NMR (CDCl₃) δ (ppm): 0.88 (3H, s, Me-8), 1.30 (3H, s, Me- 8), 1.50 (IH, d, H-9, J = 11.1 Hz), 1.82-1.98 (2H, m), 2.07-2.29 (3H, m), 3.28 (IH, dd, H-5, J = 10.32

20 Hz), 3.95-4.00 (IH, m), 5.87 (IH, s, NH). ¹³C NMR (CDCl₃) δ (ppm): 19.7 (Me-6), 23.3 (CH₂), 24.7 (CH₂), 26.8 (Me-6), 40.0 (C_q), 41.9 (CH), 43.7 (CH), 44.9 (CHC=O), 51.8 (CHN), 173.9 (C=O). b) Ethyl (lS^S^SSVl-amino-ό^-dimethylbicyclop.l.l.lheptane-S-carboxylate hydrochloride

1.16 g (7.0 mmol) of (15,25,5Λ,75)-8,8-dimethyl-3-azatricyclo[5.1.1.0²'⁵]nonan-4-one, prepared

25 according to Example 6a, was stirred in solution of 10% hydrochloric acid in dry ethanol (20 ml) at room temperature. After 1.5 h stirring the solution was evaporated to dryness and the crystalline product obtained was recrystallized from diisopropyl ether/ethyl acetate mixture.

Isolated compound: 1.54 g (89%); mp: 138-139 ⁰C; [α£° = -19.6 (c = 0.5, MeOH); IR = 2918, 1729, 1373, 1189 cm^'1. Anal. Calcd. for C₁₂H₂₂ClNO₂ (247.76): C, 58.17; H, 8.95; N, 5.65. Found: C,

30 58.43; H, 9.26; N, 5.51. ¹H NMR (CDCl₃) δ (ppm): 0.97 (3H, s, Me-6), 1.36 (3H, s, Me-6), 1.35 (3H, t, CH₂-CH₃, J = 7.3 Hz), 1.56 (IH, d, H-4, J = 11.1 Hz), 2.08-2.23 (3H, m), 2.35-2.46 (2H, m), 3.51 (IH, dt, J = 10.1; 3.0 Hz), 4.08 (IH, d, J = 9.6 Hz), 4.24-4.35 (2H, m, CH₂-CU₃). ¹³C NMR (CDCl₃) δ (ppm): 13.7 (Me), 19.7 (Me), 24.4 (CH₂), 25.8 (CH), 29.4 (CH₂), 34.7 (CH), 39.4 (C_q), 39.6 (Me), 44.2 (CH), 50.4 (CH), 63.0 (CH₂), 176.7 (C=O).

35 c) (15,25,3jR,55)-(2-Amino-6,6-diinethyl-bicyclo[3.1.1hept-3-yl)inethanol hydrochloride

To a slurry of 0.93 g (24.5 mmol) of LiAlH₄ in 150 ml of dry THF, 2.00 g (9.5 mmol) of (\S,2S,3R,5S) amino ester liberated from its hydrochloride salt (prepared according to Example 6b) was added dropwise at 0 ⁰C. After stirring at room temperature for 1.5 h (the reduction was monitored by means of TLC), the mixture was decomposed with the mixture of 10 ml of THF and 2.0 ml of water under ice cooling. After 1 h standing the inorganic material was filtered off and washed with THF. After drying (Na₂SO₄) and evaporation, a pale-yellow oil was obtained. The amino alcohol obtained was purified as the hydrochloride with recrystallizing from diethyl ether/ethanol mixture.

Isolated compound: 1.45 g (70%); mp: 179-183 ⁰C; [αβ⁰ = +13.4 (c = 0.5, MeOH); IR = 3123, 2917, 1529, 1457, 1051 cm^'1. Anal. Calcd. for C₁₀H₂₀C1NO (205.72): C, 58.38; H, 9.80; N, 6.81. Found: C, 58.61; H, 10.11; N, 6.49. ¹H NMR (CDCl₃) δ (ppm): 0.95 (3H, s, Me-6), 1.15 (IH, d, H-4, J = 11.1 Hz), 1.28 (3H, s, Me-6), 1.44 (IH, dt, J = 4.0; 14.1 Hz, H-7), 1.97-2.03 (IH, m, H-5), 2.09-2.18 (2H, m, H-I, H-7), 2.27-2.34 (IH, m, H-4), 2.59-2.70 (IH, m, H-3), 3.73 (2H, ddd, J = 5.0; 11.58; 40.2 Hz, CH₂- OH), 3.98 (IH, d, J = 9.6 Hz, H-2). ¹³C NMR (CDCl₃) δ (ppm): 19.9 (Me), 25.2 (CH₂), 25.8 (Me), 29.0 (CH₂), 30.3 (CH), 38.6 (C_q), 40.0 (CH), 45.0 (CH), 52.9 (CH), 65.0 (CH₂). d) (15,25,7Λ,95)-(10,10-Dimethyl-5-oxa-3-azatricyclo[7.1.1.0^2>7]-undec-4-ylidene)phenyl- amine (compound 15)

0.46 g (2.7 mmol) of (15,25,3Λ,55)-(2-amino-6,6-dimethylbicyclo[3.1.1]hept-3-yl)methanol, prepared according to Example 6c, and 0.39 g (2.8 mmol) of phenyl isothiocyanate were dissolved in 40 ml of toluene. The mixture was stirred at room temperature for 3 h, then evaporated to dryness, and the white crystalline substance was filtered and washed with M-hexane. The resulting 0.52 g (1.7 mmol) of thiourea intermediate compound was dissolved in 20 ml of dry methanol, then 0.58 ml (9.3 mmol) of iodomethane was added to the solution in one portion. The reaction mixture was stirred at room temperature for 3 h, then evaporated to dryness. The resulting semisolid material was dissolved in 20 ml of 2,5N methanolic solution of potassium hydroxide. The mixture was stirred for further 4 h at room temperature, then evaporated to dryness. The remaining crude product was dissolved in 30 ml of water and extracted with chloroform (3x30 ml). The combined organic layer was dried (Na₂SO₄), then filtered and evaporated resulting in a white crystalline product which was purified by recrystallization from n- hexane. Isolated compound: 0.38 g (78%); mp: 164-166 ⁰C; [aft⁰ = -9 (c = 0.25, MeOH); IR = 2915,

