WO2003014117A1

WO2003014117A1 - 3-substituted pyrrolo[2.1-a]isoquinoline derivatives

Info

Publication number: WO2003014117A1
Application number: PCT/US2002/024877
Authority: WO
Inventors: Marcus Bauser; Jens-Kerim ERGÜDEN; Dietmar Flubacher; Paul Naab; Thorsten-Oliver Repp; Jürgen Stoltefuss; Nils Burkhardt; Andrea Sewing; Michael Schauer; Karl-Heinz Schlemmer; Olaf Weber; Stephen J. Boyer; Mark Miglarese; Shihong Ying; Ulrich Niewöhner
Original assignee: Bayer Corporation; Bayer Aktiengesellschaft; NIEWÖHNER, Maria
Priority date: 2001-08-06
Filing date: 2002-08-05
Publication date: 2003-02-20
Also published as: WO2003014115A1

Abstract

The present invention relates to 3-substituted pyrrolo[2.1-a]dihydroisoquinolines which are in-hibitors of phosphodiesterase 10a and can be used for combating cancer.

Description

3-SUBSTITUTED PYRROLO[2.1-a]ISOQUINOLINE DERIVATIVES

This application claims priority from U.S. Provisional Application 60/310,384, filed 6 August 2001.

BACKGROUND OF THE INVENTION

The present invention relates to 3 -substituted pyrrolo[2.1-a]isoquinoline derivatives which are inhibitors of phosphodiesterase 10a, a process for preparing those com- pounds and a method of treating cancer in humans and animals by administering those compounds.

Cyclic AMP metabolism is regulated by the opposing activities of adenylyl cyclase, which generates cAMP in response to extracellular stimuli (e.g. engagement of G- protein coupled receptors by their cognate ligands), and 3 ',5' cyclic nucleotide phos- phodiesterases (PDEs), which hydrolyze cAMP to 5 '-AMP. Signal transduction via cAMP is associated with transcriptional events that can result in the inhibition of cellular proliferation (TJ. Shaw et al, Exp. Cell Res. 273, 95 (2002); T.W. Moody et al, Ann. N.Y. Acad. Sci. 921, 26 (2000); W.L. Lowe et al, Endocrinology. 138, 2219 (1997)); D.A. Albert, J. Clin. Invest. 95, 1490 (1995); M.I. Mednieks et al., FEBS

Lett. 254, 83 (1989). indeed, elevation of intracellular cAMP concentration is growth inhibitory for several human tumor cell lines, including those derived from breast, lung and colorectal carcinomas (B. Wagner et al, Biochem. Pharmacol. 63, 659 (2002); S.B. Jakowlew et al, Peptides 21, 1831 (2000); LS. Fentimen et al., Mol. Biol. Med. 2, 81 (1984); P. Cassoni et al., Int. J. Cancer 72, 340 (1997); S. Shafer et al., Biochem. Pharmacol. 56, 1229 (1998); N.M. Hoosein et al., Regul. Peptides 24, 15 (1989)). In several human breast carcinoma cell lines, increased cAMP production through stimulation of adenylate cyclase activity and/or reduction in cAMP catabolism through inhibition of phosphodiesterase activity has been shown to result in increased steady state levels of cAMP and growth inhibition (D. Melck et al, FEBS Letters 463, 235 (1999); N. Veber et al, Eur. J. Cancer. 30A, 1352 (1994); J.A. Fontana et al, J. Natl. Cancer Just. 78, 1107 (1987); T.A. Slotkin et al., Breast Cancer Res. and Treatment. 60, 153 (2000)). In contrast to breast tumor cell lines, normal human mammary epithelial cells are stimulated to proliferate by elevation of intracellular cAMP (LS. Fentimen et al., Mol. Biol. Med. 2, 81 (1984)). These observations suggest that elevation of intracellular cAMP may selectively inhibit breast tumor cell proliferation. Interestingly, it has been reported that neoplastic mammary tissues have higher levels of low-Km phosphodiesterase activity compared to normal breast tissue, suggesting that tumors may gain a growth or survival advantage by keeping intracellular cAMP levels in check (A.L. Singer et al., Cancer

Res. 36, 60 (1976)).

The ICAST (Inhibitor of Cyclic AMP Signal Transduction) gene encodes a specific 3 ',5 '-cyclic nucleotide phosphodiesterase. Compared to corresponding normal tissues, ICAST mRNA is overexpressed in breast carcinoma specimens, liver metastases of colorectal carcinoma and non-small cell lung carcinomas. The ICAST cDNA was also recently cloned by other groups and named PDElOa (K. Fujishige et al., J. Biol. Chem. 274, 18 438 (1999); S.H. Soderling et al., Proc. Natl. Acad. Sci. USA 96, 7071 (1999); K. Loughney et al., Gene 234, 109 (1999)). Published expression data for ICAST mRNA show a very limited distribution across adult human tissues, with highest levels observed in the testis, caudate nucleus and putamen (K. Fujishige et al., 1999). Increased expression of ICAST mRNA in human tumor specimens indicates that ICAST may play an important role in tumor cell growth and/or survival under conditions of elevated cAMP generation. Selective inhibition of ICAST activity in tumor cells should lead to increased cAMP concentrations and growth inhibition. The expression profile of ICAST and the published reports indicating that breast, lung and colon carcinomas are particularly sensitive to elevation of intracellular cAMP indicate that ICAST may play critical roles specifically in those tumor types. Ln addition to elevation of cAMP, inhibition of ICAST activity should also decrease the intracellular concentration of 5-AMP, which could limit purine pools and DNA synthesis in rapidly dividing tumor cells.

Pyrrolo[2.1-a]isoquinoline derivatives of the formula (A) are described in J. Med. Chem. 27, 1321 (1984) and in J. Med. Chem. 31, 2097 (1988):

R' = H, OMe, CI

R" = H, CI

R^m = H, Me

R"", R" = Me, Et, i -Pr, C_gH^

These compounds are described as having antineoplastic activity, which however is stated to be due to the carbamate moieties being electrophilic centers enabling the compounds (A) to react via an alkyl-oxygen cleavage mechanism. It is not mentioned that these compounds have any PDE 10a inhibitory activity.

Tetracyclic compounds of the formula (B) containing a pyrrolo[2.1-a]isoquinoline moiety are described in Arch. Pharm. 321, 481 (1988):

The compounds B are described as having anti-tumor activity due to their ability to intercalate into DNA. It is not mentioned that these compounds have any PDE 10a inhibitory activity.

The synthesis of pyrrolo[2.1-a]isoquinoline derivatives of formula (C) is described in H. Meyer, Liebigs Ann. Chem. 9, 1534-1544 (1981):

R = H, OCH₃ R" = CH₃, C₆H₅ R"' = C₆H₅

These compounds are not described as having any biological activity, and it is not mentioned that these compounds have any PDE 10a inhibitor activity.

Compounds ofthe formula (D) are described in GB 1 153 670 A:

R = H, C0₂H, C0₂R"" R" = H, C0₂H, C0₂R"" R'" = C₆H_5l CH₃, C0₂R""

These compounds are described as having hypotensive, sympathicolytic and psycho- tropic properties, but it is not mentioned that these compounds have any PDE 10a inhibitory activity. The synthesis of compounds ofthe formula (E) is described in US Patent 4,694,085:

R = H, CH₃, OCH₃ R ⁼ H, CH₃

R- = C₆H₅, CH₃, C0₂R'"" R"" = H, CH₃

It is not mentioned that these compounds have any PDE 10a inhibitory activity.

Derivatives ofthe formula (F) are described in WO 98/55118:

R = H, CI, OCH₃ R" = CH₃

R'" = OR""', CH₃, NH₂ R"" = H, CH₃, OR'""

These compounds are described as useful for the treatment of diseases such as psoriasis. However, the compounds disclosed in WO 98/55118 are described as having virtually no cytotoxic activity. It is not mentioned that these compounds have any PDE 10a inhibitor activity. BRIEF SUMMARY OF THE INVENTION

Surprisingly, it has been found that the pyrrolo[2.1-a]isoquinoline derivatives of the present invention inhibit PDE 10a and exhibit an antiproliferative activity.

The present invention relates to a compound ofthe formula

wherein

x and y independently from each other denote zero or 1 and x+y is 1 or 2;

R¹ and R² independently from each other denote hydrogen, C_1-4-alkyl or CF₃₎ or R¹ and R² together form a C_1- -alkylene bridge;

R³ denotes hydrogen, formyl, (C_1- -alkyl)-carbonyl, (C_1-4-alkoxy)-carbonyl, NO₂, NR⁶R⁷, C_1-4-alkyl-NR⁶R⁷, C_1-4-alkyl-OR⁸, C^-alkyl-COOR⁸, C_6-10-aryl-C_1-4-alkyl wherein the aryl moiety is optionally substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl, and C_1-4-alkoxy;

wherein

R⁶ and R⁷ independently from each other denote hydrogen, C_1- -alkyl, C_3-8- cycloalkyl, or C_6-10-aryl-C_1-4-alkyl wherein the aryl moiety is optionally substituted with 1 to 3 radicals selected from the group consisting of OH, Cj.

₄-alkyl and C_1- -alkoxy; or

R and R together with the nitrogen atom to which they are attached form a 5- to 7-membered heterocyclyl which may contain up to 2 further hetero atoms selected from the group consisting of N, O and S, which heterocyclyl can further be substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl, C_1- -alkoxy, C_6-10-aryl and aromatic 4- to 9- membered heterocyclyl with 1 to 4 hetero atoms selected from the group consisting of N, O and S;

R denotes hydrogen or C_1-4-alkyl;

R⁴ denotes C_1-4-alkyl;

R⁵ is

i) C_6-14-aryl optionally containing 1 to 3 further substituents selected from the group consisting of

halogen;

C_1-6-alkyl which can be further substituted with one or more radicals selected from the group consisting of Cι_-6-alkoxy, OH, and NH₂;

C_1-6-alkoxy which can be further substituted with one or more radicals selected from the group consisting of C_1-6-alkoxy, OH, and NH₂;

C_6-10-aryloxy-C_1-6-alkoxy; OH;

NO₂; CN; CF₃; OCF₃;

NR⁹R¹⁰; CONR⁹R¹⁰; COOR¹¹; SR¹¹; SOR¹¹;

SO₂R^u; OSO₂R^π;

-O-(CH₂)_1-4-O- wherein the oxygen atoms are bound to the aryl moiety in ortho-position to each other;

phenyloxy or benzyloxy wherein the phenyl moieties can contain one further substituent selected from the group consisting of C_1- -alkyl, C_1-6-alkoxy, halogen and NO₂;

phenyl optionally substituted with CN;

aromatic 4- to 9-membered heterocyclyl with 1 to 4 hetero atoms selected from the group consisting of N, O and S;

and

saturated 5- to 7-membered nitrogen-containing heterocyclyl which is linked to the C_6-10-aryl moiety via the nitrogen atom and may contain up to 2 further heteroatoms selected from the group consisting of N, O and S and which saturated heterocyclyl can be further substituted with one or more radicals selected from the group consisting of C_1-6-alkoxy, OH and NH₂;

wherein

R and R¹⁰ independently from each other denote hydrogen, C_1-6-alkyl, (C_1-6-alkyl)-carbonyl, (C_1-6-alkoxy)-carbonyl, C_1-6- alkylsulfonyl, (C_1-6-alkylamino)-carbonylamino, (C_6-1o-aryl- amino)-carbonylamino,

or

R⁹ and R¹⁰ together with the nitrogen atom to wliich they are attached, form a 5- to 7-membered saturated, partially unsaturated or aromatic heterocyclyl which can contain up to 3 further hetero atoms selected from the group consisting of N, O and S, and which heterocyclyl can contain 1 to 3 substituents selected from the group consisting of C_1-6-alkyl, C-_1-6-alkoxy, C_6-10-aryl and aromatic 4- to 9-membered heterocyclyl with 1 to 4 hetero atoms selected from the group consisting of N, O and S;

