WO2002030906A1 - Anti-tumour compounds - Google Patents

Abstract

A compound of formula (II) in which positional numbering, where mentioned, refers to the system illustrated above, and one or more W and one or more U are attached to a ring carbon or carbons at any of positions 1-11 or to a ring nitrogen at position 7 if present, and in which: W is C(=Q)NRCHJ(CH2)nR1 where Q is O or S, R is hydrogen or a C¿1-4? alkyl group which is optionally substituted with one or more OH or NH2 groups, J is H or a C1-C6 alkyl group optionally substituted with OH, OMe, NH2, NHMe or NMe2 functions, R?1¿ is C(=NR2)NH2, NHC(=NR3)NH2 or NR4R5, where each of R?2 and R3¿ are independently hydrogen or a C¿1-4? alkyl group which is optionally substituted with one or more OH or NH2 groups, and R?4 and R5¿ are independently hydrogen or a C¿1-4? alkyl group which is optionally substituted with one or more OH or NH2 groups, or R?4 and R5¿ together with the nitrogen atom to which they are attached form an optionally substituted saturated or unsaturated heterocyclic group; and n is an integer from 0 to 5; X, Y, V are independently CH or N; Z is CH¿2?, CH-C1-4 alkyl, CO, O, S, SO, SO2, N-C1-4 alkyl or NH; and U is H, halo, OH, CO2H, NR?6R7¿, nitro, cyano, C¿1-6? alkyl, C1-6 haloalkyl, C1-6 alkoxy, C1-6 haloalkoxy, C1-6 aminoalkyl or C1-6 aminoalkoxy in which R?6 and R7¿ have the same definitions as R?4 and R5¿ above; or a pharmaceutically-acceptable salt, N-oxide, hydrate, solvate, pharmaceutically acceptable derivative, pro-drug, tautomer or isomer thereof, with the proviso that W at position 4 cannot be CONH(CH¿2?)2NMe2, CONH(CH2)2NEt2, CONH(CH2)3NMe2, CONH(CH2)3NEt2, CONH(CH2)2Npiperidyl, or CONH(CH2)2Nmorpholinyl, when X and Y are N, V is CH, U is H and Z is CO. 'Bis' compounds and carboxylic acid precursors are also claimed. The compounds are useful in the treatment of neoplastic conditions such as cancer.

Description

ANTI-TUMOUR COMPOUNDS

This invention relates to compounds with topoisomerase-inhibitory activity, to pharmaceutical compositions comprising these compounds, and to the use of the compounds in the treatment of cancers .

BACKGROUND OF THE INVENTION

Despite the great advances in chemotherapy of cancer which have been made over the last decades, there are still a number of types of cancers for which the response rates are poor, and the agents which are available to treat these conditions have very significant toxic side-effects. In particular, solid tumours such as cancers of the colon, breast and lung are extremely common, and although each can be treated with presently- available cytotoxic agents, response rates are poor, relapse is common, and the five-year survival rate is poor. Since the initial discovery of the enzymes DNA topoisomerase I (topo I) and DNA topoisomerase II (topo II; DNA gyrase) , intensive effort has been directed at identifying inhibitors of these enzymes and evaluating their activity as potential anti-cancer agents. In particular, inhibitors of topo II have been viewed as attractive targets for drug development. Because DNA topoisomerases are essential for many aspects of cell multiplication, they are potentially very useful as anti- tumour agents . Compounds which have the ability to intercalate into the DNA double helix represent a major class of' inhibitors of topoisomerases. These agents include synthetic intercalating drugs, such as the aminoacridines, antibiotics such as anthracylines, including doxorubicin, and plant-derived agents such as the ellipticines and camptothecins . Following the clinical success of the DNA- intercalating topo II inhibitors doxorubicin, mitoxantrone and their analogues as anticancer drugs, a great deal of work has been devoted towards other classes of compounds with similar overall topology, eg. polycyclic chromophores bearing a flexible cationic side chain, as topo II inhibitors. The benzoisoquinolinediones amonafide (1) (Asbury et al . , 1998; Leaf et al . , 1997) and the anthrapyrazoles, such as losoxantrone (2) (Diab et al . , 1999; Judson, 1992), have undergone extensive clinical trials, while other compounds such as the phenazine-1- carboxamides ( eg. 3) (Rewcastle et al , 1987) have been studied in animal models .

More recently, interest has focused on compounds with the ability to inhibit both topo I and topo II enzymes. Examples of such "mixed" inhibitors which show broad- spectrum activity against solid tumours and are in clinical trial include N- [2- (dimethylamino) ethyl] cridine-4- carboxamide (DACA) (4) (Atwell et al , 1987; Baguley et al , 1995; U.S. Patent No. 4,590,277 and International Patent Application No. WO93/24096) , and various tetracyclic chromophores, including Tas-103 (5) (Utsugi et al, 1996) .

While the majority of DNA-intercalating anticancer drugs possess linear polycyclic chromophores, an increasing number of examples of "fused" tetracyclic systems have been reported, including the azonafides (e.g, 6) (Sami et al, 1993, 1996), imidazoacridinones (e.g., 7) (Cholody et al 1990, 1996), pyrimido [5, 6, 1-de] acridines (e.g., 8) (Antonini et al, 1995) and benzo [e]pyrimidines (e.g., 9) (Stefanska et al , 1993). Many of these compounds have been shown to be potent topoisomerase inhibitors and cytotoxins . For example, the azonafide analogue 7 was on average 40-fold more cytotoxic in a panel of human tumour cell lines than a related tricyclic analogue, the clinical drug amonafide (1) (Sami et al , 1993). Formulae of these compounds are presented in Figure 1. In the examples of "fused" compounds 6-9 above, the basic side-chain is linked to the ring system through an alkyl or amino function.