1667, 1587, 1223, 763 cm ¹). Anal. Calcd. for C₁₇H₂₂N₂O (270.37): C, 75.52; H, 8.20; N, 10.36. Found: C, 75.87; H, 8.25; N, 10.11. ¹H NMR (CDCl₃) δ (ppm): 0.91 (3H, s, Me-6), 1.24 (3H, s, Me-6), 1.34 (IH, d, H-4, J = 10.7 Hz), 1.37-1.41 (m, IH), 1.92-2.17 (4H, m), 2.45-2.65 (IH, m), 3.86-4.12 (3H, m), 6.95- 7.25 (5H, m). ¹³C NMR (CDCl₃) δ (ppm): 20.8 (Me), 25.9 (CH₂), 26.9 (Me), 28.6 (CH₂), 28.7 (CH), 39.4 (C_q), 40.9 (CH), 47.2 (CH), 52.2 (CH), 71.5 (CH₂), 122.6 (CH_2n), 123.5 (CH₃₁), 129.1 (CH₃,), 131.6 (C_q), 151.6 (C=N).

Example 7

(lΛ^ΛJ^PΛJ-ta-Methyl-lO^O-dimethyl-S-oxa-S-azatricyclolXl.l.O^^undec^-ylidene)- phenylamine (compound 8b) (Scheme 5)

The synthesis of compound 8b was accomplished by analogy with Example 5, starting from 0.37 g (2.02 mmol) of (lΛ,2Λ,35,5Λ)-(2-methylamino-6,6-dimethylbicyclo[3.1.1]hept-3-yl)-methanol (compound 7b) and 0.29 g (2.12 mmol) of phenyl isothiocyanate. The resulting crude product was recrystallized from n-hexane/ethyl acetate mixture to a white crystalline product. Isolated compound: 0.31 g (54%); mp: 89-91 ⁰C; [αβ° = -168 (c = 0.25, MeOH); IR: 3062,

2948, 2870, 1924, 1636, 1584, 1056, 750, 693. Anal. Calcd. for C₁₈H₂₄N₂O (284.19): C, 76.02; H, 8.51; N, 9.85. Found: C, 75.67; H, 8.40; N, 10.03. ¹H NMR (CDCl₃) δ (ppm): 0.96 (3H, s), 1.21 (IH, d, J = 10.6 Hz), 1.28 (3H, s), 1.65-1.73 (IH, m), 1.93-1.97 (IH, m), 2.11-2.21 (2H, m), 2.35 (IH, q, J = 5.5, 9.1 5 Hz), 2.67-2.76 (IH, m), 2.88 (3H, s), 3.70 (IH, ddd, J = 1.5, 3.5, 10.6 Hz), 3.81 (IH, dd, J = 4.0, 10.6 Hz), 4.08 (IH, dd, J = 4.5, 10.6 Hz), 6.90-6.96 (3H, m), 7.19-7.25 (2H, m). ¹³C NMR (CDCl₃) δ (ppm): 21.1 (Me), 26.6 (CH₂), 27.7 (Me), 29.8 (CH), 31.7 (CH₂), 36.0 (CH), 39.6 (C_q), 40.9 (CH), 44.7 (CH), 60.1 (Me), 70.0 (CH₂), 122.2 (CH_a,), 124.1 (CH₂₁), 129.0 (CH₃₁), 149.2 (C_q), 154.7 (C=N). MS (m/z): 284.2 (M⁺), 149.1, 106.1, 93.1, 41.0. 10

The starting material (l/?,2Λ,35,5Λ)-(2-methylamino-6,6-dimethylbicyclo[3.1.1]hept-3- yl)methanol (7b) was prepared as below:

To a slurry of 2.82 g (74.32 mmol) OfLiAlH₄ in 150 ml of dry THF 10 ml THF solution of 4.78 g (16.9 mmol) of N-Boc amino acid were added dropwise at room temperature. After stirring at room 15 temperature for 6 h (the reduction was monitored by means of TLC), the mixture was decomposed with the mixture of 30 ml of THF and 6.0 ml of water under ice cooling. The inorganic material was filtered off and washed with THF. After drying (Na₂SO₄) and evaporation, a pale-yellow oil was obtained. The amino alcohol was purified as the hydrochloride with recrystallizing from diethyl ether/ethanol mixture.

1.44 g (39%): mp: 192-193 ⁰C; [D]*⁰ = -15.7 (c = 0.5, MeOH); IR = 3308, 3123, 2916, 2475, 20 1595, 1458, 1049 cm^"'.

Example 8

(lΛ,2Λ,75,9Λ)-(3-Benzyl-10,10-dimethyl-5-oxa-3-azatricyclo[7.1.1.0^2>7]undec-4-ylidene)- phenylamine (compound 8c) (Scheme 5)

25 The synthesis of compound 8c was accomplished by analogy with Example 5, starting from 0.80 g (3.1 mmol) of (lΛ,2Λ,35,5Λ)-(2-benzylamino)-6,6-dimethylbicyclo[3.1.1]heptan-3-yl)methanol (7c) and 0.44 g (3.26 mmol) of phenyl isothiocyanate. The resulting crude product was recrystallized from n- hexane/ethyl acetate mixture.