R¹¹ is hydrogen, C_1-6-alkyl or C_6-1o-aryl;

or

ii) C_1-12-alkyl wliich can contain 1 to 3 substituents selected from the group consisting of C_1-6-alkyl, C-_1-6-alkoxy, C_6-1o-aryl and aromatic 4- to 9-membered heterocyclyl with 1 to 4 hetero atoms selected from the group consisting of N, O and S;

or

iii) C_3-8-cycloalkyl which can contain 1 to 3 substituents selected from the group consisting of C_1-6-alkyl, C-_1-6-alkoxy, COOR¹¹ wherein R¹¹ is as defined above, C_6-10-aryl and aromatic 4- to 9-membered heterocyclyl with 1 to 4 hetero atoms selected from the group consisting of N, O and S;

or

iv) aromatic C_2-9-heterocyclyl with 1 to 4 hetero atoms selected from the group consisting of N, O and S which aromatic heterocyclyl can contain 1 to 3 further substituents selected from the group consisting of OH, C_1-6-alkyl, C-_1-6-alkoxy, C_6-1o-aryl which can contain 1 to 3 halogen radicals, COR¹¹ or COOR¹¹ wherein R¹¹ is as defined above, halogen, CN, and saturated 5- to 7-membered nitrogen-containing heterocyclyl wliich is bound to the C _-10-aryl moiety via the nitrogen atom and can contain up to 2 further hetero atoms selected from the group consisting of N, O and S and which saturated heterocyclyl can be further substituted with one or more radicals selected from the group consisting of C_1-6-alkoxy, OH and NH₂;

with the proviso that ethyl 8,9-dimethoxy-2-phenyl-5,6-dihydropyrrolo[2.1-a]iso- quinoline-1-carboxylate is excluded;

and an isomer, a pharmaceutically acceptable salt, a hydrate or a hydrate of a pharmaceutically acceptable salt thereof. An alternative embodiment of the present invention relates to a compound of the formula (I), wherein

x and y independently from each other denote zero or 1 and x+y is 1 or 2;

R¹ and R² independently from each other denote C_1-4-alkyl or CF ;

R³ denotes hydrogen, formyl, (C_1-4-alkyl)-carbonyl, (C_1- -alkoxy)-carbonyl, NO₂, NR₆R₇, C_1-4-alkyl-NR⁶R⁷, C_1-4-alkyl-OR⁸, C_1-4-alkyl-COOR⁸, or C_6-10-aryl-C_1-4-alkyl wherein the aryl moiety can be substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl and C_1-4-alkoxy;

wherein

R⁶ and R⁷ independently from each other denote hydrogen, C_1- -alkyl, C_3-8- cycloalkyl, or C_6-10-aryl-C_1-4-alkyl wherein the aryl moiety can be substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl, and C_1-4-alkoxy;

or

R⁶ and R⁷ together with the nitrogen atom to wliich they are attached form a 5- to 7-membered heterocyclyl which may contain up to 2 further hetero atoms selected from the group consisting of N, O, and S and which heterocyclyl can be further substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl, C_1- -alkoxy, C_6-10-aryl and aromatic 4- to 9-membered heterocyclyl with 1 to 3 hetero atoms selected from the group consisting of N, O, and S; R⁸ denotes hydrogen or C_1- -alkyl;

R⁴ denotes C_1-4-alkyl;

R⁵ is

i) phenyl optionally having 1 to 3 further substituents selected from the group consisting of F, CI, Br; C_1-6-alkyl; C_1-6-alkoxy; OH; NR⁹R¹⁰ and COOR¹¹;

or

ii) naphthyl optionally containing one further OH group;

or

iii) indolyl optionally having 1 to 3 further substituents selected from the group consisting of F, CI, Br; C_1-6-alkyl; C_1-6-alkoxy; OH; NR⁹R¹⁰ and COOR¹¹;

wherein R⁹ to R¹¹ independently from each other denote C_1-6-alkyl;

and an isomer, a pharmaceutically acceptable salt, a hydrate or a hydrate of a pharmaceutically acceptable salt thereof.

A further alternative embodiment of the present invention relates to a compound of the formula (I), wherein x and y independently from each other denote zero or 1 and x+y is 1 or 2;

R¹ and R² independently from each other denote CH₃ or C₂H₅;

R³ denotes hydrogen, formyl, (C_1-4-alkyl)-carbonyl, (C_1-4-alkoxy)-carbonyl, NO₂, NR⁶R⁷, C_1-4-alkyl-NR⁶R⁷, C_1-4-alkyl-OR⁸, C_1-4-alkyl-COOR⁸, or C_6-10-aryl-C_1-4-alkyl wherein the aryl moiety can be substituted with 1 to 3 radicals selected from the group consisting of OH, C_1- -alkyl and C_1-4-alkoxy;

wherein

independently from each other denote hydrogen, C_1-4-alkyl, C_3-8- cycloalkyl, or C_6-10-aryl-C_1- -alkyl wherein the aryl moiety can be substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl and

C_1-4-alkoxy;

or

together with the nitrogen atom to which they are attached form a

5- to 7-membered heterocyclyl which may contain up to 2 further hetero atoms selected from the group consisting of N, O, and S and which heterocyclyl can be further substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl, C_1-4-alkoxy, C_6-10-aryl and aromatic 4- to 9-membered heterocyclyl with 1 to 3 hetero atoms selected from the group consisting of N, O, and S;

R denotes hydrogen or C_1- -alkyl;

R⁴ denotes CH₃ or C₂H₅; R⁵ is

i) phenyl optionally having 1 to 3 further substituents selected from the group consisting of CI; C_1-4-alkyl; C_1-4-alkoxy; OH; NR⁹R¹⁰, and

COOR¹¹;

or

ii) naphthyl optionally containing one further OH group;

or

iii) indolyl optionally having 1 to 3 further substituents selected from the group consisting of F, CI, Br; C_1-6-alkyl; C_1-6-alkoxy; OH; NR⁹R¹⁰, and COOR¹¹;

wherein R⁹ to R¹¹ independently from each other denote C_1-4-alkyl;

A further alternative embodiment of the present invention relates to a compound of the formula (I), wherein

x and y independently from each other denote zero or 1 and x+y is 1 or 2; R and R independently from each other denote CH₃ or C H₅;

R denotes hydrogen, formyl, (C_1-4-alkyl)-carbonyl, (C_1-4-alkoxy)-carbonyl, NO₂, NH₂, C_1-4-alkyl-NR⁶R⁷, C_1-4-alkyl-OR⁸, C_1-4-alkyl-COOR⁸, or phenyl-C_1-4-alkyl wherein the phenyl moiety can be substituted with 1 to 3 C_1-4-alkyl or C_1- -alkoxy moieties;

wherein

R⁶ and R⁷ independently from each other denote hydrogen, C_1-4-alkyl, C_3-6- cycloalkyl, or phenyl-C_1-4-alkyl wherein the phenyl moiety can be substituted with 1 to 3 C_1-4-alkyl or C_1-4-alkoxy radicals;

or

R and R together with the nitrogen atom to which they are attached, form a saturated 5- to 7-membered heterocyclyl which may contain up to 2 further hetero atoms selected from the group consisting of N, O, and S and which saturated heterocyclyl can be further substituted with 1 to 3 radicals selected from the group consisting of C_1-4-alkyl, C_1-4-alkoxy, phenyl and pyridyl;

R⁸ denotes hydrogen or C_1-4-alkyl;

R⁴ denotes CH₃ or C₂H₅;

R⁵ is

i) phenyl optionally having 1 to 3 further substituents selected from the group consisting of CI; C_1-4-alkyl; C_1- -alkoxy; OH; NR⁹R¹⁰; and COOR¹¹; wherein R >9 ₊ to. R .11 independently from each other denote C_1-4-alkyl;

or

ii) naphthyl optionally containing one further OH group;

or

iii) indolyl;

A bond with a dotted line thereunder, _ ττ denotes a bond which alternatively is a single bond or a double bond.

Compounds (I) wherein the radicals (R¹O)_x and (R²O)_y are attached to the phenyl ring in the following positions, are particularly preferred:

Furthermore, according to the present invention the respective 5,6-dihydro pyrrolo derivatives of formula (I) are preferred.

Furthermore, the compounds of Examples 2, 5, and 29 are particularly preferred.

DETAILED DESCRIPTION OF THE INVENTION

Pharmaceutically acceptable salts according to the invention are non-toxic salts which in general are accessible by reaction ofthe compounds (I) with an inorganic or organic base or acid conventionally used for this purpose. Non-limiting examples of pharmaceutically acceptable salts of compounds (I) include the alkali metal salts, e.g. lithium, potassium and sodium salts, the alkaline earth metal salts such as the magnesium and calcium salts, the quaternary ammonium salts such as, for example, the triethyl ammonium salt, acetates, benzene sulphonates, benzoates, dicarbonates, disulphates, ditartrates, borates, bromides, carbonates, chlorides, citrates, dihydro- chlorides, fumarates, gluconates, glutamates, hexyl resorcinates, hydrobromides, hydrochlorides, hydroxynaphthoates, iodides, isothionates, lactates, laurates, malates, maleates, mandelates, mesylates, methylbromides, methylnitrates, methylsulphates, nitrates, oleates, oxalates, palmitates, pantothenates, phosphates, diphosphates, poly- galacturonates, salicylates, stearates, sulphates, succinates, tartrates, tosylates, valerates, and other salts used for medicinal purposes.

The present invention includes both the individual enantiomers or diastereomers and the corresponding racemates, diastereomer mixtures and salts of the compounds ac- cording to the invention. In addition, all possible tautomeric forms ofthe compounds described above are included according to the present invention. The diastereomer mixtures can be separated into the individual isomers by chromatographic processes. The racemates can be resolved into the respective enantiomers either by chromatographic processes on chiral phases or by resolution. In the context of the present invention, the substituents, if not stated otherwise, in general have the following meaning:

Alkyl per se as well as the prefixes "alkyl" and "alk" in the terms "alkylcarbonyl", "alkylsulphonyl", "alkylaminocarbonylamino", "alkoxy", and "alkoxycarbonyl" represent a linear or branched alkyl radical preferably having 1 to 12, more preferably 1 to 6 carbon atoms. Non-limiting examples include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, pentyl, isopentyl, hexyl, and isohexyl.

Non-limiting examples of "alkylcarbonyl" include acetyl, ethylcarbonyl, propyl- carbonyl, isopropylcarbonyl, butylcarbonyl, and isobutylcarbonyl. The terms "alkylcarbonyl" and "acyl" are used synonymously.

Non-limiting examples of "alkylsulphonyl" include methylsulphonyl, ethylsulphonyl, propylsulphonyl, isopropylsulphonyl, butylsulphonyl, and isobutylsulphonyl.

Non-limiting examples of "alkylaminocarbonylamino" include methylaminocarbo- nylamino, ethylaminocarbonylamino, propylaminocarbonylamino, isopropylamino- carbonylamino, butylaminocarbonylamino, and isobutylaminocarbonylamino.

Non-limiting examples of "alkoxy" include methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, pentoxy, isopentoxy, hexoxy, and isohexoxy.

Non-limiting examples of "alkoxycarbonyl" include methoxycarbonyl, ethoxy- carbonyl, propoxycarbonyl, isopropoxycarbonyl, butoxycarbonyl, and isobutoxy- carbonyl.

Alkylene represents a linear or branched (bivalent) alkylene radical preferably having 1 to 4 carbon atoms. Non-limiting examples include methylene, ethylene, propylene, α-methylethylene, β-methylethylene, α-ethylethylene, β-ethylethylene, butylene, α- methylpropylene, β-methylpropylene, and γ-methylpropylene.