It is known that many topoisomerase inhibitors are bis compounds, such as jis-imidazoacridones and bis- triazenoacridones, and the compounds DMP840 and LU79553 are in Phase II clinical trials.

From the publications referred to above, it can be seen that compounds which have activity against topoisomerases show a wide variety of structures . Although there are some families of compounds, within a given family a wide variety of substituents on the central ring structure(s) is also possible.

In particular, the attachment of a basic side-chain to a ring system by way of a carboxamide link has provided compounds of current interest. The acridine-based DACA (4) referred to above, and tetracyclic 11-oxo-llH-indeno [1, 2- b] quinoline-6-carboxamides, illustrated by 10 (Deady et al 1997), which also has excellent activity against model tumour systems in vi tro and in vivo, are examples of "linear" systems, while 11 has a carboxamide linked side- chain on a "fused" ring system (Antonini et al 1992).

Because the relationships between structure and the ability to inhibit topoisomerases are still not sufficiently defined to enable predictions to be made about activity, and because of the potential utility of such compounds, further classes of such agents are needed. It will be clearly understood that, although a number of prior art publications are referred to herein, this reference does not constitute an admission that any of these documents forms part of the common general knowledge in the art, in Australia or in any other country.

SUMMARY OF THE INVENTION

We have now synthesised and evaluated a novel series of carboxamide linked compounds based on systems formed from fusion of four six-membered aromatic rings in the pattern shown in formula I.

Representative examples of these compounds show good activity against model tumour systems .

According to a first aspect, the invention provides a compound of formula II

II

in which positional numbering, where mentioned, refers to the system illustrated above, and one or more W and one or more U are attached to a ring carbon or carbons at any of positions 1-11 or to a ring nitrogen at position 7 if present, and in which:

W is C(=Q)NRCHJ(CH₂)_nR¹ where Q is 0 or S,

R is hydrogen or a C_]__ alkyl group which is optionally substituted with one or more OH or NH₂ groups,

J is H or a Ci-Cg alkyl group optionally substituted with OH, OMe, NH2, NHMe or NMe2 functions,

R¹ is C(=NR²)NH₂, NHC(=NR³)NH₂ or NR⁴R⁵, where each of R² and R³ are independently hydrogen or a C₁_₄ alkyl group which is optionally substituted with one or more OH or NH₂ groups, and R⁴ and R⁵ are independently hydrogen or a C^_^ alkyl group which is optionally substituted with one or more OH or NH₂ groups, or R⁴ and R⁵ together with the nitrogen atom to which they are attached form an optionally substituted saturated or unsaturated heterocyclic group; and n is an integer from 0 to 5;

X, Y, V are independently CH or N; Z is CH₂, CH-C₁_₄ alkyl, CO, 0, S, SO, S0₂, N-C₁_₄ alkyl or NH; and U is H, halo, OH, C0₂H,

, nitro, cyano, C _₆ alkyl, C_]__ haloalkyl, C^g alkoxy, C^-s haloalkoxy Cι_6 aminoalkyl or C-___Q aminoalkoxy in which R^δ and R⁷ have the same definitions as R⁴ and R⁵ above; or a pharmaceutically-acceptable salt, N-oxide, hydrate, solvate, pharmaceutically acceptable derivative, pro-drug, tautomer or isomer thereof, with the proviso that W at position 4 cannot be CONH(CH2)2 Me2, CONH (CH2) 2NEt2, CONH (CH2 ) 3NMe2 , CONH(CH2)3NEt2, CONH (CH2) 2Npiperidyl , or

CONH(CH₂)₂Nmorpholinyl, when X and Y are N, V is CH, -U is H. and Z is CO.

The term "alkyl" used either alone or in a compound word such as "optionally substituted alkyl" or "optionally substituted cycloalkyl" denotes straight chain, branched or mono- or poly- cyclic alkyl, alkyl or cycloalkyl . Examples of straight chain and branched alkyl include methyl, ethyl, propyl, isopropyl, butyl, isbutyl, sec-butyl, tert-butyl, amyl, isoamyl, sec-amyl, 1,2- dimethylpropyl , 1, 1-dimethylpropyl, hexyl, 4-methylpentyl, 1-methylpentyl, 2-methylpentyl, 3-methylpentyl, 1,1- dimethylbutyl, 2 , 2-dimethylbutyl, 3 , 3-dimethylbutyl, 1,2- dimethylbutyl, 1, 3-dimethylbutyl, 1, 2 , 2-trimethylpropyl, 1, 1, 2-trimethylpropyl, heptyl, 5-methylhexyl, 1- methylhexyl, 2 , 2-dimethylpentyl, 3 , 3-dimethylpentyl, 4,4- dimetylpentyl , 1, 2-dimethylpentyl, 1,3-dimethylpentyl, 1, 4-dimethylpentyl, 1, 2 , 3-trimethylbutyl, 1,1,2- trimethylbutyl, nonyl, 1-, 2-, 3-, 4-, 5-, 6- or 7- methyloctyl, 1-, 2-, 3-, 4- or 5-ethylheptyl, 1-2- or 3- propylhexyl, decyl, 1- , 2 - , 3-, 4-, 5-, 6-, 7- and 8- methylnonyl, 1-, 2-, 3-, 4-, 5- or 6-ethyloctyl, 1-, 2-, 3- or 4-propylheptyl, undecyl 1-, 2-, 3-, 4-, 5-, 6-, 7-, 8- or 9-methyldecyl, 1-, 2-, 3-, 4-, 5-, 6- or 7- ethylnonyl, 1-, 2-, 3-, 4- or 5-propyloctyl, 1-, 2- or 3- butylheptyl, 1-penylhexyl, dodecyl, 1-, 2-, 3-, 4-, 5-, 6-, 7-, 8-, 9- or 10-methylundecyl, 1-, 2-, 3-, 4-, 5-, 6- , 7- or 8-ethyldecyl, 1-, 2-, 3-, 4-, 5- or 6-propylnonyl, 1-, 2-, 3- or 4-butyloctyl, 1-2-pentylheptyl and the like. Examples of cyclic alkyl include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl and cyclodecyl and the like. The alkyl may optionally be substituted by any non-deleterious substituent .