Isolated compound: 0,41 g (36%); mp: 105-108 ⁰C; [αβ⁰ = -35 (c = 0.25, MeOH); IR: 2940,

30 2918, 2860, 1630, 1579, 1264, 1098, 993, 693. Anal. Calcd. for C₂₄H₂₈N₂O (360.22): C, 79.96; H, 7.83; N, 7.77. Found: C, 79.83; H, 8.01; N, 7.52. ¹H NMR (CDCl₃) δ (ppm): 0.84 (3H, s), 1.25 (3H, s), 1.32 (IH, d, J = 11.1 Hz), 1.72-1.78 (IH, m), 1.91-1.96 (IH, m), 2.12-2.23 (2H, m), 2.32-2.37 (IH, m), 2.63- 2.73 (IH, m), 3.69-3.73 (IH, m), 3.83 (IH, dd, J= 3.0, 10.6 Hz), 4.08 (IH, d, J = 15.1 Hz), 4.15 (IH, dd, 7 = 4.5, 10.6 Hz), 5.13 (IH, d, J = 15.1 Hz), 6.90-6.98 (3H, m), 7.19-7.38 (7H, m). ¹³C NMR (CDCl₃) δ

35 (ppm): 21.0 (Me), 27.0 (CH₂), 27.6 (Me), 30.0 (CH), 32.0 (CH₂), 39.8 (C_q), 40.8 (CH), 44.7 (CH), 50.3 (CH₂), 57.1 (CH), 70.1 (CH₂), 122.3 (CH₃,), 124.1 (CH₃₁), 127.7 (CH₃₁), 128.6 (CH₃,), 129.0 (CH₃,), 139.1 (C,), 149.0 (C,), 154.3 (C=N). MS (m/z): 360.2 (M⁺), 224.1, 132.1, 106.1, 91.1. The starting material 7c (lΛ,2Λ,35,5/?)-(2-benzylamino)-6,6-dimethylbicyclo[3.1.1]heptan-3- yl)methanol was prepared as below:

To a slurry of 0.78 g (20.4 mmol) OfLiAlH₄ in 50 ml of dry THF 25 ml THF solution of 3.06 g (10.2 mmol) of (IR,2R,3S,5R) N-benzyl amino ester were added dropwise at room temperature. After 5 stirring at room temperature for 4 h (the reduction was monitored by means of TLC), the mixture was decomposed with the mixture of 20 ml of THF and 2.0 ml of water under ice cooling. After 1 h standing the inorganic material was filtered off and washed with THF. After drying (Na₂SO₄) and evaporation of the filtrate, a pale-yellow oil was obtained. The amino alcohol was purified as the hydrochloride with recrystallizing from diethyl ether/ethanol mixture.

10 Isolated compound: 1.82 g (61%); mp: 252-253 ⁰C; [a]™ = -8.5 (c = 0.5, MeOH); IR = 3177,

2927, 2741, 1597, 1457, 1048 cm^"1.

Example 9

(l/-,2Λ,7-?,9Λ)-(10,10-Dimethyl-5-oxa-3-azatricydo[7.1.1.0^2>7]undec-4-yIidene)-(3-chloro- 15 phenyl)amine (compound 9a) (Scheme 5)

The synthesis of compound 9a was accomplished by analogy with Example 5, starting from 0.60 g (3.5 mmol) of (17?,2Λ,35,5Λ)-(2-methylamino-6,6-dimethylbicyclo[3.1.1]hept-3-yl)methanol (7a), prepared according to Example 5, and 0.66 g (3.85 mmol) of 3-chlorophenyl isothiocyanate.

Isolated compound: 0.54 g (51%); mp: 157-160 ⁰C; [α£° = +4 (c = 0.25, MeOH); IR = 2904, 20 1670, 1584, 1219, 781 cm^'1. Anal. Calcd. for C₁₇H₂₁ClN₂O (304.81): C, 66.99; H, 6.94; N, 9.19; Found: C, 67.27; H, 6.78; N, 9.35. ¹H NMR (CDCl₃) δ (ppm): 0.89 (3H, s), 1.18 (3H, s), 1.32 (IH, d, J = 10.7 Hz), 1.33-1.45 (m, IH), 1.72-1.81 (IH, m), 1.90-1.99 (IH, m), 2.01-2.18 (2H, m), 2.49-2.65 (IH, m), 3.74-4.12 (3H, m), 6.87 (IH, d, J = 7.3 Hz), 6.93 (IH, d, J = 7.3 Hz), 7.03 (IH, s), 7.14 (IH, t, J = 7.5 Hz). 25

Example 10

(lΛ,2Λ,75,9Λ)-(3-Methyl-10,10-dimethyl-5-oxa-3-azatricyclo[7.1.1.0²'⁷]undec-4-ylidene)-3- chlorophenylamine hydrochloride (compound 9b) (Scheme 5)

The synthesis of compound 9b was accomplished by analogy with Example 5, starting from 0.37

30 g (2.02 mmol) of (l/?,2Λ,35,5Λ)-(2-methylamino-6,6-dimethylbicyclo[3.1.1]hept-3-yl)methanol (7b), prepared according to Example 7, and 0.36 g (2.12 mmol) of 3-chlorophenyl isothiocyanate. The prepared crude product was purified as the hydrochloride with recrystallizing from diethyl ether/ethanol mixture to a trasparent product.

Isolated compound: 0.35 g (55%); mp: 187-191 ⁰C; [α£° = -181 (c=0.25, MeOH); IR: 2988,

35 2914, 2582, 1654, 1593, 1521, 1055, 736. Anal. Calcd. for Ci₈H₂₃ClN₂O (318.15): C, 67.81; H, 7.27; N, 8.79; C, 67.92; H, 7.10; N, 8.88. ¹H NMR (CD₃OD) δ (ppm): 0.83 (IH, d, J = 11.08 Hz), 1.12 (3H, s), 1.40 (3H, s), 1.78 (IH, ddd, J = 2.52, 6.55, 13.60 Hz), 2.05-2.10 (IH, m), 2.34-2.43 (2H, m), 2.53 (IH, q, J= SM, 9.57 Hz), 3.15-3.32 (4H, m), 4.25-4.32 (IH, dd, J = 3.02, 10.58 Hz), 4.32-4.37 (IH, dd, J= 2.52, 10.58 Hz), 4.55-4.60 (IH, dd, J = 4.03, 10.58 Hz), 7.33-7.41 (2H, m), 7.43-7.50 (2H, m). ¹³C NMR (CD₃OD) δ (ppm): 21.1 (Me), 27.6 (CH₂), 27.8 (Me), 29.5 (CH), 32.1 (CH₂), 37.2 (CH), 40.6 (C_q), 41.7 (CH), 45.8 (CH), 63.2 (Me), 73.3 (CH₂), 124.2 (CHJ, 125.8 (CU₃₁), 128.4 (CH₃₁), 131.8 (CH₃₁), 135.8 (CCl), 137.4 (C_q), 161.5 (C=N). MS (m/z): 318.2 (M⁺), 183.0, 149.1, 140.0, 93.1, 78.0, 63.0, 36.0.