Cycloalkyl represents a saturated cycloalkyl radical preferably having 3 to 8 carbon atoms. Non-limiting examples include cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl; cyclopropyl, cyclopentyl and cyclohexyl are preferred.

Aryl per se and in the terms "aryloxy", "aryl-alkyl", and "arylaminocarbonylamino" represents an aromatic radical preferably having from 6 to 14, more preferably 6 to 10 carbon atoms. Non-limiting examples of aryl radicals include phenyl, benzyl, naphthyl, and phenanthrenyl. Non-limiting examples of aryloxy radicals include phenyloxy and benzyloxy. Non-limiting examples of aryl-alkyl radicals include benzyl. Non-limiting examples of arylaminocarbonylamino radicals include phenyl- aminocarbonylamino, benzylaminocarbonylamino, naphthylaminocarbonylamino, and phenanthrenylaminocarbonylamino.

Heterocyclyl in the context ofthe invention represents a saturated, partially saturated or aromatic 4- to 9-membered, for example 5- to 6-membered ring which can contain from 1 to 3 hetero atoms selected from the group consisting of S, N and O and wliich ring can be bound via a carbon atom or a nitrogen atom, if such an atom is present.

Non-limiting heterocyclyl examples include: oxadiazolyl, thiadiazolyl, pyrazolyl, pyridyl, pyrimidinyl, pyridazinyl, pyrazinyl, quinolinyl, isoquinolinyl, indolyl, thienyl, furyl, pyrrolyl, N-methylpyrrolyl, indazolyl, benzimidazolyl, pyrrolidinyl, piperazinyl, tetrahydropyranyl, tetrahydrofuranyl, 1,2,3 triazolyl, thiazolyl, oxazolyl, imidazolyl, morpholinyl, thiomorpholinyl or piperidyl. Preferred examples include thiazolyl, furyl, oxazolyl, pyrazolyl, triazolyl, pyridyl, pyrimidinyl, pyridazinyl and tetrahydropyranyl. The terms "heteroaryl" and "hetaryl" denotes an aromatic heterocyclic radical. Halogen in the context of the invention represents fluorine, chlorine, bromine, and iodine.

The present invention also relates to a process for manufacturing the compounds according to the invention comprising the reaction of a compound ofthe formula

wherein x, y, R¹, R² and R⁴ are as defined above,

[A] with the compounds ofthe formulae

R⁵-CHO and R³-CH₂-NO₂

(II) (in) wherein R³and R⁵ are as defined above, or

[B] with a compound ofthe formula

wherein R³ and R⁵ are as defined above, and optionally

[C] the conversion of the compound obtained through either process [A] or [B] into an isomer, a pharmaceutically acceptable salt, a hydrate or a hydrate of a pharmaceutically acceptable salt thereof. The compounds (II) are commercially available or can be synthesized according to methods commonly known to those skilled in the art (LT. Harrison and S. Harrison, Compendium of Organic Synthetic Methods, pp. 132-176, Wiley-Interscience; T.D. Harris and G.P. Roth, J. Org. Chem. 44, 146 (1979); E. Mϋller (ed.), "Methoden der Organischen Chemie" (Houben-Weyl), Vol. VII/1 Sauerstoff-Verbindungen IL, Georg Thieme Verlag, Stuttgart 1954).

The compounds (III) are commercially available.

The compounds (FV) can be synthesized by reacting compounds ofthe formula

wherein x, y, R¹ and R² are as defined above,

with compounds ofthe formula

wherein R⁴ is as defined above and

L is a leaving group, for example a halogen radical such as CI, or a radical of the formula

to give compounds ofthe formula

wherein x, y, R¹, R² and R⁴ are as defined above,

and reacting compound (VIII) with a dehydrating agent.

The compounds (VI) are commercially available or can be synthesized according to methods commonly known to those skilled in the art (H. Mayer et al., Heterocycles 31, 1035 (1990); E. Mύller (ed.), "Methoden der Organischen Chemie" (Houben- Weyl), 4^th ed., Vol. 11/1 Stickstoff-Verbindungen II, Georg Thieme Verlag, Stuttgart 1957; Shepard et al. in J. Org. Chem. 17, 568 (1952) and in J. Am. Chem. Soc. 72, 4364 (1950)).

The compounds (VII) are commercially available or can be synthesized according to methods commonly known those skilled in the art [e.g. via acylation of acetic acid with an alkyl chloroformate or dialkyl carbonate (March, Advanced Organic Chemistry, 3^rd ed., p. 440-441, Wiley 1985) and converting the resulting monoester of malonic acid into e.g. the corresponding acid chloride or anhydride by methods commonly known to those skilled in the art (see e.g. March, Advanced Organic Chemistry, 3^rd ed., p. 355, 388, Wiley 1985)].

The reaction between the compounds (VT) and (Nil) is preferably carried out in a solvent. Suitable solvents comprise the customary organic solvents which are inert under the reaction conditions. Νon-limiting examples include ethers such as diethyl ether, dioxane, tetrahydrofuran, 1,2-dimethoxy ethane; hydrocarbons such as benzene, toluene, xylene, hexane, cyclohexane, mineral oil fractions; halogenated hydrocarbons such as dichloromethane, trichloromethane, carbon tetrachloride, dichloroethane, tri- chloroethylene, chlorobenzene; ketones such as acetone; esters such as ethyl acetate; nitriles such as acetonitrile; heteroaromatics such as pyridine; optionally N-alkylated carboxylic acid amides such as dimethyl formamide and dimethyl acetamide; alkyl sulphoxides such as dimethyl sulphoxide; optionally alkylated phosphoric acid amides such as hexamethyl phosphoric acid tris-amide; and mixtures of the above-mentioned solvents. Dichloromethane is particularly preferred.

The compound (NH) is generally employed in an amount of from 1 to 4 mol per mol of compound (NL); an equimolar amount or slight excess of compound (VH) is preferred.

The reaction between the compounds (VI) and (VH) is preferably carried out in the presence of a base. Νon-limiting examples include alkali metal hydrides and alkali metal alkoxides such as, for example, sodium hydride and potassium tert-butoxide; C₁-C₄-alkyl amines such as, for example, triethyl amine; cyclic amines such as, for example, piperidine, pyridine, dimethylamino pyridine and - preferably - 1,8-diaza- bicyclo[4.3.0]undec-7-ene (DBU). The base is generally employed in an amount of from 1 to 4 mol per mol of compound (VI); an equimolar amount or slight excess ofthe base is preferred.

The reaction of the compounds (VI) and (NIL) can generally be carried out within a relatively wide temperature range. In general, the reaction is carried out within a range of from -20 to 200°C, preferably from 0 to 70°C, and more preferably at room temperature.

For the cyclization of the compounds (VHJ) to yield compounds (IN), dehydrating agents such as, for example, P₂O_5; POCl₃, or methane sulfonic anhydride are generally employed in an amount of from 1 to 10 mol, preferably from 1 to 2 mol of methane sulfonic anhydride or 4 to 8 mols of P₂O₅ and POCl₃, respectively, per mol of compound (NIH) in each case.

The cyclization reaction of the compounds (VLJI) to yield the compounds (LV) is also preferably carried out in a solvent. Νon-limiting examples comprise the customary organic solvents which are inert under the reaction conditions. They preferably include ethers such as diethyl ether, dioxane, tetrahydrofuran, 1,2-dimethoxy ethane; hydrocarbons such as benzene, toluene, xylene, hexane, cyclohexane, mineral oil fractions; halogenated hydrocarbons such as dichloromethane, trichloromethane, carbon tetra- chloride, dichloroethane, trichloroethylene, chlorobenzene; esters such as ethyl acetate; ketones such as acetone; nitriles such as acetonitrile; heteroaromatics such as pyridine; optionally N-alkylated carboxylic acid amides such as dimethyl formamide and dimethyl acetamide; alkyl sulphoxides such as dimethyl sulphoxide; optionally alkylated phosphoric acid amides such as hexamethyl phosphoric acid tris-amide; and mixtures thereof. Toluene is preferred if the reaction is carried out with P₂O₅ or methane sulfonic anhydride; and acetonitrile is preferred if the reaction is carried out with POCl₃ (Benovsky, Stille, Tetrahedron Lett. 38, 8475-8478 (1997)).

The temperature for the cyclization reaction of compounds (VLIL) is preferably within a range of from 60 to 200°C and more preferably within a range of from 80 to 120°C.

The above process steps are generally carried out under atmospheric pressure. However, it is also possible to carry them out under superatmospheric pressure or under reduced pressure (for example, in a range of from 0.5 to 5 bar). The reaction time can generally be varied within a relatively wide range. In general, the reaction is finished after a period of from 2 to 24 hours, preferably from 6 to 12 hours.

The reaction ofthe compounds (IN) with either compounds (II) and (HI) or with compound (N) can be carried out as a one-pot synthesis, preferably in a solvent. Suitable solvents comprise the customary organic solvents wliich are inert under the reaction conditions. Non-limiting examples include ethers such as diethyl ether, dioxane, tetra- hydrofuran, 1,2-dirnethoxy ethane; hydrocarbons such as benzene, toluene, xylene, hexane, cyclohexane, mineral oil fractions; halogenated hydrocarbons such as dichloromethane, trichloromethane, carbon tetrachloride, dichloroethane, trichloro- ethylene, chlorobenzene; alcohols such as methanol, ethanol, n-propanol, isopropanol; esters such as ethyl acetate; ketones such as acetone; nitriles such as acetonitrile; heteroaromatics such as pyridine; optionally N-alkylated carboxylic acid amides such as dimethyl formamide and dimethyl acetamide; alkyl sulphoxides such as dimethyl sulphoxide; optionally alkylated phosphoric acid amides such as hexamethyl phosphoric acid tris-amide; and mixtures thereof. Ethanol/isopropanol (approximately

1 : 1 vol/vol) mixtures are preferred.

The compounds (Hi) are generally employed in an amount of from 1 to 3 mol per mol of compound (H); an equimolar amount or slight excess of compound (HI) is particu- larly preferred. The compounds (IN) are generally employed in an amount of from 0.1 to 1 mol, preferably from 0.3 to 1 mol, per mol of compounds (H).

The reactions ofthe compounds (IN) with either compounds (H) and (HI) or with compound (V) are preferably carried out in the presence of a base. Νon-limiting examples include alkali metal hydrides and alkali metal alkoxides such as, for example, sodium hydride and potassium tert-butoxide; C_1-4-alkyl amines such as, for example, triethyl amine; cyclic amines such as, for example, pyridine, dimemylamino pyridine, 1,8-di- azabicyclo[4.3.0]undec-7-ene (DBU) and - preferably - piperidine. The base is generally employed in an amount of from 0.1 to 1 mol, preferably from 0.3 to 1 mol, per mol of compound (H) or compound (V), respectively.

The reactions of the compounds (LV) with either compounds (H) and (HI) or with compound (V) are generally carried out within a relatively wide temperature range. In general, they are carried out in a range of from -20 to 200°C, preferably from 0 to 100°C, and more preferably from 50 to 90°C. The steps of this reaction are generally carried out under atmospheric pressure. However, it is also possible to carry them out under superatmospheric pressure or under reduced pressure (for example, in a range of from 0.5 to 5 bar). The reaction time can generally be varied within a relatively wide range. In general, the reaction is finished after a period of from 2 to 24 hours, preferably from 6 to 12 hours.

The compounds (V) are commercially available or can be synthesized in analogy to the reaction of compounds (H) and (IH) described above (in the absence of compound

(IN)).

The following scheme illustrates a representative process for the synthesis of the compounds according to the present invention:

(VIII)

wherein x, y, R¹ to R⁵ and L are as defined above.