"Me" refers to methyl.

The term "heterocyclic group" used either alone or in compound words such as "optionally substituted saturated or unsaturated heterocyclic group" denotes monocyclic or polycyclic heterocyclic groups containing at least one heteroatom atom selected from nitrogen, sulphur and oxygen. Suitable heterocyclic groups include N- containing heterocyclic groups, such as, unsaturated 3 to 6 membered heteromonocyclic groups containing 1 to 4. nitrogen atoms, for example, pyrrolyl, pyrrolinyl, imidazolyl, pyrazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, triazolyl or tetrazolyl; saturated 3 to 6-membered heteromonocyclic groups containing 1 to 4 nitrogen atoms, such as, pyrrolidinyl, imidazolidinyl, piperidino or piperazinyl; unsaturated condensed heterocyclic groups containing 1 to 5 nitrogen atoms, such as indolyl, isoindolyl, indolizinyl, benzimidazolyl, quinolyl, isoquinolyl, indazolyl, benzotriazolyl or tetrazolopyridazinyl; unsaturated 3 to 6-membered heteromonocyclic group containing an oxygen atom, such as, pyranyl or furyl; unsaturated 3 to 6-membered heteromonocyclic group containing 1 to 2 sulphur atoms, such as, thienyl; unsaturated 3 to 6-membered heteromonocyclic group containing 1 to 2 oxygen atoms and 1 to 3 nitrogen atoms, such as, oxazolyl, isoxazolyl or oxadiazolyl; saturated 3 to 6-membered heteromonocyclic group containing 1 to 2 oxygen atoms and 1 to 3 nitrogen atoms, such as, morpholinyl; unsaturated condensed heterocyclic group containing 1 to 2 oxygen atoms and 1 to 3 nitrogen atoms, such as, benzoxazolyl or benzoxadiazolyl; unsaturated 3 to 6-membered heteromonocyclic group containing 1 to 2 sulphur atoms and 1 to 3 nitrogen atoms, such as, thiazolyl or thiadiazolyl; saturated 3 to 6-membered heteromonocyclic group containing 1 to 2 sulphur atoms and 1 to 3 nitrogen atoms, such as, thiazolidinyl; and unsaturated condensed heterocyclic group containing 1 to 2 sulphur atoms and 1 to 3 nitrogen atoms, such as, benzothiazolyl or benzothiadiazolyl .

In this specification "optionally substituted" means that a group may or may not be further substituted with one or more groups selected from alkyl, alkenyl, alkynyl, aryl, halo, haloalkyl, haloalkenyl, haloalkynyl, haloaryl, hydroxy, alkoxy, alkenyloxy, aryloxy, benzyloxy, haloalkoxy, haloalkenyloxy, haloaryloxy, nitro, nitroalkyl, nitroalkenyl, nitroalkynyl, nitroaryl, nitroheterocyclyl, amino, alkylamino, dialkylamino, alkenylamino, alkynylamino, arylamino, diarylamino, benzylamino, dibenzylamino, acyl, alkenylacyl, alkynylacyl, arylacyl, acylamino, diacylamino, acyloxy, alkylsulphonyloxy, arylsulphenyloxy, heterocyclyl, heterocycloxy, heterocyclamino, haloheterocyclyl, alkylsulphenyl, arylsulphenyl, carboalkoxy, carboaryloxy, mercapto, alkylthio, benzylthio, acylthio, phosphorus- containing groups and the like. In some instances in this specification, where substitutents may be present, preferred substitutents have been mentioned. The term "halo" or "halogen" denotes fluorine, chlorine, bromine, or iodine. Where halogen substitution is present, preferred halogens are chlorine or bromine.

Preferably X is N; Z is CO, CH₂, 0 or S; V and Y are CH, N or C-CONH (CH₂) ₂N(CH₃ ) ₂ ; U is H, Cl, methyl or methoxy; and n is 2.

In a preferred embodiment, X is N, V and Y are CH, U is 2 -methyl, and Z is CO. In a particularly preferred embodiment the compound is N- [2- (dimethylamino) ethyl] -2 -methyl-7-oxo-7H- dibenz [ £, ij ] isoquinoline-11-carboxamide or a pharmaceutically acceptable salt or N-oxide thereof.

The present invention also provides a compound of the formula III :

(III)

in which: one or more W' and U' may be present at any of positions 1-11 of the structure;

W' has the same definition as W in formula II, or W is H;

U', V, X¹, Y¹ and Z¹ have the same definitions as U, V, X, Y and Z in formula II;

L is a linker group of valency x,

L' is a direct bond or a linker group attached to the ring system at one or two positions; and x is an integer from 2 to 4, or a pharmaceutically-acceptable salt, N-oxide, hydrate, solvate, pharmaceutically acceptable derivative, pro-drug, tautomer or isomer thereof.

For the avoidance of any doubt, it is to be understood that each bracketed portion of the compound may be different, provided that in each bracketed portion the substituents are individually within the definitions provided. In other words, compounds with two or more different units of formula II linked together are to be considered to be within the definition of formula III above .

The term "linker group" is used herein in its broadest sense to refer to any organic group that links together the adjacent units of the compound together. The linker groups L' in each repeated unit of the compound may be the same or different. Accordingly, the linker may be symmetrical or non-symmetrical.