Example 11

(lΛ,2Λ,75,9Λ)-(3-Benzyl-10,10-dimethyI-5-oxa-3-azatricyclo[7.1.1.0^2l7]undec-4-ylidene)-3- chlorophenylamine (compound 9c) (Scheme 5)

The synthesis of compound 9c was accomplished by analogy with Example 5, starting from 0.42 g (1.63 mmol) of (l/?,2/?,35,5/?)-(2-benzylamino-6,6-dimethylbicyclo[3.1.1]hept-3-yl)methanol (7c), prepared according to Example 8, and 0.29 g (1.71 mmol) of rø-chlorophenyl isothiocyanate. The obtained crude product was purified by flash chromatography (silica gel, n-hexane/ethyl acetate 4:1) resulting in a white crystalline product.

Isolated compound: 0.25 g (39%); mp: 95-97 ⁰C; [aft⁰ = -64 (c=0.25, MeOH); IR: 2988, 2925,

2364, 1635, 1585, 1239, 773. Anal. Calcd. for C₂₄H₂₇ClN₂O (394.18): C, 72.99; H, 6.89; N, 7.09; Found: C, 73.21; H, 6.55; N, 7.17. ¹H NMR (CDCl₃) δ (ppm):0.85 (3H, s), 1.22-1.29 (4H, m), 1.76 (IH, ddd, J =

2.5, 5.0, 13.6 Hz), 1.92-1.98 (IH, m), 2.13-2.23 (2H, m), 2.34 (IH, q, J = 5.5, 8.6 Hz), 2.65-2.75 (IH, 7«,),

3.72 (IH, dd, J = 2.0, 10.6 Hz), 3.86 (IH, dd, J = 3.0, 10.6 Hz), 4.08 (IH, d, J= 15.1 Hz), 4.15 (IH, dd, J

= 4.5, 10.6 Hz), 5.10 (IH, d, J= 15.1 Hz), 6.80 (IH, d, J= 8.1 Hz), 6.87-6.96 (2H, m), 7.12 (IH, t, J= 8.1

Hz), 7.23-7.37 (5H, m). ¹³C NMR (CDCl₃) δ (ppm): 21.0 (Me), 27.0 (CH₂), 27.6 (Me), 29.8 (CH), 32.0 (CH₂), 39.8 (C_q), 40.8 (CH), 44.7 (CH), 50.4 (CH₂), 57.2 (CH), 70.2 (CH₂), 122.2 (CH₃,), 122.5 (CH₃₁),

124.2 (CH₃,), 127.8 (CH₃,), 128.5 (CH₈₁.), 129.1 (CH₃,), 129.9 (CH₃,), 134.4 (CCl), 138.8 (C_q), 150.4 (C_q),

154.7 (C=N). MS (m/z): 394.2 (M⁺), 259.1, 167.0, 149.0, 132.1, 106.1, 91.0, 57.1, 32.0.

Example 12 (lΛ,2Λ,75,9Λ)-(10,10-DimethyI-5-oxa-3-azatricyclo[7.1.1.0²'⁷]undec-4-ylidene)-(3-methoxy- phenyl)amine (compound 10a) (Scheme 5)

The synthesis of compound 10a was accomplished by analogy with Example 5, starting from 0.50 g (2.95 mmol) of (lΛ,2Λ,35,5Λ)-(2-amino-6,6-dimethylbicyclo[3.1.1]hept-3-yl)methanol (7a), prepared according to Example 5, and 0.49 g (3.25 mmol) of 3-methoxyphenyl isothiocyanate. Isolated compound: 0.39 g (44%): mp: 139-140 ⁰C; [a]™ = -7 (c = 0.25, MeOH); IR = 3202,

2904, 1665, 1596, 1263, 1091, 773 cm^'1. Anal. Calcd. for C₁₈H₂₄N₂O₂ (300,40): C, 71.97; H, 8.05; N, 9.33; Found: C, 71.81; H, 8.39; N, 9.31. ¹H NMR (CDCl₃) δ (ppm): 0.91 (3H, s), 1.23 (3H, s), 1.34 (IH, d, J = 10.7 Hz), 1.34-1.47 (m, IH), 1.76-2.20 (4H, m), 2.51-2.74 (IH, m), 3.77 (3H, s), 3.81-4.20 (3H, m), 6.45-6.73 (3H, m), 7.09-7.22 (IH, m).

Example 13

(l/?,2Λ,75,9Λ)-(10,10-Dimethyl-5-oxa-3-azatricyclo[7.1.1.0^2>7]undec-4-ylidene)-(4-methyl- phenyl)amine (compound Ha) (Scheme 5) The synthesis of compound 11a was accomplished by analogy with Example 5, starting from 0.50 g (2.95 mmol) of (lΛ,2Λ,35,5Λ)-(2-amino-6,6-dimethylbicyclo[3.1.1]hept-3-yl)methanol (7a), prepared according to Example 5, and 0.44 g (2.95 mmol) of 4-methylphenyl isothiocyanate.

Isolated compound: 0.56 g (67%); mp: 188-189 ⁰C; [Ct]J!,⁰ = +8 (c = 0.25, MeOH); IR = 2902, 1684, 1509, 1223 cm^"1. Anal. Calcd. for C₁₈H₂₄N₂O (284.40): C, 76.02; H, 8.51; N, 9.85; Found: C,

75.81; H, 8.27; N, 9.56. ¹H NMR (CDCl₃) δ (ppm): 0.90 (3H, s), 1.22 (3H, s), 1.34 (IH, d, J = 10.7 Hz),

1.35-1.48 (IH, m), 1.85-2.19 (4H, m), 2.28 (3H, s), 2.49-2.73 (IH, m), 3.68-4.1 (3H, m), 6.80-7.10 (4H, m).