If the compounds (I) are not directly obtained by reacting the compounds (H), (HI) and (LV) or (IN) and (N), the compounds thus obtained have to be converted into the compounds (I) by further reactions known to the man skilled in the art.

For example, compounds (I) wherein R³ is C_1-4-alkyl-ΝR⁶R⁷, C_1-4-alkyl-OR⁸, C_1-4- alkyl-COOR⁸ or C_6-1o-aryl-C_1-4-alkyl can be synthesized from compounds wherein R³ is C_1-4-alkyl (which themselves can be obtained according to one of the above processes A or B) by reaction with a halogenating agent such as sulfuryl chloride

(SO₂Cl₂), thionyl chloride (SOCl₂) or N-chlorosuccinimide (NCS), preferably in an organic solvent commonly used for such reactions, for example in a halogenated alkane such as dichloromethane, under conditions known to the skilled man, and a consecutive nucleophilic substitution reaction with an appropriate nucleophile such as the respective amine HNR⁶R⁷, the respective alcoholate R⁸O ^" or OH ^", under conditions commonly used for such reactions and known to the skilled man. The halogenated intermediate obtained after the first ofthe above reaction steps can either be isolated and then reacted with the desired nucleophile or directly be converted into the desired product by reaction with a respective nucleophile.

Compounds (I) wherein R³ is hydrogen can be synthesized by process A or B using compound (III) or (V) respectively wherein R³ is hydrogen.

Compounds (I) wherein R³ is formyl can be synthesized from the compounds wherein R³ is methyl (which themselves can be obtained according to one of the above processes A or B) by reaction with manganese dioxide in an organic solvent commonly used for such reactions such as, for example, an ether such as dioxane under conditions known to the skilled man. The formyl compounds thus obtained can also be converted into compounds (I) wherein R³ is CH₂NR⁶R⁷ by a reductive animation reaction commonly known to the skilled man. Compounds (I) wherein R³ is C_1-4-alkylcarbonyl can be synthesized preferably by reaction of compounds (IN) with compounds (N), wherein R³ is C_1-4-alkylcarbonyl (these derivatives can be prepared from nitromethyl-alkylketones (compare D.C. Baker et al., Synthesis 1978; 478-479) and activated aldehydes, e.g. benzylidene- butyl-amines (see Dornow et al., Liebigs Ann. Chem. 602: 14, 19 (1957)).

Compounds (I) wherein R³ is ΝO₂ can be synthesized from the compounds wherein R³ is methyl (which themselves can be obtained according to one of the above processes A or B) by reaction with HNO₃ in acetic acid under conditions commonly used for such reactions and known to the skilled man. These nitro compounds can further be converted into compounds wherein R³ is NR⁶R⁷ by a hydrogenation of the nitro group to the respective amino group under conditions commonly used for such reactions and known to the skilled man, and optionally alkylating the amino group under conditions commonly used for such reactions and known to the skilled man.

The compounds ofthe present invention are inhibitors of phosphodiesterase 10a (PDE 10a). As outlined above, the inhibition of PDE 10a is a promising approach for the treatment of cancer. The biological tests described below show that the compounds (I) exhibit a pronounced anti-proliferation activity against tumor cells; they are therefore useful for the treatment of cancer. Furthermore, our investigations showed that they are also useful for treatment of conditions of pain and/or for the lowering of the temperature ofthe body in fever conditions.

The compounds according to the invention can be used as active ingredients for the production of medicaments against carcinomatous disorders. For this, they can be converted into the customary formulations such as tablets, coated tablets, aerosols, pills, granules, syrups, emulsions, suspensions and solutions using inert, non-toxic, pharmaceutically suitable excipients or solvents. Preferably, the compounds accord- ing to the invention are used in an amount such that their concentration is approxi- mately 0.5 to approximately 90% by weight, based on the ready-to-use formulations, the concentration being dependent, inter alia, on the indication ofthe medicament.

The formulations can be produced, for example, by extending the active compounds with solvents and/or excipients having the above properties, where, if appropriate, additionally emulsifiers or dispersants and, in the case of water as the solvent, an organic solvent can additionally be added.

Administration can be carried out in a customary manner, preferably orally, trans- dermally or parenterally, for example perlingually, buccally, intravenously, nasally, rectally or inhalationally.

For human use, in the case of oral administration, it is recommended to administer doses of from 0.001 to 50 mg/kg, preferably from 0.01 to 20 mg/kg. In the case of parenteral administration such as, for example, intravenously or via mucous membranes nasally, buccally or inhalationally, it is recommended to use doses of from 0.001 to 0.5 mg/kg.

If appropriate, it may be necessary to depart from the amounts mentioned above, namely depending on the body weight or the type of administration route, on the individual response towards the medicament, the manner of its formulation and the time or interval at which administration takes place. Thus, in some cases it may be sufficient to manage with less than the above mentioned minimum amount, while in other cases the upper limit mentioned must be exceeded. In the case ofthe adniinistration of relatively large amounts, it may be recommended to divide these into several individual doses over the course ofthe day.

The compounds according to the invention are also suitable for use in veterinary medicine. For use in veterinary medicine, the compounds or their non-toxic salts can be ad- ministered in a suitable formulation in accordance with general veterinary practice. Depending on the kind of animal to be treated, the veterinary surgeon can determine the nature of use and the dosage.

The present invention provides compounds and salts thereof for the use in a medicinal application, in particular for combating cancer.

The invention further provides a method of manufacturing a pharmaceutical composition by combining at least one of the compounds ofthe invention with at least one pharmacologically acceptable formulating agent.

The invention further provides a pharmaceutical composition comprising as an active ingredient an effective amount of at least one ofthe compounds ofthe invention and at least one pharmacologically acceptable formulating agent.

The invention further provides a pharmaceutical composition comprising as an active ingredient an effective amount of at least one ofthe compounds ofthe invention and at least one pharmaceutically active ingredient wliich is different from the compounds ofthe invention.

The invention further provides a medicament in dosage unit form comprising an effective amount of a compound according to the invention together with an inert pharmaceutical carrier.

The invention further provides a method of combating cancer in humans and animals comprising the administration of an effective amount of at least one compound according to the invention either alone or in admixture with a diluent or in the form of a medicament.

The percentages in the description above, in the following tests and in the Examples are - if not stated otherwise - percentages by weight; parts are parts by weight. Solvent ratios, dilution ratios and concentrations in solutions of liquids in liquids are ratios and concentrations by volume.

Biological tests

In vitro Enzyme Inhibition Assay

Full-length recombinant PDE 10a was expressed in Sf9 insect cells (Invitrogen, Carlsbad, CA, U.S.A.) using the Bac-to-Bac™ Baculovirus Expression System (Life Technologies, Gaithersburg, MD, U.S.A.). 48 hours post infection, cells were harvested and resuspended in 20 mL (per IL culture) of Lysis Buffer (50 mM Tris- HCl, pH 7.4, 50 mM NaCl, 1 mM MgCl₂, 1.5 mM EDTA, 10% glycerol plus 20 μL

Protease Inhibitor Cocktail Set III [CalBiochem, La Jolla, CA, U.S.A.]). Cells were sonicated at 4°C for 1 minute and centrifuged at 10,000 RPM for 30 minutes at 4°C. Supernatant was removed and stored at -20°C for activity assays.

The test compounds were serially diluted in DMSO using two-fold dilutions to stock concentrations ranging typically from 200 μM to 1.6 μM (final concentrations in the assay range from 4 μM to 0.032 μM). 96-well assay isoplates (Wallac Inc., Atlanta, GA, U.S.A.) were loaded with 2 μL ofthe serially diluted individual test compounds followed by 50 μL of a dilution of crude recombinant PDE lOa-containing Sf9 cell lysate. The dilution ofthe lysate was selected such that less than 70% ofthe substrate is converted during the later incubation (typical dilution: 1:10 000; dilution buffer: 50 mM Tris/HCl pH 7.5, 8.3 mM MgCl₂, 1.7 mM EDTA, 0.2% BSA). The substrate, [5',8-³H] adenosine 3',5'-cyclic phosphate (1 μCi/μL; Amersham Pharmacia Biotech., Piscataway, NJ, U.S.A.), was diluted 1:2000 in assay buffer (assay buffer: 50 mM Tris/HCl pH 7.5, 8.3 mM MgCl₂, 1.7 mM EDTA) to give a final working concentration of 0.0005 μCi/μL. The enzymatic assay was initiated by addition of 50 μL (0.025 μCi) of diluted substrate. Reactions were incubated at room temperature for 60 minutes and terminated by addition of 25 μL of 18 mg/mL Yttrium Scintillation Proximity Beads (Amersham Pharmacia Biotech., Piscataway, NJ, U.S.A.). Plates were sealed and incubated at room temperature for 60 minutes. Plates were read for 30 seconds/well using a Microbeta counter (Wallac Inc., Atlanta, GA, U.S.A.). The IC₅₀ values were determined by plotting compound concentration versus percent inhibition. Representative results are shown in Table 1 :

Table 1

In vitro Proliferation Inhibition Assay:

MDA-MB-231 human breast carcinoma cells (ATCC # HTB26) were cultured in standard growth medium (DMEM), supplemented with 10% heat-inactivated FBS, lO mM HEPES, 2 mM glutamine, 100 U/mL penicillin, and 100 μg/mL streptomycin) at 37°C in 5% CO₂ (vol/vol) in a humidified incubator. Cells were plated at a density of 3000 cells per well in 100 μL growth medium in a 96 well culture dish. 24 hours after plating, LDH activity was determined using the Cytotox 96 Non-radioactive Cytotoxicity Kit (Promega, Madison, Wl, U.S.A.) to yield T_0h LDH values. Briefly, cells were lysed with the addition of 200 μL of Lysis Buffer (included in the Promega Kit) and lysates were further diluted in Lysis Buffer so that LDH values fell within the standard curve. 50 μL of diluted cell lysate were transferred to a fresh 96 well culture plate. The assay was initiated with the addition of 50 μL of substrate per well. Color development was allowed to proceed for 10-15 minutes. The assay was terminated with the addition of 50 μL of Stop Solution (included in Promega Kit). Optical densities were determined spectrophotometrically at 490 nm in a 96 well plate reader (SpectraMax 250, Molecular Devices, Sunnyvale, CA, U.S.A.). Test compounds were dissolved in 100% DMSO to prepare 10 mM stocks. Stocks were further diluted 1 :250 in growth medium to yield working stocks of 40 μM test compound in 0.4% DMSO. Test compounds were serially diluted in growth medium containing 0.4% DMSO to maintain constant DMSO concentrations for all wells. 50 μL of fresh growth medium and 50 μL of diluted test compound were added to each culture well to give a final volume of 200 μL. The cells with and without individual test compounds were incubated for 72 hours at which time LDH activity was measured to yield T _1l values. Optionally, the ICso values can be determined with a least squares analysis program using compound concentration versus percent inhibition.

% Inhibition = [l-(T_72ht_est-To )/(T₇₂h _ctri-T_oh)] x 100

wherein

L 72htest LDH activity at 72 hours in the presence of test compound,

T_{72h ctr}i = LDH activity at 72 hours in the absence of test compound and

Toh LDH activity at Time Zero

Representative results are shown in Tables 2A and 2B below:

Table 2A

Table 2B

In vivo Tumor Growth Inhibition Assay: MDA-MB-231 Tumor Xenograft Model

Inhibition of tumor growth in vivo is readily determined via the following assay:

MDA-MB-231 cells are cultured as described above. The cells are harvested by trypsinization, washed, counted, adjusted to 2.5 x 10⁷ cells/mL with ice-cold PBS, and subsequently stored on ice until transplantation. Xenograft experiments are conducted using eight-to-ten week-old female athymic mice with an average body mass of 20-25 g. Approximately 5 x 10⁶ cells in a total volume of 0.2 mL of PBS are injected subcutaneously in the flank region. Thereafter the mice are randomized and divided into several groups that reflect different dosages or schedules, respectively (n= 10 mice/ group). The test compounds are administered starting at day 1 at different dosages (e.g. 10, 20 and 40 mg/kg) and different schedules (e.g. QlDxl5, Q2Dx7, Q3Dx5). Test compounds are formulated for oral administration in a vehicle for oral administration composed of polyethylene glycol-400, ™Cremophor, ethanol and 0.9% saline (40:5:5:50). Tumor measurements are performed twice per week. Tumor weights are calculated using the formula (a x w²)/!. Animals are sacrificed on day 15 after transplantation and plasma was harvested for pharmacokinetic analyses.