L¹ is preferably -C ( =Q ' ) NR⁸CHJ ' (CH₂ ) _m- , in which: Q¹ has the same definition as Q in formula II above ;

R⁸ has the same definition as R in formula II above, or R⁸ is -C(=0)- or -C(=S)- in which the carbon atom of this group is attached to the ring structure at a second carbon atom on the ring;

J¹ has the same definition as J in the compound of formula II above, and m has the same definition as n in the compound of formula II above. L may be any suitable linking group that links the bracketed portions of the compound together.

Preferably L is a nitrogen-containing linker group. For example, L may be N (x=3) or -NR⁹-[ (CHR¹⁰)oNRⁿ]_p- wherein: p is 0 or 1; o is an integer from 0 to 5; R⁹ and R¹¹ have the same definition as R in formula II above, or when p is 1, R⁹ and R' ' may together form an optionally substituted branched or straight chained alkylene; and R¹⁰ has the same definition as J in formula II above .

Preferably R⁹ and R¹¹ are each independently H, CH₃, or C₂H₅, or if p is 1, R⁹ and R¹¹ is preferably -CH₂CH₂-

However, L could alternatively be 0, S, an optionally substituted C_.-₂o alkylene, alkenylene or alkynylene chain, which may optionally be interspersed with one or more aryl or heterocyclic groups (which may also be substituted) and/or one or more 0, S or N atoms; or L may be an optionally substituted saturated or unsaturated aryl or heterocyclic group.

The terms "alkylene", "alkenylene" and "alkynylene" are the divalent radical equivalents of the terms "alkyl", "alkenyl" and "alkynyl", respectively. The two bonds connecting the alkylene, alkenylene or alkynylene to the adjacent groups may come from the same carbon atom or different carbon atoms in the divalent radical .

Preferred optional substituents in the linker group L are selected from halogen; oxy; hydroxy; alkoxy; alkylthio; cyano; azido; acyloxy; alkyl sulphonyl ; aryl and heteroaryl .

It will be appreciated by persons skilled in the art of the invention that the compounds of formula III include bis compound forms of the compound of formula II in which W is replaced in each subunit of the bis compound by a linker group. The linker group of the bis compounds preferably has a structure that is based on that of W. The bis form of the compounds within the scope of formula III therefore include compounds having two units of formula II in which the NRCHJ (CH₂ ) _nR^α portion of W in each unit of formula II is replaced with a group selected from the following:

-NH (CH₂ ) ₂NH (CH₂ ) ₂NH-

-NH (CH₂ ) ₃-NMe- (CH₂ ) ₃NH- -NH(CH₂)₂NH(CH₂)₂NH(CH₂)₂NH-

-NH (CH₂ ) ₂NH (CH₂ ) ₃NH (CH₂ ) ₂NH-

-NH (CH₂ ) ₂NMe (CH₂ ) ₂NMe (CH₂ ) ₂NH-

-NH (CH₂ ) ₂NMe (CH₂ ) ₃NMe (CH₂ ) ₂NH-

-N, N' -Bis (2-aminoethyl ) piperazine- -N,N' -Bis (3-aminopropyl) iperazine-,

-NH (CH₂ ) _mNH (CH₂ ) _pNH-

-NH (CH₂) _mNAlkyl (CH₂ ) _pNH-

-NH (CH₂ ) _mNH (CH₂ ) _pNH (CH₂ ) _rNH- , and

-NH (CH₂ ) _mNAlkyl (CH₂ ) _pNAlkyl (CH₂ ) _rNH- , where m, p and r are integers from 2 to 6.

The linker may alternatively be of the type disclosed in International Patent Application No. WO 96/25400 by The Du Pont Merck Pharmaceutical Company, the entire disclosure of which is incorporated by this cross- reference. The linkers disclosed in this application include two sites of attachment of the linker L' to the ring structure, each via -C(=Q)-. Formula III includes such compounds within its scope when R⁸ is -C(=0)- or - C(=S)- as described above. In a second aspect, the invention provides a pharmaceutical composition comprising a compound of formula II or III as described above, together with a pharmaceutically-acceptable carrier .

In a third aspect, the invention provides a method of treatment of a neoplastic condition, comprising the step of administering a therapeutically effective dose of a compound of formula II or formula III or a pharmaceutically acceptable derivative thereof, pro-drug thereof, tautomer and/or isomer thereof to a subject in need of such treatment.

The term "therapeutically-effective amount" means an amount of a chemotherapeutic agent to yield a desired therapeutic response, for example, treat or prevent a neoplastic disease.

The specific "therapeutically-effective amount" will, obviously, vary with such factors as the particular condition being treated, the physical condition of the subject, the type of animal being treated, the duration of the treatment, the nature of concurrent therapy (if any) , and the specific formulations employed and the structure of the che otherapeutic agent or its derivatives . The compound of the invention may be administered in conjunction with one or more other anti-neoplastic agents, including but not limited to anti-mitotic agents such as taxol, anti-metabolites such as 5-fluorouracil, hormonal regulators such .as tamoxifen, DNA-reactive agents such as cisplatin, or biological agents such as interleukin- 2 (IL-2) or antibodies. The compound of the invention and the second agent may be administered together or sequentially.

A second DNA-binding anti-cancer therapeutic agent could be used in conjunction with administration of the compound of formula II or formula III in order to reduce toxicity to the recipient of either or both of the compound of formula II or formula III or the other anticancer agent. The compound formula II or formula III and the other agent may be administered together or sequentially.

It is contemplated that compounds of the invention may also be administered in the form of tumour- activated prodrugs, in which the active agent is linked to a 'trigger' domain; such compounds may for example be designed to be activated by local hypoxia within a tumour mass. Suitable methods are known the in the art; see for example Denny, 1996; McFadyen et al , 1996.