Example 14

(lΛ,2iϊ,75,9Λ)-(10,10-Dimethyl-5-oxa-3-azatricyclo[7.1.1.0^2l7]undec-4-ylideneH4-fluoro- phenyl)amine (compound 12a) (Scheme 5)

The synthesis of compound 12a was accomplished by analogy with Example 5, starting from 0.60 g (3.5 mmol) of (lΛ,2/?,35,5Λ)-(2-amino-6,6-dimethylbicyclo[3.1.1]hept-3-yl)methanol (7a), prepared according to Example 5, and 0.59 g (3.85 mmol) of 4-fluorophenyl isothiocyanate.

Isolated compound: 0.57 g (56%); mp: 224-225 ⁰C; [<xf° = +17 (c = 0.25, MeOH); IR = 2930, 1666, 1505, 1208, 849 cm^"1. Anal. Calcd. for C₁₇H₂₁FN₂O (288.36): C, 70.81; H, 7.34; N, 9.71; Found: C, 71.13; H, 7.02; N, 9.56. ¹H NMR (CDCl₃) δ (ppm): 0.92 (3H, s), 1.25 (3H, s), 1.35 (IH, d, J = 10.7 Hz), 1.37-1.47 (TM, IH), 1.87-2.24 (4H, m), 2.53-2.71 (IH, m), 3.80-4.18 (3H, m), 6.87-7.07 (4H, m).

Example 15

(1 Jt,2i-,75,9il)-Ethyl-(l 0,10-dimethyl-5-oxa-3-azatricyclo [7.1.1.0²'⁷] undec-4-ylidene)amine hydrochloride (compound 13a) (Scheme 6)

The synthesis of compound 13a was accomplished by analogy with Example 5, starting from 0.60 g (3.5 mmol) of (l/?,2^,35,5/;)-(2-amino-6,6-dimethylbicyclo[3.1.1]hept-3-yl)methanol (7a), prepared according to Example 5, and 0.34 g (3.85 mmol) of ethyl isothiocyanate. The prepared crude product was purified from the base as the hydrochloride salt with recrystallizing from ethanolic hydrochloric solution.

Isolated compound: 0.49 g (54%); mp: 180-183 ⁰C; [α]*⁰ = +6 (c = 0.25, MeOH); IR = 2911, 1690, 1589 cm^"1. Anal. Calcd. for C₁₃H₂₃ClN₂O (258.79): C, 60.33; H, 8.96; N, 10.82; Found: C, 60.57; H, 8.71; N, 10.97. ¹H NMR (CDCl₃) δ (ppm): 1.00 (3H, s), 1.24 (3H, t, J = l.l Hz), 1.34 (3H, s), 2.03- 2.36 (5H, m), 2.83-2.99 (IH, m), 3.25-3.45 (2H, m), 3.99-4.60 (3H, m).

Example 16 (lΛ,2Λ,7Λ,9Λ)-(10,10-DimethyI-5-oxa-3-azatricyclo[7.1.1.0^2>7]undec-4-ylidene)phenylamine

(compound 17) (Scheme 7)

The synthesis of compound 17 was accomplished by analogy with Example 5, starting from 0.46 g (2.7 mmol) of (lΛ,2Λ,3Λ,5Λ)-(2-amino-6,6-dimethylbicyclo[3.1.1]hept-3-yl)methanol (16) and 0.39 g (2.8 mmol) of phenyl isothiocyanate. Isolated compound: 0.47 g (64%); mp: 118-121 ⁰C; [α]*° = +77 (c = 0.25, MeOH); IR = 2904,

1666, 1590, 1205, 695 cm ¹. Anal. Calcd. for C₁₇H₂₂N₂O (270.37): C, 75.52; H, 8.20; N, 10.36; Found: C,

75.68; H, 8.41; N, 10.56. ¹H NMR (CDCl₃) δ (ppm): 0.84 (3H, s), 1.31 (3H, s), 1.46 (IH, t, J = 12.0 Hz),

1.76 (IH, d, J = 10.6 Hz), 1.80-1.87 (IH, m), 1.97-2.31 (4H, w), 3.51 (IH, d, J= 9.7 Hz), 4.18 (IH, dd, J = 12.4, 9.2 Hz), 4.36 (IH, dd, J = 8.7, 5.1 Hz), 6.86-7.31 (5H, tή).

The starting material (l/?,2Λ,3Λ,5Λ)-(2-amino-6,6-dimethyl-bicyclo[3.1.1]hept-3-yl)methanol (16) was prepared as below:

To a slurry of 0.93 g (24.5 mmol) of LiAlH₄ in 150 ml of dry THF 2.00 g (9.5 mmol) of (IR,2R,3R,5R) amino ester liberated from its hydrochloride salt was added dropwise at 0⁰C. After stirring at room temperature for 1.5 h (the reduction was monitored by means of TLC), the mixture was decomposed with the mixture of 10 ml of THF and 2.0 ml of water under ice cooling. After 1 h standing the inorganic material was filtered off and washed with THF. After drying (Na₂SO₄) and evaporation, a pale-yellow oil was obtained. The amino alcohol obtained was purified as the hydrochloride with recrystallizing from diethyl ether/ethanol mixture. Isolated compund: 1.62 g (78%); mp: 199-202 ⁰C; [a]™ = -7.9 (c = 0.52, MeOH); IR = 3298,

2905, 1512, 1040 cm^"1.