In vivo Tumor Growth Inhibition Assay: MX-1 Tumor Xenograft Model

An MX-1 breast, tumor xenograft model is maintained by serial passage in NCr nu/nu female mice (Taconic Farms, Germantown, NY, U.S.A.). Tumors are aseptically harvested from mice when they weigh approximately lg. The envelope and any non- viable areas are dissected and the viable tissue is cut into 3 x 3 x 3 mm cubes. These fragments are implanted in the axilary region of the flank of recipient mice using a trochar.

Treatment in anti-tumor efficacy studies is intiated when all mice have tumors ranging in size from 75-125 mg. There are typically 10 mice in each experimental group. Each experiment contains an untreated control group to monitor tumor growth kinetics, a vehicle-treated control group, and a positive agent control group to assess the response of the model in each experiment to an agent with an expected degree of anti-tumor efficacy. Lack of conformance of any of the controls to the historical ranges for the model constitutes a reason to nullify the study. The test compounds were administered starting at different dosages (e.g. 75 and 150 mg/kg) and different schedules (e.g. qld x 10, bid x 10). Test compounds are formulated for oral administration once per day in a vehicle composed of 51% PEG400/ 12% ethanol/ 12% Cremophor EL/ 0.1 N HCl. Tumor size is recorded in whole mm as measured in two perpendicular dimensions. Animal body weights are recorded in tenths of grams. Both measurements are collected two to three times per week. Animals are sacrificed on day 10 after the last dose and last measurements.

Tumor weights are calculated using the equation (/ x w²)/2, where / and w refer to the larger and smaller dimensions collected at each measurement. Efficacy is measured as the percent suppression of tumor growth expressed as %ΔT/ΔC, where ΔT and ΔC represent the change in the size of the average tumor in the treated and control groups, respectively, over the treatment period. Significance is evaluated using a Student's t-test with a p<0.05. Abbreviations used in this specification

BSA bovine serum albumin calc. calculated

©Cremophor non-ionic emulsifyer from BASF

DBU l,8-diazabicyclo[5.4.0]undec-7-ene

DMEM Dulbecco's Modified Eagle Medium, Life Technologies,

Gaithersburg, MD, U.S.A.

DMF N,N-dimethyl formamide

DMSO dimethyl sulphoxide

EDTA ethylene diamine tetraacetate

FBS fetal bovine serum

HEPES N-(2-hydroxyethyl)-piperazine-N'-(2-ethane sulphonic acid)

HPLC high pressure liquid chromatography

HPLC-ES high pressure liquid chromatography - coupled elecfrospray mass spectroscopy

LC-MS liquid chromato raphy - coupled mass spectroscopy

LC RT liquid chromatography retention time

LDH lactate dehydrogenase

MP melting point

NMR nuclear resonance spectroscopy

PBS phosphate-buffered saline

TFA trifluoroacetic acid tic thin layer chromatography

Tris/HCl tris(hydroxymethyl)-aminomethane hydrochloride

©Triton X-100 tert.-octylphenoxypolyethoxyethanol

The yield percentages of the following Examples refer to the starting component which was used in the lowest molar amount. Examples

LC-MS /HPLC methods

Method A

MS equipment: Micromass Quattro LCZ ionisation mode: ESI positive / negative HPLC equipment: HP 1100

UN detection: 208-400 nm temperature: 40°C

Column: ™Symmetry C 18

50 mm x 2.1 mm 3.5 μm

Supplier: Waters Gradient: Time A: % B: % Flow

[min.] [mL/min.]

0.00 10.0 90.0 0.50

4.00 90.0 10.0 0.50

6.00 90.0 10.0 0.50

6.10 10.0 90.0 1.00

7.50 10.0 90.0 0.50

A: 0.1% sfrength solution of formic acid in acetonitrile

B : 0.1% strength aqueous formic acid

Method B

Column: ™Kromasil C 18 60 mm x 2.0 mm

Gradient: Time A: % B: % Flow [min.] [mL/min.]

0.00 90.0 10.0 0.75

0.50 90.0 10.0 0.75

4.50 10.0 90.0 0.75

6.50 10.0 90.0 0.75

7.50 90.0 10.0 0.75

A: 0.001 % sfrength aqueous H₃PO₄

B: acetonitrile

Method C

MS equipment: Micromass TOF-MUX-Interface 4-fold parallel injection ionisation mode: ESI positive HPLC equipment: Waters 600

UN detection: 210 nm temperature: 40°C

Column: Symmetry C 18

50 mm x 2.1 mm 3.5 μm

Supplier: Waters Gradient: Time A: % B: % Flow

[min.] [mL/min.]

0.00 10.0 90.0 0.75

0.50 10.0 90.0 0.75

4.00 90.0 10.0 0.75

5.50 90.0 10.0 0.75

5.60 10.0 90.0 1.25

6.50 10.0 90.0 0.75

A: 0.1 % strength solution of formic acid in acetonitrile

B : 0.1%) strength aqueous formic acid Method D

MS equipment: Micromass Platform LCZ ionisation mode: ESI positive / negative HPLC equipment: HP 1100

UN detection: . 208-400 nm temperature: 40°C

Column: Symmetry C 18

50 mm x 2.1 mm 3.5 μm

Supplier: Waters Gradient: Time A: % B: % Flow

[min.] [mL/min.]

0.00 10.0 90.0 0.50

4.00 90.0 10.0 0.50

6.00 90.0 10.0 0.50

6.10 10.0 90.0 1.00

7.50 10.0 90.0 0.50

A: 0.1% sfrength solution of formic acid in acetonitrile

B : 0.1% strength aqueous formic acid

Method E

Column: Kromasil C 18 60 mm x 2.0 mm

Gradient: Time A: % B: % Flow

[min.] [mL/min.]

0.00 98.0 2.0 0.75

4.50 10.0 90.0 0.75

6.50 10.0 90.0 0.75

6.70 98.0 2.0 0.75 7.50 98.0 2.0 0.75

A: 0.5% strength aqueous HClO₄

B: acetonitrile

Method F

MS equipment: Finnigan LCQ Ion Trap Mass Spectrometer ionisation mode: ESI

HPLC equipment: HP 1100

UV detection: 254 nm

Column: YMC pro C-18

23 mm x 2 mm 120 A

Supplier: YMC Gradient: Time A: % B: % Flow

[min.] [mL/min.]

0.50 90.0 10.0 1.0

3.50 5.0 95.0 1.0

4.00 5.0 95.0 1.0

4.01 90.0 10.0 1.0 6.50 90.0 10.0 1.0

A: 0.02% strength solution of trifluoroacetic acid in 2 % acetonitrile / 98 % water

B: 0.018% sfrength solution of trifluoroacetic acid in 98 % acetonitrile / 2 % water

The present invention is illusfrated below with the aid of the following non-limiting examples: Starting Materials/Intermediates

Starting Material 1. Phenethylamines

The substituted 2-phenethyl amines are commercially available or can be prepared in analogy to any one of the following procedures, e.g. starting from the corresponding benzaldehydes (see also Shepard et al. in J. Org. Chem. 17, 568 (1952) and in J. Am. Chem. Soc. 72, 4364 (1950)).

Intermediates 1: Amides

Intermediate la: Ethyl 2-{N-[2-(3,4-dimethoxyphenyl)-ethyl]-carbamoyl}-acetate

To a solution of 50.0 g (275.88 mmol) of 3,4-dimethoxy- phenethylamine in 500 mL of dichloromethane was added 42.0 g (275.88 mmol) of l,8-diazabicyclo[5.4.0]undec-7-ene, followed by dropwise addition of 35.0 mL (41.62 g, 276.43 mmol) of ethyl malonyl chloride at a rate that kept the internal temperature below 30°C. The resultant clear yellow solution was stirred at room temperature under an argon atmosphere for 16 hours, at which time TLC analysis (silica gel 60, methanol/dichloromethane (5:95), UV detection) suggested complete reaction. The organics were washed with brine (3 x 1000 mL), dried over sodium sulfate and concentrated. The residue was dried under high vacuum at 30°C for 24 hours to provide 80.55 g (272.75 mmol, 99%) of a yellow oil. 1H-NMR (OMSO-d₆): δ = 1.16 (t, J= 7.0 Hz, 1.5H); 1.18 (t, J

= 7.0 Hz, 1.5H); 2.63 (t, J= 7.7 Hz, 2H); 3.18 (s, 2H); 3.25 (m,

2H); 3.70 (s, 3H); 3.73 (s, 3H); 4.05 (q, J= 7.0, 2H); 6.69 (dd,

J= 2.2 Hz, 8.4 Hz, IH); 6.79 (d, J= 2.2 Hz, IH); 6.83 (d, J =

8.4 Hz, IH); 8.1 (bt, J= 5.4 Hz, IH).

MS (HPLC/ES): m/z = 296 (M + 1).

TLC (10:90 methanol/dichloromethane): R = 0.70.

The following amides were obtained in analogy to the described procedure:

Intermediate lb: Ethyl 3- { [2-(3 -methoxyphenyl)-ethyl] -amino } -3-oxopropanoate Intermediate lc: Ethyl 3- {[2-(4-methoxyphenyl)-ethyl] -amino} -3-oxopropanoate

Intermediates 2 : 3 ,4-Dihydro- 1 (2H)-isoquinolinylidene-ethanoates

intermediate 2a: Ethyl (2E)-(6,7-dimethoxy-3 ,4-dihydro- 1 (2H)-isoquinolinyli- dene)-ethanoate

To a refluxing solution of methane sulfonic anhydride (648.83 g, 3.72 mol) in toluene (4 L) was added intermediate la, ethyl 2- {N-[2-(3,4-dimethoxyphenyl)-ethyl] -carbamoyl }- acetate, (1000 g, 3.39mol) portionwise over 20 minutes. The reaction was stirred at reflux for 30 minutes at which point the heat was removed and the toluene was decanted. The resulting dark oil was then dissolved in water (3000 ml) and freated portionwise with solid potassium carbonate until a pH of about 5 8 was achieved. The organic material was extracted from the dark biphasic mixture using ethyl acetate (3000 mL). The combined organic extracts were washed with brine (3 x 2000 mL) and concentrated to 1/3 volume. The resultant dark oil was placed on a pad of silica gel 60 (400 cc) and eluted

10 using ethyl acetate/hexanes (1:1). The desired fractions were concentrated to a yellow oil which was seeded with a small amount of crystals of the title compound and placed in a refrigerator overnight. The yellow crystalline solid which formed was filtered, washed with ethyl acetate / hexanes (1:1)

15 (2 x 50 ml), and vacuum dried for 12 hours to give the desired product (533.26 g). The filtrate was concentrated to a dark oil and seeded a second time. After 1 hour, the newly formed yellow solid was filtered and washed with ethyl acetate/ hexanes (1:1) (2 x 50 mL) and vacuum dried for 12 hours to

20 provide a second crop (106.23 g). The two batches of crystals were combined to provide the title compound (639.49 g, 68 %).