The compound of the invention may also be used in combination with agents which relieve side effects caused by drug treatment such as granulocyte-macrophage-colony stimulating factor (GM-CSF), or anti-emetics. Optionally the compound of the invention is administered in a divided dose schedule, such that there are at least two administrations in total in the schedule. Administrations are given preferably at least every two hours for up to four hours or longer; for example the compound may be administered every hour or every half hour. In one preferred embodiment, the divided-dose regimen comprises a second administration of the compound of the invention after an interval from the first administration sufficiently long that the level of active compound in the blood has decreased to approximately from 5-30% of the maximum plasma level reached after the first administration, so as to maintain an effective content of active agent in the blood. Optionally one or more subsequent administrations may be given at a corresponding interval from each preceding administration, preferably when the plasma level has decreased to approximately from 10-50% of the immediately-preceding maximum.

The compounds of the invention may be administered by any suitable route, for example orally, buccally, topically or parenterally, for example by intravenous, sub-cutaneous, intramuscular, intra- peritoneal, or intratumoral injection. The dose and route of administration will depend on the condition to be treated, and will be at the discretion of the attending physician or veterinarian. It is contemplated that each administration will supply between 0.1 and 500 mg, preferably 1 to 200, more preferably 1 to 50 mg of active compound . The compounds of the invention are suitably presented in unit dosage form.

Methods and pharmaceutical carriers for preparation of pharmaceutical compositions are well known in the art, as set out in textbooks such as Remington's Pharmaceutical Sciences, 17th Edition, Mack Publishing Company, Easton, Pennsylvania, USA. Further information on formulations is also provided in the more detailed description of the invention set out below.

The term "subject" as used herein refers to any animal having a disease or condition which requires treatment with an anti-neoplastic agent. Preferably the subject is suffering from a cellular proliferative disorder (eg., a neoplastic disorder). Subjects for the purposes of the invention include, but are not limited to, mammals (eg., bovine, canine, equine, feline, porcine) and preferably humans .

By "cell proliferative disorder" is meant that a cell or cells demonstrate abnormal growth, typically aberrant growth, leading to a neoplasm, tumor or a cancer. Cell proliferative disorders include, for example, cancers of the breast, lung, prostate, kidney, skin, neural, ovary, uterus, liver, pancreas, epithelial, gastric, intestinal, exocrine, endocrine, lymphatic, haematopoietic system or head and neck tissue. Generally, neoplastic diseases are conditions in which abnormal proliferation of cells results in a mass of tissue called a neoplasm or tumor. Neoplasms have varying degrees of abnormalities in structure and behaviour. Some neoplasms are benign while others are malignant or cancerous. An effective treatment of neoplastic disease would be considered a valuable contribution to the search for cancer preventive or curative procedures . The compounds of the invention are preferably used in the treatment of leukaemias, ly phomas, sarcomas, and brain tumours, and for cancers of the lung, breast, ovary, testes, and colon.

In a fourth aspect, the present invention provides for the use of a compound of formula II or III in the manufacture of a medicament for the treatment or prophylaxis of a neoplastic condition.

It will be clearly understood that methods of synthesis of compounds of the invention also form part of the invention. Certain intermediate compounds described herein are new, in particular precursor acids to the amides of the invention, and also form part of the invention. The present invention therefore also provides compounds of formula IV:

(IV)

in which: the compound includes one or more U' ' groups and one or more carboxylic acid groups are attached to a ring carbon or carbons or nitrogen when present in the ring structure at positions 1 - 11; and U' ' , V ' , X' ' , Y' ' and Z'' have the same definitions as U, V, X, Y and Z, respectively, in formula II.

According to the present invention, there is also provided a method of synthesising a compound of formula II or formula III, comprising the step of converting the carboxylic acid of formula IV into the target amide of formula II or formula III.

The conversion may involve an intermediate step in which the compound of formula IV is converted into an imidazolide, and reacting the imidazolide intermediate with the appropriate amine to obtain the target amide of formula II or formula III. Alternatively, the reaction involves converting the carboxylic acid of formula IV into an acid halide, and reacting the acid halide with an amine to obtain the target amide of formula II or formula III . The reagent in this second route is preferably thionyl chloride. It will be clearly understood in the above description that in the case of the bis compounds, two units of the carboxylic acid will be reacted with the appropriate diamine to form the target diamide.

For the purposes of this specification it will be clearly understood that the word "comprising" means "including but not limited to", and that the word "comprises" has a corresponding meaning.

BRIEF DESCRIPTION OF THE DRAWINGS:

Figure 1 shows the structures of prior art compounds 1 to 11 referred to herein.

Figure 2 shows reaction schemes for the preparation of aminoanthraquinone precursors of the tetracyclic systems.

Scheme 2a — l-aminoanthraquinone-2 -carboxylic acid;

Scheme 2b — 4-aminoanthraquinone-l-carboxylic acid; Scheme 2c — 5- and 8-aminoanthraquinone-l- carboxylic acids;

Scheme 2d — 5- and 8-chloro-l-aminoanthraquinone and an alternative route to 5- and 8-aminoanthraquinone-l- carboxylic acids. Figure 3 shows reaction schemes leading to tetracyclic compounds .

Scheme 3a — Route to the precursor 2-methyl-7H- benzo [ e] perimidin-7 -one

Scheme 3b — General route from aminoanthraquinones

Scheme 3c — Route to carbocyclic example

Scheme 3d — Route to diaza example

Figure 4 shows the structure of the imidazolide intermediate from acid 21a. Figure 5 shows a comparison between the growth of transplanted tumour in the colon 38 in control mice (•) and those receiving a single dose of 24h (65 mg/kg; O) . Figure 6 shows a comparison between the growth of transplanted tumour in the colon 38 in control mice (•) and those receiving multiple doses of 24f (150 mg/kg q4dx3 ; O) . Figure 7 shows a comparison between the growth of transplanted tumour in the colon 38 in control mice (•) and those receiving multiple doses of 24i (65 mg/kg q4dx3 ; ■) .