Example 17 (15^l,25,75,95)-(10,10-Dimethyl-5-oxa-3-azatricyclo[7.1.1.0²'⁷]undec-4-ylidene)phenylainine (compound 19) (Scheme 8) a) (15,25,5Λ,75)-8,8-Dimethyl-3-a2atricyclo[5.1.1.0²'⁵]nonan-4-one

A mixture of 12.21 g (100.0 mmol) of (+)-(lΛ,5/?)-apopinene (compound b), prepared via literature method [Lightner, D.A.; Crist, B.V. Tetrahedron, 41, 3021-3028 (1985)], and 14.30 g (101.2 mmol) of chlorosulfonyl isocyanate (CSI) was stirred in 300 ml of dry diethyl ether for 48 h at room temperature. 20.4 g (162 mmol) of dry sodium sulfite in 140 ml water was then cautiously added dropwise to the solution while the pH was held at 7-8 by the addition of 20% aqueous potassium hydroxide. After 2 h stirring at the appropriate pH, the organic phase was separated and the aqueous layer was extracted with diethyl ether (2x100 ml). The combined organic layer was dried (Na₂SO₄), then after filtration evaporated and the white crystalline product obtained was recrystallized from diisopropyl ether. Isolated compound: 13.54 g (82%); mp: 68-72 ⁰C; [α]*⁰ = +61.5 (c = 0.5, MeOH; ee = 90%);

IR = 3247, 2914, 1710, 1380, 1256, 1189 cm ¹. Anal. Calcd. for C₁₀H₁₅NO (165.23): C, 72.69; H, 9.15; N, 8.48. Found: C, 72.52; H, 9.27; N, 8.31. ¹H-NMR (CDCl₃) δ (ppm): 0.88 (3H, s, Me-8), 1.30 (3H, s, Me- 8), 1.50 (IH, d, H-9, ^ = 11.1 Hz), 1.82-1.98 (2H, m), 2.07-2.29 (3H, m), 3.28 (IH, dd, H-5, J = 10.32 Hz), 3.95-4.00 (IH, m), 5.87 (IH, s, NH). ¹³C NMR (CDCl₃) δ (ppm): 19.7 (Me-6), 23.3 (CH₂), 24.7 (CH₂), 26.8 (Me-6), 40.0 (C,), 41.9 (CH), 43.7 (CH), 44.9 (CHC=O), 51.8 (CHN), 173.9 (C=O). b) Ethyl (lS^S^^SS^-airύno-δ^-dimethylbicycloβ.l.l.lheptane-S-carboxylate hydrochloride

1.16 g (7.0 mmol) of (l-?,25,5Λ,75)-8,8-dimethyl-3-azatricyclo[5.1.1.0^2>5]nonan-4-one, prepared according to Example 17 a, was stirred in solution of 10% hydrochloric acid in dry ethanol (20 ml) at room temperature. After 1.5 h stirring the solution was evaporated to dryness and the crystalline product obtained was recrystallized from diethyl ether/ethyl acetate mixture.

Isolated compound: 1.54 g (89%); mp: 138-139 ⁰C; [α]∞ = -19.6 (c = 0.5, MeOH); IR = 2918,

1729, 1373, 1189 cm ¹. Anal. Calcd. for C₁₂H₂₂ClNO₂ (247.76): C, 58.17; H, 8.95; N, 5.65. Found: C, 58.43; H, 9.26; N, 5.51. ¹H NMR (CDCl₃) δ (ppm): 0.97 (3H, s, Me-6), 1.36 (3H, s, Me-6), 1.35 (3H, t,

CH₂-CH₃, J = 7.3 Hz), 1.56 (IH, d, H-4, J = 11.1 Hz), 2.08-2.23 (3H, m), 2.35-2.46 (2H, m), 3.51 (IH, dt, J = 10.1; 3.0 Hz), 4.08 (IH, d, J = 9.6 Hz), 4.24-4.35 (2H, m, CH₂-CH₃). ¹³C NMR (CDCl₃) δ (ppm):

13.7 (Me), 19.7 (Me), 24.4 (CH₂), 25.8 (CH), 29.4 (CH₂), 34.7 (CH), 39.4 (C_q), 39.6 (Me), 44.2 (CH), 50.4 (CH), 63.0 (CH₂), 176.7 (C=O). c) Ethyl (l^S^iS^S/yj-l-ainino-όjό-dimethylbicyclop.l.ljheptane-S-carboxylate hydrochloride

To a solution of 0.23 g (10 mmol) of sodium in 30 ml of dry ethanol 1.05 g (5 mmol) of the base of ethyl (15,25,3/?,55)-2-amino-6,6-dimethylbicyclo[3.1.1]_neptane.3._{carboxylate was added} J_{n one} portion. The solution was stirred at room temperature until isomerisation was accomplished (approx. 4 h, the isomerisation process was monitored by means of TLC). The solution was evaporated to approx. 5 ml, diluted with ice-cold water (50 ml) and extracted with ethyl acetate (3x50 ml). The combined organic layer was dried (Na₂SO₄), filtered and evaporated. The hydrochloride salt, prepared from the resulted amino ester base with 15% solution of hydrochloric acid in dry ethanol, was recrystallized from isopropyl ether. Isolated compound: 0.85 g (69%); mp: 147-148°C; [α]^⁰ = +31.0 (c = 0.5, MeOH); IR = 2926,

1734, 1509, 1292, 1193 cm ¹. Anal. Calcd. for C₁₂H₂₂ClNO₂ (247.76): C, 58.17; H, 8.95; N, 5.65; Found: C, 58.35; H, 8.78; N, 5.79. ¹H NMR (CDCl₃) δ (ppm): 0.88 (3H, s, Me-6), 1.34 (3H, s, Me-6), 1.34 (3H, s, Me-6), 1.36 (3H, t, CH₂-CH₃, J = 7.1 Hz), 1.56 (IH, d, H-4, J = 11.1 Hz), 2.05 (IH, dd, J = 13.6, 9.1 Hz), 2.10-2.23 (2H, m), 2.35-2.46 (2H, m), 3.05 (IH, dd, J = 9.1, 18.1 Hz), 4.10 (IH, d, J = 89.6 Hz), 4.33 (2H, dd, J = 7.1; 14.1 Hz, OZ₂-CH₃). ¹³C NMR (CDCl₃) δ (ppm): 13.8 (Me), 19.1 (Me), 23. (CH₂),