1H-NMR (OMSO-d₆): δ 1.18 (t, J = 7.0 Hz, 3H); 2.76 (t, J = 6.5 Hz, 2); 3.36 (m, 2H); 3.78 (s, 6H); 4.02 (q, J = 7.0 Hz, 2H);

25 5.05 (s, IH); 6.87 (s, IH); 7.15 (s, IH); 8.95 (bs, IH). MS

(HPLC/ES): m/z = 278 (M + 1). TLC (ethyl acetate/ hexanes (1:1)): R = 0.63

Instead of methane sulfonic anhydride also phosphorous pent- 30 oxide can be used according to this method. The following 3,4-dihydro-l(2H)-isoquinolinylidene-ethanoates were obtained in analogy to the described procedure:

Intermediate 2b: Ethyl (2E,Z)-(6-memoxy-3,4-dihydro-l(2H -isoqumolinylidene)- ethanoate

intermediate 2c: Ethyl (2E,Z)-(7-methoxy-3,4-dihydro- 1 (2H)-isoquinolinylidene)- ethanoate

intermediates 3: 5.6-Dihvdro-pyrrolo[2, 1 -a]isoquinoline- 1 -carboxylates

Intermediate 3 a: Ethyl 2-(3-chlorophenyl)-8,9-dimethoxy-3-methyl-5,6-dihydro- pyrrolo[2, 1 -a]isoquinoline-l -carboxylate

A mixture of 10.06 g (0.04 mol) of Intermediate 2a, 5.1 g (0.04 mol) of 3-chloro-benzaldehyde, 2.72 g (0.04 mol) of nifroethane and 0.54 mL (0.01 mol) of piperidine in 45 mL of ethanol was refluxed for 80 hours. It was cooled, the obtained crystals were sucked off and washed carefully with isopropanol. 10.74 g of a nearly colorless solid compound of melting point 132-133 °C were obtained. The following 5,6-dihydro-pyrrolo[2,l-a]isoqumoline-l-carb- oxylates were obtained in analogy to the described procedure :

Intermediate 3b: Ethyl 2-(4-hydroxy-3,5-dimethylphenyl)-8,9-dimethoxy-3- methyl-5,6-dihydropyrrolo[2, 1 -a]isoquinoline- 1 -carboxylate using 3,5-dimethyl-4-hydroxy-benzaldehyde instead of 3- chloro-benzaldeyde

Intermediate 3c: Ethyl 8,9-dimethoxy-2-[4-(methoxycarbonyl)-phenyl]-3-meth- yl-5,6-dihydropyrrolo[2, 1 -a]isoquinoline- 1 -carboxylate using 4-methoxycarbonyl-benzaldehyde instead of 3-chloro-benzaldehyde

Intermediate 3d: Ethyl 2-(3-chlorophenyl)-8-methoxy-3-methyl-5,6-dihydro- pyrrolo[2,l-a]isoquinoline-l -carboxylate using ethyl (6-meth- oxy-3,4-dihydro-l(2H)-isoquinolinylidene-ethanoate instead of ethyl (6,7-dimethoxy-3 ,4-dihydro- 1 (2H)-isoquinolinylidene- ethanoate

Intermediate 3e: Ethyl 8,9-dimethoxy-2-(3-methoxyphenyl)-3-methyl-5,6-dihy- dropyrrolo [2, 1 -a] isoquinoline- 1 -carboxylate using 3 -methoxy- benzaldehyde instead of 3-chloro-benzaldehyde

Intermediate 4: Ethyl 2-(3-chlorophenyl)-8,9-dimethoxy-3-chloromethyl-5,6- dihydro-pyrrolo[2, 1 -ajisoquinoline- 1 -carboxylate

200 mg (0,47 mmol) of ethyl 2-(3-chlorophenyl)-8,9-dimeth- oxy-3 -methyl-5,6-dihydropyrrolo[2, 1 -ajisoquinoline- 1 -carboxylate (Intermediate 3 a) were dissolved in 1,3 mL of dichloromethane, and a solution of 65,28 mg (0,48 mmol) of sulfuryl chloride in 1 mL of dichloromethane was added dropwise at 0°C. The color of the solution changed from yellow to red and finally to brown. It was stirred for 10 minutes under ice cooling and for 1 hour at room temperature. The solvent was evaporated, and after addition of dichloromethane the solvent was evaporated again. The raw chloromethyl compound thus obtained was further reacted without purification.

Instead of sulfuryl chloride also thionyl chloride or N-chloro- succinimide can be used according to this method.

Intermediate 5: Ethyl 3 -(chloromethyl)- 8 ,9-dimethoxy-2-(3 -methoxyphenyl)- 5,6-dihydropyrrolo[2, 1 -a]isoquinoline- 1 -carboxylate

To a 0°C solution of 200 mg (0.475 mmol) of ethyl 8,9-di- methoxy-2-(3-methoxyphenyl)-3-methyl-5,6-dihydropyrrolo- [2, l-a]isoquinoline-l -carboxylate (Intermediate 3e) in 2 mL of dry dichlomethane were added 0.475 mL (0.475 mmol) sulfuryl chloride (1 M in dichlomethane ). The reaction was stirred at 0°C for 10 minutes and then at room temperature for 1 hour. The solvent was removed in vacuo and the residue was used without further purification.

Intermediate 6: Ethyl 2-(3-chlorophenyl)- 1 -nitroethene-carboxylate

A solution of 28,5 g (150 mmol) of titanium tefrachloride in 40 mL of tefrachloromethane was added dropwise to 300 mL of ice-cooled THF under an argon atmosphere. Into the suspension thus obtained 10,56 g (75,13 mmol) of 3-chloro- benzaldehyde and 10,0 g (75,13 mmol) of nitroacetic acid ethyl ester were added simultaneously from two dropping funnels at 0°C. Thereafter, 30,4 g (300.52 mmol) of N-methyl morpho- line were added dropwise within 2 hours at 0°C. It was stirred overnight at 0°C and then allowed to warm up until room temperature. The solution was carefully reacted with water 5 under cooling. 400 mL of diethyl ether were added, the layers separated, the aqueous layer extracted twice with diethyl ether, the combined organic layers washed with aqueous sodium chloride solution and dried, and the solvent was evaporated. The residue was crystallized with ethanol/petrolether (1:1). 10 5,05 g (26,4 %) of crystals having a melting point of 65-66°C were obtained.

Preparation Examples

Example 1

Ethyl 2-(3-chlorophenyl)-8,9-dimethoxy-3-N-morpholinomethyl-5,6-dihydro-pyr- rolo[2, 1 -a]isoquinoline- 1 -carboxylate

The unpurified chloromethyl derivative Intermediate 4 (raw material obtained from

0.47 mmol of Intermediate 3a) was dissolved in 3 mL of dichloromethane and reacted dropwise with a solution of 2 mmol (174 mg) of morpholine in 2 mL of dichloromethane. The mixture was stirred overnight, diluted with dichloromethane, washed with water and dried, and the solvent was evaporated. After a preliminary purification by means of a cartridge 118 mg (49,2 %, based on 0.47 mmol of

Intermediate 3a) of colorless crystals having a melting point of 186-187°C were obtained.

Example 2

Ethyl 2-(3-chlorophenyl)-8,9-dimethoxy-3-methoxymethyl-5,6-dihydro-pyrrolo[2, 1 - a]isoquinoline- 1 -carboxylate

The unpurified chloromethyl derivative intermediate 5 (51 mg, 0,11 mmol) in methanol was reacted with 0,2 mL of a 30 % solution of sodium methylate in methanol. The solvent was evaporated after 2 hours, the residue was taken up in ethyl acetate, the solution was washed with water, and the solvent was evaporated. Purification was conducted by means of a cartridge. 20 mg (40 %) of colorless crystals having a melting point of 116-117°C were obtained. 1H NMR (300 MHz, DMSO-d₆): δ = 0.89 (t, 3H), 2.97 (t, 2H), 3.20 (s, 3H), 3.73 (s, 3H), 3.81 (s, 3H), 3.93-4.08 (m, 4H), 4.27 (s, 2H), 6.98 (s, IH), 7.13-7.19 (m, IH),

7.22 (s, IH), 7.33-7.48 (m, 2H), 7.72 (s, IH). MS: 455 (M⁺) HPLC retention time [min]: 5 (method E)

The following Examples (Nos. 3-12) were carried out in analogy to the description of

Examples 1 and 2: D)

D)

(t,

D)

Example 13

Ethyl 2-(3-chlorophenyl)-8-methoxy-3-nitro-5,6-dihydro-pyrrolo[2, 1 -a]isoquinoline- 1 -carboxylate

840 mg (2,12 mmol) of ethyl 2-(3-chlorophenyl)-8-methoxy-3-methyl-5,6-dihydro- pyrrolo[2,l-a]isoquinoline-l -carboxylate (Intermediate 3d) were suspended in 16,27 mL of glacial acetic acid, and a mixture of 0,31 mL (4,45 mmol) of 65 % nitric acid in 1 mL of glacial acetic acid was added dropwise at 20°C. The reaction solution changed from green to orange red. After 2 hours the solution was poured into ice water, the mixture was extracted twice with dichloromethane, the combined organic layers were washed with water and dried, and the solvent was evaporated. The purification was made by column chromatography with dichloromethane. 153 mg of crystals having a melting point of 189- 190°C were obtained.

Starting from intermediate 3 a, the following compound was prepared in analogy to Example 13):

Example 15

Ethyl 3-amino-8-methoxy-2-(3-chlorophenyl)-5,6-dihydro-pyrrolo[2, 1 -a]isoquino- line- 1 -carboxylate

80 mg (0,18 mmol) of the compound of Example 13 were dissolved in a mixture of 1 mL of pyridine and 5 mL of ethanol and, after addition of 40 mg of Pd/C (10%), hydrogenated with hydrogen gas at normal pressure for 4 hours. The mixture was filtrated, the solvent was evaporated, and the compound was purified with cyclo- hexane/ethyl acetate gradient mixtures over a silica gel column. 35 mg (49,6 %) of little yellowish crystals having a melting point of 150-152°C were obtained.

n analogy to Example 15, the following compound was prepared from the compound of Example 14:

Examples 17a and 17b

Ethyl 3-formyl-8,9-dime oxy-2-(3-cUorophenyl)-5,6-άO ydro-pyrrolo[2, 1 -a]isoquino- line- 1 -carboxylate

Example 17 a: A mixture of 100 mg (0,234 mmol) of ethyl 2-(3-chlorophenyl)-8,9- dimethoxy-3-methyl-5,6-dihydro-pyrrolo[2, 1 -a]isoquinoline- 1 -carboxylate (Inter- mediate 3a) and 500 mg of manganese dioxide in 3 mL of dioxane was stirred for 2 hours at 100°C. The mixture was cooled, filtrated, and the solvent was evaporated.

The resulting mixture of products was separated with toluene/ ethyl acetate (until 4:1) over a silica gel column. 27 mg (26,2 %) of colorless crystals having a melting point of 180-181 °C were obtained. 1H NMR (200 MHz, CDC1₃): δ = 0.94 (t, 3H), 3.02 (t, 2H), 3.89 (s, 3H), 3.93 (s, 3H),

4.08 (q, 2H), 4.69 (t, 2H), 6.78 (s, IH), 7.13-7.44 (m, 4H), 7.87 (s, IH), 9.35 (s, IH). Example 17b: As a second compound the respective dehydro compound ethyl 3- formyl-8,9-dimethoxy-2-(3-chlorophenyl)-pyrrolo[2, 1 -a]isoquinoline- 1 -carboxylate having a melting point of 155-157°C could be obtained:

Starting from Intermediate 3 c, the following compound was prepared in analogy to Example 17a:

Example 19

Ethyl 3-N-morpholinomethyl-8,9-dimethoxy-2-(3-chlorophenyl)-pyrrolo[2, 1 -a]iso- quinoline- 1 -carboxylate

100 mg (0,23 mmol) of ethyl 2-(3-chlorophenyl)-8,9-dimethoxy-3-methyl-5,6-di- hydro-pyrrolo[2,l-a]isoquinoline-l-carboxylate (Intermediate 3a) were stirred in 0,4 mL of thionyl chloride for 70 minutes under moderate reflux conditions. The solvent was evaporated, the residue was taken up with dichloromethane, the solvent was evaporated and the residue was dissolved in 4 mL of dichloromethane. 87 mg (1 mmol) of morpholine were added dropwise;, the reaction mixture was stirred for 3 hours at 20°C, diluted with dichloromethane and shaken twice with water. The solvent was evaporated and the compound was purified over a short silica gel column with dichloromethane/ethyl acetate (until 1:1). 43 mg (36,7 %) of colorless crystals having a melting point of 176-177°C were obtained.