DETAILED DESCRIPTION OF THE INVENTION

Before the present compounds, compositions, and methods are described, it is understood that this invention is not limited to the particular materials and methods described, as these may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the present invention which will be limited only by the appended claims . It must be noted that as used herein and in the appended claims, the singular forms "a," "an," and "the" include plural reference unless the context clearly dictates otherwise. Thus, for example, a reference to "a compound" includes compounds of similar formula and equivalents thereof known to those skilled in the art, and so forth. Unless defined otherwise, all technical and scientific terms used herein have the same meanings as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any materials and methods similar or equivalent to those described herein can be used to practice or test the present invention, the preferred materials and methods are now described.

The description that follows makes use of a number of terms used in pharmaceutical chemistry and cell biology. In order to provide a clear and consistent understanding of the specification and claims, including the scope given such terms, the following definitions are provided. The salts of the compound of Formula II are preferably pharmaceutically acceptable, but it will be appreciated that non-pharmaceutically acceptable salts also fall within the scope of the present invention, since these are useful as intermediates in the preparation of pharmaceutically acceptable salts. Examples of pharmaceutically acceptable salts include salts of pharmaceutically acceptable cations such as sodium, potassium, lithium, calcium, magnesium, ammonium and alkylammonium; acid addition salts of pharmaceutically acceptable inorganic acids such as hydrochloric, orthophosphoric, sulphuric, phosphoric, nitric, carbonic, boric, sulfamic and hydrobromic acids; or salts of pharmaceutically acceptable organic acids such as acetic, propionic, butyric, tartaric, maleic, hydroxymaleic, fumaric, citric, lactic, mucic, gluconic, benzoic, succinic, oxalic, phenylacetic, methanesulphonic, trihalomethanesulphonic, toluenesulphonic, benzenesulphonic, salicyclic, sulphanilic, aspartic, glutamic, edetic, stearic, palmitic, oleic, lauric, pantothenic, tannic, ascorbic and valeric acids.

By "pharmaceutically acceptable derivative" is meant any pharmaceutically acceptable salt, hydrate or any other compound which, upon administration to the subject, is capable of providing (directly or indirectly) a compound of Formula II or an antivirally active metabolite or residue thereof.

The term "pro-drug" is used herein in its broadest sense to include those compounds which are converted in vivo to compounds of Formula II.

The term "tautomer" is used herein in its broadest sense to include compounds of Formula II which are capable of existing in a state of quilibrium between two isomeric forms. Such compounds may differ in the bond connecting two atoms or groups and the position of these atoms or groups in the compound. For example, it will be clearly understood that when U is 2-OH and Y and/or X are N, the invention includes within its scope the keto tautomeric form where V is CO and Y or X is NH.

The term "isomer" is used herein in its broadest sense and includes structural, geometric and stereo isomers . As the compound of Formula II may have one or more chiral centres, it is capable of existing in enantiomeric forms .

The term "toxic side effects" or "side effects" means the deleterious, unwanted effects of chemotherapy on the subject's normal, non-diseased tissues and organs. For example, toxic side effects may include bone marrow suppression (including neutropenia) , cardiac toxicity, hair loss, gastrointestinal toxicity (including nausea and vomiting), neurotoxicity, lung toxicity and asthma. The aldehyde-releasing compound and/or chemotherapeutic agents may be administered orally, topically, or parenterally in dosage unit formulations containing conventional non-toxic pharmaceutically acceptable carriers, adjuvants, and vehicles. The term parenteral as used herein includes subcutaneous injections, aerosol, intravenous, intramuscular, intrathecal, intracranial, injection or infusion techniques.

The present invention also provides suitable topical, oral, and parenteral pharmaceutical formulations for use in the novel methods of treatment of the present invention. The compounds of the present invention may be administered orally as tablets, aqueous or oily suspensions, lozenges, troches, powders, granules, emulsions, capsules, syrups or elixirs. The composition for oral use may contain one or more agents selected from the group of sweetening agents, flavouring agents, colouring agents and preserving agents in order to produce pharmaceutically elegant and palatable preparations . The tablets contain the active ingredient in admixture with non-toxic pharmaceutically acceptable excipients which are suitable for the manufacture of tablets . These excipients may be, for example, (1) inert diluents, such as calcium carbonate, lactose, calcium phosphate or sodium phosphate; (2) granulating and disintegrating agents, such as corn starch or alginic acid; (3) binding agents, such as starch, gelatin or acacia; and (4) lubricating agents, such as magnesium stearate, stearic acid or talc. These tablets may be uncoated or coated by known techniques to delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period. For example, a time delay material such as glyceryl monostearate or glyceryl distearate may be employed. Coating may also be performed using techniques described in the U.S. Pat. Nos. 4,256,108; 4,160,452; and 4,265,874 to form osmotic therapeutic tablets for control release .

The compounds and compositions useful in the methods of the invention can be administered, for in vivo application, parenterally by injection or by gradual perfusion over time independently or together.

Administration may be intravenously, intra-arterial, intraperitoneally, intramuscularly, subcutaneously, intracavity, or transdermally. For in vitro studies the agents may be added or dissolved in an appropriate biologically acceptable buffer and added to a cell or tissue .

Preparations for parenteral administration include sterile aqueous or non-aqueous solutions, suspensions, and emulsions. Examples of non-aqueous solvents are propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable organic esters such as ethyl oleate. Aqueous carriers include water, alcoholic/aqueous solutions, emulsions or suspensions, including saline and buffered media. Parenteral vehicles include sodium chloride solution,

Ringer's dextrose, dextrose and sodium chloride, lactated Ringer's intravenous vehicles include fluid and nutrient replenishers, electrolyte replenishers (such as those based on Ringer's dextrose), and the like. Preservatives and other additives may also be present such as, for example, anti-microbials, anti-oxidants, chelating agents, growth factors and inert gases and the like.