25.8 (CH), 27.5 (CH₂), 38.4 (CH), 39.3 (Me), 39.7 (C_q), 43.4 (CH), 52.3 (CH), 63.0 (CH₂), 175.9 (C=O). d) (l-y,25,3iS,55)-(2-Amino-6,6-dimethylbicyclo[3.1.1]hept-3-yl)methanol hydrochloride To a slurry of 0.93 g (24.5 mmol) of LiAlH₄ in 150 ml of dry THF 2.00 g (9.5 mmol) of (IS,2S,3S,5S) amino ester liberated from its hydrochloride salt (prepared according to Example 17c) was added dropwise at 0 ⁰C. After stirring at room temperature for 1.5 h (the reduction was monitored by means of TLC), the mixture was decomposed with the mixture of 10 ml of THF and 2.0 ml of water under ice cooling. The inorganic material was filtered off and washed with THF. After standing for 1 h the inorganic material was filtered and then washed with THF. The filtrate was dried (Na₂SO₄) and then evaporated. The pale-yellow oily amino alcohol was purified as the hydrochloride by recrystallizing from diethyl ether/ethanol mixture.

Isolated compound: 1.62 g (78%); mp: 199-202 ⁰C; [αβ° = +8.1 (c = 0.5, MeOH); IR = 3298, 2905, 1512, 1040 cm^"1. Anal. Calcd. for C₁₀H₂₀C1NO (205.72): C, 58.38; H, 9.80; N, 6.81; Found: C, 58.49; H, 9.71; N, 6.93. ¹H NMR (CDCl₃) δ (ppm): 0.86 (3H, s), 1.32 (3H, s), 1.50-1.59 (IH, m), 1.62 (IH, d, J = 10.6 Hz), 2.03-2.20 (4H, m), 2.28-2.36 (IH, m), 3.63 (IH, d, J= 8.1 Hz), 3.73 (2H, ddd, J = 2.5, 5.5; 11.1 Hz). ¹³C NMR (CDCl₃) δ (ppm): 19.1 (Me), 23.1 (CH₂), 26.2 (Me), 26.3 (CH₂), 35.1 (CH), 89.7 (CH), 40.0 (C_q), 44.2 (CH), 54.6 (CH), 64.5 (CH₂). ^■ e) (15',25,75,95)-(10,10-Dimethyl-5-oxa-3-azatricyclo[7.1.1.0²'⁷]uπdec-4-ylidene)phenyl- amine

0.46 g (2.7 mmol) of (lS,25,35,5S)-(2-amino-6,6-dimethylbicyclo[3.1.1]hept-3-yl)methanol (18) and 0.39 g (2.8 mmol) of phenyl isothiocyanate were dissolved in 40 ml of toluene. The reaction mixture was stirred at room temperature for 3 h, then evaporated to dryness. The resulting white crystalline product was washed with «-hexane and filtered off. The resulting 0.52 g (1.7 mmol) of thiourea intermediate compound was dissolved in 20 ml of dry methanol, and 0.58 ml (9.3 mmol) of iodomethane was added to the solution in one portion. The reaction mixture was stirred at room temperature for 3 h, then evaporated to dryness. The resulting semisolid material was dissolved in 20 ml of 2,5N methanolic solution of potassium hydroxide. The mixture was stirred for further 4 h at room temperature, then evaporated to dryness. The remaining crude product was dissolved in 30 ml of water and extracted with chloroform (3x30 ml). The combined organic layer was dried (Na₂SCj), filtered and evaporated. The obtained crude product was purified by recrystallization from «-hexane, resulting in a white crystalline product.

Isolated compound: 0.47 g (64%); mp: 118-121 ⁰C; [αβ⁰ = -76 (c = 0.25, MeOH); IR = 2904, 1666, 1590, 1205, 695 cm^"1. Anal. Calcd. for C₁₇H₂₂N₂O (270.37): C, 75.52; H, 8.20; N, 10.36; Found: C, 75.68; H, 8.41; N, 10.56. ¹H NMR (CDCl₃) δ (ppm): 0.84 (3H, s), 1.31 (3H, s), 1.46 (IH, t, J= 12.0 Hz)₅ 1.76 (IH, d, J= 10.6 Hz), 1.80-1.87 (IH, m), 1.97-2.31 (4H, m), 3.51 (IH, d, J = 9.7 Hz), 4.18 (IH, dd, J = 12.4, 9.2 Hz), 4.36 (IH, dd, J = 8.7, 5.1 Hz), 6.86-7.31 (5H, in).

In the enclosed Fig. 2 relating to the Summarizing Formula Table the formulae of the compounds described in the Examples are given together with the formula numbers.

BIOLOGICAL ASSAYS

A) Cytostatic effect assay on multiple cell lines

The cytostatic effect of the compounds according to the invention was evaluated in vitro in a concentration range of 3-90 μM on three human tumour cell lines (A431, HeLa, MCF7*). * A431 : human squamosus carcinoma HeLa: human cervix adenocarcinoma MCF7: human breast adenocarcinoma.

The cytostatic MTT-assay is a widely used antiproliferative method in which the test compounds are screened for cytostatic property [Sobottka, S. B. and Berger, M. R.: Cancer Chemotherapy and Pharmacology 30, 385-393 (1992)]. This test is a highly reproducible and high-throughput method carried out on 96-well microplates. A limited amount of human cancer cells (5000/well) was seeded onto the microplate. During an overnight preincubation period the cells sedimented on the bottom of the well. On the second day of the test the original medium was removed and 200 μl of a new medium containing the test substances in a final concentration of 3-90 μM were added. A 30 μM stock solution of each compound to be tested was prepared with dimethyl sulfoxide (DMSO). The final DMSO concentration of the medium was never higher than 0.3% by mass since this value has no substantial background effect on cell proliferation. After an incubation period of 72 hours living cells were investigated by adding 22 μl of an 5 mg/ml MTT solution (methylthiazolyldiphenyl-tetrazolium bromide dissolved in phosphate-buffered sterile saline). MTT is a yellowish dye which was converted by intact mitochondrial reductase and precipitated as blue crystals during a 4-hour contact period. Then the medium was removed and the precipitated crystals were dissolved in 100 μl of DMSO during a 60-minute period of shaking. Finally the reduced MTT was determined at 550 run using a microplate reader. The absorbance values were compared to values obtained for control wells containing only medium. AU in vitro experiments were carried out on two microplates with at least 5 parallel wells.