Example 20

Diethyl 8,9-dimethoxy-2-(3-chlorophenyl)-5,6-dihydro-pyrrolo[2, 1 -a]isoquinoline- 1,3-dicarboxylate

1,0 g (3,93 mmol) of Intermediate 6 was added to a solution of 1,09 g (3,93 mmol) of ethyl (6,7-dimethoxy-3 ,4-dihydro- 1 (2H)-isoquinolinylidene)-ethanoate (Intermediate 2a) in 40 mL of isopropanol. The solution was stirred for 17 hours under moderate reflux conditions and then cooled with an ice bath. The precipitated crystals were sucked off and washed with isopropanol. 1,179 g (62 %) of crystals having a melting point of 140-141 °C were obtained.

Example 21

Ethyl 8,9-dimethoxy-2-(3-chlorophenyl)-5,6-dihydro-pyrrolo[2, 1 -ajisoquinoline- 1 - carboxylate

In analogy to the procedure described for Intermediate 3 a, the title compound was obtained using nitromethane instead of nifroethane. MS: 412.2 (M+H)

HPLC retention time [min]: 4.86 (method C)

Example 22

Ethyl 9-methoxy-2-(2-methylphenyl)-3-methylcarbonyl-5,6-dihydro-pyrrolo[2,l-a]- isoquinoline- 1 -carboxylate

150 mg (0,61 mmol) of ethyl (7-methoxy-3,4-dihydro-l(2H)-isoquinolinylidene)- ethanoate (Intermediate 2c) and 248,95 mg (1,21 mmol) of 4-(2-methylphenyl)-3- nifro-3-buten-2-one (prepared from 2-methyl-benzaldeyde and l-nitro-propane-2- one) in 2 mL of ethanol were refluxed for 24 hours. The solvent was evaporated to dryness, and the desired compound was separated on a silica gel column with dichloromethane. A yield of 52 mg (21,25 % of theory) was obtained.

1H NMR (200 MHz, CDC1₃): δ = 0.84 (t, J = 7.2 Hz, 3H), 1.79 (s, 3H), 2.18 (s, 3H),

2.98 (t, J = 6.4 Hz, 2H), 3.81 (s, 3H), 3.98 (q, J = 7.2 Hz, 2H), 4.43-4.78 (m, 2H),

6.86 (dd, J = 8.2 Hz, J = 2.5 Hz, IH), 7.03-7.30 (m, 4H), 7.58 (d, J = 2.5 Hz, IH).

MS: 404.3 (M+H)

HPLC retention time [min]: 4.7 (method C)

Example 23

Ethyl 2-(4-hydroxy- 1 -naphthyl)-8,9-dimethoxy-3-(3-methoxy-3-oxopropyl)-5,6-di- hydropyrrolo[2, 1 -ajisoquinoline- 1 -carboxylate

In analogy to the procedure described for Intermediate 3 a, the title compound was obtained using ethyl (6,7-dimethoxy-3,4-dihydro-l(2H)-isoquinolinylidene)-ethano- ate (Intermediate 2a), 4-hydroxy-l-naphthaldehyde and methyl 4-nitrobutanoate. MS: 530.2 (M+H) HPLC retention time [min]: 4.33 (method A)

Example 24

Ethyl 2-(4-hydroxy-3,5-dimethylphenyl)-8,9-dimethoxy-3-(3-methoxy-3-oxopropyl)- 5,6-dihydropyrrolo[2,l-a]isoquinoline-l-carboxylate

In analogy to the procedure described for Intermediate 3a, the title compound was obtained using ethyl (6,7-dimethoxy-3,4-dihydro-l(2H)-isoquinolinylidene)-ethano- ate (Intermediate 2a), 4-hydroxy-3,5-dimethyl-benzaldehyde and methyl 4-nitro- butanoate.

^!H NMR (300 MHz, DMSO-d₆): δ = 0.90 (t, J = 7.0 Hz, 3H), 2.15 (s, 6H), 2.38-2.53 (m, 2H), 2.73-2.84 (m, 2H), 2.93 (t, J = 6.1 Hz, 2H), 3.54 (s, 3H), 3.70 (s, 3H), 3.78 (s, 3H), 3.89-4.02 (m, 4H), 6.70 (s, 2H), 6.94 (s, IH), 7.52 (s, IH), 8.09 (s, IH) MS: 508.4 (M+H), 525.4 (M+NH₄) HPLC retention time [min]: 4.54 (method B)

Example 25

Ethyl 2-(lH-indol-3-yl)-8,9-dimethoxy-3-(3-methoxy-3-oxopropyl)-5,6-dihydro- pyrrolo [2, 1 -a]isoquinoline- 1 -carboxylate

In analogy to the procedure described for intermediate 3 a, the title compound was obtained using ethyl (6,7-dimethoxy-3,4-dihydro-l(2H)-isoquinolinylidene)-ethano- ate (Intermediate 2a), lH-indole-3-carbaldehyde and methyl 4-nitrobutanoate. MS: 503.2 (M+H) HPLC retention time [min]: 4.34 (method A) Example 26

Ethyl 2-[3-(dimethylamino)-phenyl]-8,9-dimethoxy-3-(4-methoxybenzyl)-5,6-di- hydropyrrolo[2, 1 -ajisoquinoline- 1 -carboxylate

In analogy to the procedure described in Intermediate 3a, the title compound was obtained using ethyl (6,7-dimethoxy-3,4-dihydro-l(2H)-isoquinolinylidene)-ethano- ate (Intermediate 2a), 3-(dimethylamino)-benzaldehyde and l-methoxy-4-(3-nifropro- pyl)-benzene.

1H NMR (300 MHz, DMSO-d₆): δ = 0.91 (t, J = 7.2 Hz, 3H), 3.24-3.38 (m, 8H), 3.62-3.81 (m, 11H), 3.86-4.07 (m, 4H), 6.44-6.54 (m, 2H), 6.57-6.66 (m, IH), 6.79- 7.01 (m, 5H), 7.05-7.19 (m, IH), 7.62 (s, IH)

MS: 541.0 (M+H) HPLC retention time [min]: 4.5 (method B) Example 27

Ethyl 8,9-dimethoxy-3-(4-methoxybenzyl)-2-(3,4,5-trimethoxyphenyl)-5,6-dihydro- pyrrolo[2, 1 -a]isoquinoline- 1 -carboxylate

In analogy to the procedure described for intermediate 3a, the title compound was obtained using ethyl (6,7-dimethoxy-3,4-dihydro-l(2H)-isoquinolinylidene)-ethano- ate (intermediate 2a), 3,4,5-trimethoxy-benzaldehyde and l-methoxy-4-(3-nitro- propyl)-benzene.

1H NMR (300 MHz, DMSO-d₆): δ = 0.92 (t, J = 7.0 Hz, 3H), 2.79-2.90 (m, 2H), 3.57-3.81 (m, 20H), 3.87-4.09 (m, 4H), 6.43 (s, 2H), 6.85 (d, J = 8.7 Hz, 2H), 6.90 (s, IH), 6.98 (d, J = 8.7 Hz, 2H), 7.68 (s, IH)

MS: 588.0 (M+H), 610.0 (M+Na) HPLC retention time [min]: 5.12 (method B) Example 28

Ethyl 2-(3,5-dimethyl-4-hydroxy-phenyl)-8,9-dimethoxy-3-N-morpholinomethyl-5,6- dihydro-pyrrolo[2, 1 -a]isoquinoline- 1 -carboxylate

10.73 mg (0.08 mmol) of N-chlorosuccinimide were added to a 0°C solution of 35 mg (0.08 mmol) of ethyl 2-(4-hydroxy-3,5-dimethylphenyl)-8,9-dimethoxy-3- methyl-5,6-dihydro-pyrrolo[2,l-a]isoquinoline-l -carboxylate (intermediate 3b) in

1 mL of dichloromethane . The mixture was stirred for 1 hour at room temperature, the solvent was evaporated in vacuo, the residue was solved again in 1 mL of dichloromethane and freated with 3 drops of morpholine. The mixture was diluted with dichloromethane after 3 hours, washed with water, dried, and the solvent was evaporated. After chromatography on silicagel 10 mg of crystals having a melting point of 195°C were obtained.

Example 29

Ethyl 8,9-dimethoxy-3-(methoxymethyl)-2-(3-methoxyphenyl)-5,6-dihydropyrrolo- [2, 1 -a]isoquinoline- 1 -carboxylate

647.4 mg (1.420 mmol) of ethyl 3-(chloromethyl)-8,9-dimethoxy-2-(3-methoxy- phenyl)-5,6-dihydropyrrolo[2,l-a]isoquinoline-l-carboxylate (intermediate 5) were dissolved in methanol and stirred for 15 minutes at which time the reaction looked complete by LC-MS. Purification by chromatography [Biotage System, 40S column, 32-63 μm silica, eluant: ethyl acetate/ hexanes (1:3)] followed by recrystallization with methanol provided 130 mg (0.29 mmol, 27.29%) of the title compound as a white solid. MS (HPLC/ES): m/z = 452.5 (M + 1).

HPLC RT (Method F): 3.33 min. TLC [ethyl acetate/ hexanes (3:7)]: R_f = 0.18.

Numerous modifications and variations in the invention as described in the above illustrative examples are expected to occur to those skilled in the art and consequently only those limitations as appear in the appended claims should be placed thereon.

Accordingly it is intended in the appended claims to cover all such equivalent variations wliich come within the scope ofthe invention as claimed.