Generally, the terms "treating", "treatment" and the like are used herein to mean affecting a subject, tissue or cell to obtain a desired pharmacologic and/or physiologic effect. The effect may be prophylactic in terms of completely or partially preventing a disease or sign or symptom thereof, and/or may be therapeutic in terms of a partial or complete cure of a disease. "Treating" as used herein covers any treatment of, or prevention of a disease in a vertebrate, a mammal, particularly a human, and includes: (a) preventing the disease from occurring in a subject that may be predisposed to the disease, but has not yet been diagnosed as having it; (b) inhibiting the disease, ie., arresting its development; or (c) relieving or ameliorating the effects, ie., cause regression of the effects of the disease.

The invention includes various pharmaceutical compositions useful for treating a disease. The pharmaceutical compositions according to one embodiment of the invention are prepared by one or more compounds according to the invention into a form suitable for administration to a subject using carriers, excipients and additives or auxiliaries . Frequently used carriers or auxiliaries include magnesium carbonate, titanium dioxide, lactose, mannitol and other sugars, talc, milk protein, gelatin, starch, vitamins, cellulose and its derivatives, animal and vegetable oils, polyethylene glycols and solvents, such as sterile water, alcohols, glycerol and polyhydric alcohols. Intravenous vehicles include fluid and nutrient replenishers . Preservatives include antimicrobial, anti-oxidants, chelating agents and inert gases. Other pharmaceutically acceptable carriers include aqueous solutions, non-toxic excipients, including salts, preservatives, buffers and the like, as described, for instance, in Remington's Pharmaceutical Sciences, 15th ed. Easton: Mack Publishing Co., 1405-1412,1461-1487 (1975) and The National Formulary XIV., 14th ed. Washington:

American Pharmaceutical Association (1975) , the contents of which are hereby incorporated by reference. The pH and exact concentration of the various components of the pharmaceutical composition are adjusted according to routine skills in the art. See Goodman and Gilman's The Pharmacological Basis for Therapeutics (7th ed.).

The pharmaceutical compositions are preferably prepared and administered in dose units. Solid dose units are tablets, capsules and suppositories. For treatment of a subject, depending on activity of the chemotherapeutic agent, manner of administration, nature and severity of the disorder, age and body weight of the subject, different daily doses can be used. Under certain circumstances, however, higher or lower daily doses may be appropriate. The administration of the daily dose can be carried out both by single administration in the form of an individual dose unit or else several smaller dose units and also by multiple administration of subdivided doses at specific intervals. The pharmaceutical compositions according to the invention may be administered locally or systemically in a therapeutically effective dose. Amounts effective for this use will, of course, depend on the severity of the disease and the weight and general state of the subject. Typically, dosages used in vitro may provide useful guidance in the amounts useful for in situ administration of the pharmaceutical composition, and animal models may be used to determine effective dosages for treatment of particular disease. Various considerations are described, eg., in Langer, Science, 249: 1527, (1990). Formulations for oral use may be in the form of hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate or kaolin. They may also be in the form of soft gelatin capsules wherein the active ingredient is mixed with water or an oil medium, such as peanut oil, liquid paraffin or olive oil.

Aqueous suspensions normally contain the active materials in admixture with excipients suitable for the manufacture of aqueous suspension. Such excipients may be (1) suspending agent such as sodium carboxymethyl cellulose, methyl cellulose, hydroxypropylmethylcellulose, sodium alginate, polyvinylpyrrolidone, gum tragacanth and gum acacia; (2) dispersing or wetting agents which may be (a) naturally occurring phosphatide such as lecithin; (b) a condensation product of an alkylene oxide with a fatty acid, for example, polyoxyethylene stearate; (c) a condensation product of ethylene oxide with a long chain aliphatic alcohol, for example, heptadecaethylenoxycetanol; (d) a condensation product of ethylene oxide with a partial ester derived from a fatty acid and hexitol such as polyoxyethylene sorbitol monooleate, or (e) a condensation product of ethylene oxide with a partial ester derived from fatty acids and hexitol anhydrides, for example polyoxyethylene sorbitan monooleate. The pharmaceutical compositions may be in the form of a sterile injectable aqueous or oleagenous suspension. This suspension may be formulated according to known methods using those suitable dispersing or ^• wetting agents and suspending agents which have been mentioned above. The sterile injectable preparation may also a sterile injectable solution or suspension in a non- toxic parenterally-acceptable diluent or solvent, for example, as a solution in 1, 3-butanediol . Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution, and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose, any bland fixed oil may be employed including synthetic mono-or diglycerides . In addition, fatty acids such as oleic acid find use in the preparation of injectables.

In addition, some of the compounds of the instant invention may form solvates with water or common organic solvents. Such solvates are encompassed within the scope of the invention.

The compounds of the present invention may additionally be combined with other compounds to provide an operative combination.

The invention will now be described in detail by way of reference only to the following non-limiting examples, and to the drawings.

The structure of representative compounds of the invention is summarised in Table 1. It will be evident that forms A and B represent mono and bis forms, respectively, of compounds of general formula II

Table 1

A B Representative Compounds of the Invention

R¹ = (CH₂)₂NMe₂, R = ( CH₂ ) ₃NMe₂ , R = (CH₂) ₂N-ρiperazine, R⁴= (CH₂)₂NH(CH₂)₂OH, R⁵=CH ( S-Me) CH₂NMe₂, R⁶=CH(i?- Me)CH₂NMe₂. Note: R²-R^s do not relate to the corresponding references in formulae II and III

Synthesis of aminoanthraquinones

The tetracyclic systems of general formula II with X=N were constructed from precursor aminoanthraquinones, prepared by known methodology from available materials .