The cytostatic effect of compounds 8a, 9a, 9b, 10a, 11a, 12a, 15, 17, and 19 was assayed in a concentration range of 3 to 90 μM. Sigmoidal curves were fitted on the results and the IC₅₀ values, that is the concentration of the tested compounds at which 50% of the maximal cytostatic effect could be detected, were calculated by GraphPad Prism 2.01 method. The thus-obtained diagrams are presented for compounds 9a, 9b, 10a, 1 Ia, 12a and 17 on the enclosed drawing (Figs 3 to 5).

On Fig. 6 dose-effect curves are shown for compound 8a as well as doxorubicin and cisplatin on various cell lines.

The calculated IC₅₀ values are shown in Table I. Table I

Bl) Growth inhibition assay on multiple cancer cell lines - single concentration

The inhibiting effect (ID₅₀) of the compounds according to the invention exerted on cancerous cells was tested on six tumour cell lines. The compounds were tested at 10 μM concentration, by a 72 h treatment after 16 h preincubation on the following cell lines: U937 human leukemic monocyte lymphoma

H358 RAS mutant lung cancer

Hl 650 EGFR deletion mutant lung cancer

RD rhabdomyosarcoma

HT29 colorectal cancer

HCTl 16 colorectal cancer

The cell number was 2000/well in 100 μl RPMIl 640+FBS/well, on 96well PerkinElmer black culture plates (#6005668). Method: PerkinElmer ATPlite™ Luminescence Assay System (#6016949) measured on BioTek Synergy II multiplate reader in two independent experiments with 5-5 parallels.

Score is calculated as the sum of the mean viability values of different cell lines. Score above 400 means that the drug had no significant effect, score below 400 and above 200 means that efficiency is significant (33% average growth inhibition on all cell lines), and score below 200 means that the drug has more than 66% average growth inhibitory effect on the cell lines.

The mean viability values are shown in Table II.

Table II

It appears from the Table that most significant effects were shown by compounds of formula 8a, 9a, 10a and 11a with scores below 200.

B2) Growth inhibition assay on multiple cancer cell lines - IC₅₀ determination

The growth inhibition effect (IC₅₀) of some selected compounds according to the invention was tested on six cancer cell lines given under item Bl).

The compounds were tested at 1 μM, 2 μM, 5 μM, 10 μM, and 20 μM concentration according to the protocol under item Bl). Data processing: highest and lowest data from 5 parallels were excluded. Mean values, average and standard deviations were calculated from the 3 remaining data. Viability values are calculated in percentage, they correlate to the mean control value of 100.

The calculated IC₅₀ values are shown in Table III. Table III

Claims

What we claim is:

1. Chiral compounds with monoterpene skeleton of general formula (I) - where in the formula X stands for O or H₂;

W stands for O, S, N-R² or Ph-R³;

Y stands for O or N-R⁴;

R¹ stands for H, C_1-4AIk or (CH₂)_1-4-Ph;

R² stands for C_1-4AIk or Ph-R³; R³ stands for H, C_{1- 4}Alk, C_1-4AIk-O or HIg;

R⁴ stands for H or Ph; and one of the signs — means the presence of a double bond and the other means the absence of a double bond, with the proviso that only one of W and Y may simultaneously stand for oxygen — , as well as their prodrugs and salts formed with pharmaceutically acceptable acids.

2. The following compounds of general formula (I)

- (lR,2R,7S,9R)-( 10, 10-dimethyl-S-oxa-S-aza-tricyclof?.1.1.O²⁷]undec-4-yilidene)-(3-chloro- phenyl)-amine,

- (lR,2R,7S,9R)-( 10, 1 O-dimethyl-S-oxaO-aza-tricyclolT.1.1.0^{2 7}]undec-4-yilidene)-(3-methoxy- phenyl)-amine,

- (lΛ,2Λ,7S,9Λ)-(10,10-dimethyl-5-oxa-3-aza-tricyclo[7.1.1.0²⁷]undec-4-ylidene)-(4-methyl- phenyl)-amine,

- ( lR,2R,7S,9R)-( 10, 10-dimethyl-5-oxa-3-aza-tricyclo[7.1.1.0^{2 7}]undec-4-ylidene)-(4-fluoro- phenyl)-amine, - (1R.2R, 75^r,P/?)-(3-methyl-10,10-dimethyl-5-oxa-3-aza-tricyclo[7.1.1.0^{2 7}]-undec-4-ylidene)-3- chlorophenyl-amine hydrochloride,

- (lR,2R,7R,9R)-( 10, 10-dimethyl-5-oxa-3-aza-tricyclo[7.1.1.0²-⁷]undec-4-ylidene)-phenylamine,

- (lS,2S,7R,9S)-( 10, 10-dimethyl-5-oxa-3-aza-tricyclo[7.1.1.0²-⁷]undec-4-ylidene)-phenylamine, and - (lS,2S,7S,9S)-( 10, 10-dimethyl-5-oxa-3-aza-tricyclo[7.1.1.O^{2 7}]undec-4-yilidene)-phenylamine.

3. A cytostatic pharmaceutical composition comprising as active agent one or more compounds of general formula (I) and usual inert pharmaceutical carriers and/or auxiliary agents.

4. Use of one or more compounds of general formula (I) and/or the salts of such compounds for preparing cytostatic pharmaceutical compositions.

5. Method for treating and/or curing cancerous illnesses characterized in that an effective amount of the composition according to claim 3 is administered to a patient in need of such treatment.