Claims

We claim:

1. A compound of the formula

wherein

x and y independently from each other denote zero or 1 and x+y is 1 or 2;

1 9

R and R independently from each other denote hydrogen, C_1-4-alkyl or CF₃, or R¹ and R² together form a C_1-4-alkylene bridge;

R denotes hydrogen, formyl, (C_1-4-alkyl)-carbonyl, (Cι- -alkoxy)-carbonyl, NO₂, NR⁶R⁷, C_1-4-alkyl-NR⁶R⁷, C_1-4-alkyl-OR⁸, C_1-4-alkyl-COOR⁸, C_6-10- aryl-C_1-4-alkyl wherein the aryl moiety is optionally substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl, and C_1-4- alkoxy;

wherein

R⁶ and R⁷ independently from each other denote hydrogen, C_1-4-alkyl, C_3-8-cycloalkyl, or C_6-10-aryl-C_1-4-alkyl wherein the aryl moiety is optionally substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl and C_1-4-alkoxy; or

R and R together with the nifrogen atom to wliich they are attached form a 5- to 7-membered heterocyclyl which may contain up to 2 further hetero atoms selected from the group consisting of N, O, and S, wliich heterocyclyl can further be substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl, C_1-4-alkoxy, C_6- ₁₀-aryl and aromatic 4- to 9-membered heterocyclyl with 1 to 4 hetero atoms selected from the group consisting of N, O and S;

R denotes hydrogen or C_1-4-alkyl;

R ,4 denotes C_1-4-alkyl;

R⁵ is

halogen;

C_1-6-alkyl which can be further substituted with one or more radicals selected from the group consisting of C_1-6-alkoxy, OH and NH₂;

C_1-6-alkoxy which can be further substituted with one or more radicals selected from the group consisting of C_1-6-alkoxy, OH, and NH ;

C_6-10-aryloxy-C_1-6-alkoxy; OH; NO₂; CN; 5 CF₃;

OCF₃; NR⁹R¹⁰; CONR⁹R¹⁰; COOR¹¹; 10 SR¹¹;

SOR¹¹; SO₂R^u; OSO₂R^u;

15 -O-(CH₂)_1-4-O- wherein the oxygen atoms are bound to the aryl moiety in ortho-position to each other;

phenyloxy or benzyloxy wherein the phenyl moieties can contain one further substituent selected from the group 20 consisting of C_1-6-alkyl, C_1-6-alkoxy, halogen and NO₂;

phenyl optionally substituted with CN;

aromatic 4- to 9-membered heterocyclyl with 1 to 4 hetero 25 atoms selected from the group consisting of N, O and S;

and

saturated 5- to 7-membered nitrogen-containing heterocyclyl 30 which is bound to the C_6-10-aryl moiety via the nitrogen atom and may contain up to 2 further heteroatoms selected from the group consisting of N, O and S and which saturated heterocyclyl can be further substituted with one or more radicals selected from the group consisting of C_1-6-alkoxy, OH and 5 NH₂;

wherein

R⁹ and R¹⁰ independently from each other denote 10 hydrogen, C_1-6-alkyl, (C_1-6-alkyl)-carbonyl, (C_1-6- alkoxy)-carbonyl, C_1-6-alkylsulfonyl, (C_1-6- alkylamino)-carbonylamino, (C_6-1o-arylamino)- carbonylamino,

15 or

R⁹ and R¹⁰ together with the nitrogen atom to which they are attached, form a 5- to 7-membered saturated, partially unsaturated or aromatic heterocyclyl which

20 can contain up to 3 further hetero atoms selected from the group consisting of N, O and S, and which heterocyclyl can contain 1 to 3 substituents selected from the group consisting of C_1-6-alkyl, C_1-6-alkoxy, C_6-1o-aryl and aromatic 4- to 9-membered heterocyclyl

25 with 1 to 4 hetero atoms selected from the group consisting of N, O and S;

R^u is hydrogen, C_1-6-alkyl or C_6-10-aryl;

30 or ϋ) C_1-12-alkyl which can contain 1 to 3 substituents selected from the group consisting of C_1-6-alkyl, C-_1-6-alkoxy, C_6-10-aryl and aromatic 4- to 9-membered heterocyclyl with 1 to 4 hetero atoms selected from the group consisting of N, O and S;

or iii) C_3-8-cycloalkyl wliich can contain 1 to 3 substituents selected from the group consisting of C_1-6-alkyl, C-_1-6-alkoxy, COOR¹¹ wherein R¹¹ is as defined above, C_6-10-aryl and aromatic 4- to

9-membered heterocyclyl with 1 to 4 hetero atoms selected from the group consisting of N, O and S;

or

iv) aromatic C_2-9-heterocyclyl with 1 to 4 hetero atoms selected from the group consisting of N, O and S which aromatic heterocyclyl can contain 1 to 3 further substituents selected from the group consisting of OH, C_1-6-alkyl, C-_1-6-alkoxy, C_6-10-aryl which can contain 1 to 3 halogen radicals, COR¹¹ or

COOR¹¹ wherein R¹¹ is as defined above, halogen, CN, and saturated 5- to 7-membered nitrogen-containing heterocyclyl which is bound to the C_6-10-aryl moiety via the nitrogen atom and can contain up to 2 further hetero atoms selected from the group consisting of N, O and S and which saturated heterocyclyl can be further substituted with one or more radicals selected from the group consisting of C_1-6-alkoxy, OH and NH₂; with the proviso that ethyl 8,9-dimethoxy-2-phenyl-5,6-dihydropyrrolo[2.1- a]isoquinoline-l -carboxylate is excluded;

2. A compound according to claim 1, wherein

x and y independently from each other denote zero or 1 and x+y is 1 or 2;

R and R independently from each other denote C_1-4-alkyl or CF₃;

R denotes hydrogen, formyl, (C_1-4-alkyl)-carbonyl, (C_1- -alkoxy)-carbonyl, NO₂, NR₆R₇, C -alkyl-NR^⁷, C_1-4-alkyl-OR⁸, C -alkyl-COOR⁸, or C_6-10- aryl-C_1- -alkyl wherein the aryl moiety can be substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl, and C_1-4-alkoxy;

wherein

R and R independently from each other denote hydrogen, C_1-4-alkyl,

C_3-8-cycloalkyl, or C_6-10-aryl-C_1-4-alkyl wherein the aryl moiety can be substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl, and C_1- -alkoxy;

or

R⁶ and R⁷ together with the nitrogen atom to which they are attached form a 5- to 7-membered heterocyclyl which may contain up to 2 further hetero atoms selected from the group consisting of N, O, and S and which heterocyclyl can be further substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl, C_1-4-alkoxy, C_6- ₁₀-aryl and aromatic 4- to 9-membered heterocyclyl with 1 to 3 hetero atoms selected from the group consisting of N, O, and S;

R denotes hydrogen or C_1-4-alkyl;

R⁴ denotes C_1-4-alkyl;

R⁵ is

i) phenyl optionally having 1 to 3 further substituents selected from the group consisting of F, CI, Br; C_1-6-alkyl; C_1-6-alkoxy; OH; NR⁹R¹⁰ and COOR^u;

or

ii) naphthyl optionally containing one further OH group;

or

wherein R⁹ to R¹¹ independently from each other denote C_1-6-alkyl;

with the proviso that ethyl 8,9-dimethoxy-2-phenyl-5,6-dihydropyrrolo[2.1- a]isoquinoline-l -carboxylate is excluded;

and an isomer, a pharmaceutically acceptable salt, a hydrate or a hydrate of a pharmaceutically acceptable salt thereof. A compound according to claim 1, wherein

x and y independently from each other denote zero or 1 and x+y is 1 or 2;

R¹ and R² independently from each other denote CH₃ or C₂H₅;

R³ denotes hydrogen, formyl, (C_1-4-alkyl)-carbonyl, (C_1-4-alkoxy)-carbonyl, NO₂, NR⁶R⁷, C_1-4-alkyl-NR⁶R⁷, C_1-4-alkyl-OR⁸, C_1-4-alkyl-COOR⁸, or C_6-10- aryl-C_1-4-alkyl wherein the aryl moiety can be substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl, and C_1-4-alkoxy;

wherein

R⁶ and R⁷ independently from each other denote hydrogen, C_1-4-alkyl,

C_3-8-cycloalkyl, or C_6-10-aryl-C_1-4-alkyl wherein the aryl moiety can be substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl, and C_1-4-alkoxy;

or

R⁶ and R⁷ together with the nitrogen atom to which they are attached form a 5- to 7-membered heterocyclyl which may contain up to 2 further hetero atoms selected from the group consisting of N, O, and S and which heterocyclyl can be further substituted with 1 to 3 radicals selected from the group consisting of OH, C_1-4-alkyl, C_1-4-alkoxy, C_6- io-aryl and aromatic 4- to 9-membered heterocyclyl with 1 to 3 hetero atoms selected from the group consisting of N, O, and S;

R⁸ denotes hydrogen or C_1-4-alkyl; >4 denotes CH₃ or C₂H₅;

R⁵ is i) phenyl optionally having 1 to 3 further substituents selected from the group consisting of CI; C_1-4-alkyl; C_1-4-alkoxy; OH; NR⁹R¹⁰, and COOR¹¹;

or

ii) naphthyl optionally containing one further OH group;

or

iv) indolyl optionally having 1 to 3 further substituents selected from the group consisting of F, CI, Br; C_1-6-alkyl; C_1-6-alkoxy; OH; NR⁹R¹⁰, arid COOR¹¹;

wherein R⁹ to R¹¹ independently from each other denote C_1-4-alkyl;

4. A compound according to claim 1, wherein

x and y independently from each other denote zero or 1 and x+y is 1 or 2;

R¹ and R² independently from each other denote CH₃ or C₂H₅; R³ denotes hydrogen, formyl, (C_1-4-alkyl)-carbonyl, (C_1-4-alkoxy)-carbonyl, NO₂, NH₂, C_1-4-alkyl-NR⁶R⁷, C_1-4-alkyl-OR⁸, C_1-4-alkyl-COOR⁸, or phenyl- C_1-4-alkyl wherein the phenyl moiety can be substituted with 1 to 3 C_1-4-alkyl or C^-alkoxy moieties;

wherein

R⁶ and R⁷ independently from each other denote hydrogen, C_1- -alkyl, C_3-6-cycloalkyl, or phenyl-C_1-4-alkyl wherein the phenyl moiety can be substituted with 1 to 3 C_1-4-alkyl or C_1-4-alkoxy radicals;

or

R⁶ and R⁷ together with the nitrogen atom to which they are attached, form a saturated 5- to 7-membered heterocyclyl which may contain up to 2 further hetero atoms selected from the group consisting of N, O, and S and which saturated heterocyclyl can be further substituted with 1 to 3 radicals selected from the group consisting of C_1-4-alkyl, C_1-4- alkoxy, phenyl and pyridyl;

R denotes hydrogen or C_1-4-alkyl;

R⁴ denotes CH₃ or C₂H₅;

R⁵ is

i) phenyl optionally having 1 to 3 further substituents selected from the group consisting of CI; C_1-4-alkyl; C_1-4-alkoxy; OH; NR⁹R¹⁰; and COOR¹¹; wherein R⁹ to R¹⁰ independently from each other denote d- alkyl;

or

ii) naphthyl optionally containing one further OH group;

or

iii) indolyl;

with the proviso that ethyl 8,9-dimethoxy-2~phenyl-5,6-dihydropyrrolo[2.1- a]isoquinoline-l -carboxylate is excluded;

5. A compound selected from the group consisting of the compounds of the formulae

A process for manufacturing a compound of claims 1 to 5 comprising the reaction of a compound ofthe formula

wherein x, y, R , R and R are as defined in claims 1 to 5,

[A] with the compounds ofthe formulae

R⁵-CHO and R³-CH₂-NO₂ (II) (HI)

wherein R³and R⁵ are as defined in claims 1 to 5, or

[B] with a compound ofthe formula

wherein R³ and R⁵ are as defined in claims 1 to 5, and optionally

[C] the conversion of the compound obtained through either process [A] or [B] into an isomer, a pharmaceutically acceptable salt, a hydrate or a hydrate of a pharmaceutically acceptable salt thereof.

A compound according to claims 1 to 5 for use in a medicinal application.

A compound according to claims 1 to 5 for combating cancer.

9. A method of manufacturing a pharmaceutical composition by combining at least one ofthe compounds according to claims 1 to 5 with at least one phar- rnacologicaUy acceptable formulating agent.

10. A pharmaceutical composition comprising as an active ingredient an effective amount of at least one of the compounds of claims 1 to 5 and at least one pharmacologically acceptable formulating agent.

11. A pharmaceutical composition comprising as an active ingredient an effective amount of at least one of the compounds of claims 1 to 5 and at least one pharmaceutically active ingredient which is different from the compounds of claims 1 to 5.

12. A method of combating cancer in humans and animals comprising the administration of an effective amount of at least one compound of claims 1 to

5.

13. A medicament in unit dosage form comprising an effective amount of a compound of claims 1 to 5.

14. Use of at least one ofthe compounds of claims 1 to 5 for the manufacture of a medicament for combating cancer.