Nitration of 2-methylanthraquinone (Wood and Peters, 1962) was followed by oxidation of the methyl group and borohydride reduction of the nitro function (Bennett et al, 1982) to give l-aminoanthraquinone-2-carboxylic acid 12 (Scheme 2a) .

1-Methylanthraquinone (Fieser and Heymann, 1942) (Scholl and Donat, 1931) was nitrated with potassium nitrate in concentrated sulfuric acid at < 5 °C (Scheme 2b) . Oxidation with manganese dioxide in concentrated sulfuric acid, at a higher temperature and for a longer time than reported previously (Scholl and Donat, 1931) , gave the carboxylic acid 14b in 54% yield. Reduction with aqueous sodium sulfide then gave the required amine 14c. Anthraquinone-1-carboxylic acid (14a) [from benzanthrone (Perkin, 1920)] on nitration gave a mixture of 5-nitro (14d) and 8-nitro (14e) isomers (Scheme 2c) (Sakat et al , 1973). Recrystallization from ethanol gave the former in pure form but the more soluble 8-isomer when treated as reported (Sakat et al , 1973) still contained some of the 5-nitro compound and other impurities. Compound 14e had the lowest solubility in hot toluene, and a pure sample, though in low yield, was obtained as the insoluble -residue from a toluene extraction. Reduction of each isomer with aqueous sodium sulfide gave the amines 14f and 14g.

The potassium salt of anthraquinone-1-sulfonic acid was nitrated (Scheme 2d) and, in this case, the 5-nitro (15b) and 8-nitro (15c) isomers were separated by purification of their potassium salts (Ullmann and Kertesz, 1919) . Reaction of 15b with sodium chlorate in concentrated hydrochloric acid replaced the sulfonic group with chlorine (Ullmann and Kertesz, 1919) and then reduction with aqueous sodium sulfide produced the amine 16c (Maki and Nagai, 1930) . Compound 16d.was prepared in the same way from 15c. The chloro group in 16d was also substituted by reaction with copper (I) cyanide and the nitrile 16e was hydrolyzed to provide an alternative route to carboxylic acid 14g. Since the 8-nitro isomer 15σ was the major product from the nitration of 15a, this was the preferred route to 14g. Synthesis of the tetracyclic systems from aminoanthraquinones

The reaction of 1-aminoanthraquinone (17) with dimethylacetamide and phosphoryl chloride (Fabio et al , 1978) gave an intermediate amidine 18 which was cyclised by reaction with ammonium acetate in hot ethanol (Nabar et al , 1983 ) to 2 -methyl -7iϊ-benzo [ e] pyrimidin-7 -one ( 19g)

( Scheme 3a ) .

Reaction of all aminoanthraquinones with acetone in aqueous sodium hydroxide gave substituted 2-meth.yl-7.H- dibenz [ f , ij] isoquinolin-7-ones 19 (Scheme 3b).

Surprisingly, this reaction with 14σ was accompanied by decarboxylation and 19a was formed instead of the target

19c. In the case of 14g, the product 19d also contained a small amount of the 4-methyl analogue 19h. These were separated at the later carboxamide stage.

Reaction of ester 14h (from acid 14g) with dimethylformamide dimethylacetal gave the intermediate amidine 27, which cyclized to 28a when refluxed with ammonium acetate in ethanol. Mild acidic hydrolysis afforded acid 28b (Scheme 3d) .

Further precursors to the compounds of Table 1.

A carboxylic acid function in the 2-position of both aza and diaza systems was generated by a two-stage oxidation (Scheme 3b). Aldehydes 20a,e,f,g were prepared by selenium dioxide oxidation of the corresponding methyl compounds (I9a,e,f,g). These in turn were efficiently oxidised with sodium chlorite (Lindgren and Nilsson, 1973) (Kraus and Roth, 1980) to the corresponding acids 2ia,e,f,g. The 2,8-diacid 21c was also prepared, from selenium dioxide oxidation of 19σ. In this case, prolonged reaction gave 21c directly and the intermediate aldehyde was not isolated. The 'deaza' carbocylic system, illustrated by 23 , was prepared by a literature route from methyl 2-iodobenzoate and methyl 8-bromo-l-naphthoate (Rule et al , 1934) ( Rule and Smith, 1937) (Scheme 3c) .

Compounds of Table 1

A variety of basic side-chains was then attached to the appropriate precursor by an amide link, and the range of carboxylic acids synthesised allowed the side-chains to be attached at various positions around the chromophores. ' In most cases this was achieved by way of an intermediate imidazolide, formed by reaction of the appropriate acid with 1, 1 ' -carbonyldiimidazole (CDI), e.g. 22 (Figure 4) from acid 21a. The imidazolide was isolated and then' reacted under mild conditions with the appropriate amine to give the required amides. An example of a bis amide 26 formed by reaction with N, N'~ [bis (2-aminoethyl) ] -N, N'- dimethyl-1, 3-propanediamine was also prepared by this method. The CDI route was not successful for the diamide 24e, but reaction of diacid 21c with hot thionyl chloride did give a di (acid chloride) intermediate. This was not isolated, but was reacted directly with N, N- dimethylethylenediamine. The final diamide 24e also contained a chloro substituent; nuclear chlorination occurred during the thionyl chloride reaction. The 4- orientation of the chlorine followed from ^-"C HETCOR and HMBC experiments which allowed assignment of all H and C signals. The thionyl chloride route was also used for preparation of 25.

Experimental Methods MR spectra were recorded on Brϋker AM-300 (300.13 and 75.47 MHz for ^XH and ¹³C, respectively) and Brϋker DRX- 400 (400.13 and 100.62 MHz) spectrometers, in DMSO-d6 unless stated otherwise, and are referenced to Me4Si. ΝMR signals for aromatic atoms are assigned only for 19a and diamide 24e where ^C_H (from HETCOR) and ³J_CH (from HMBC) couplings allowed identification of all C and H signals. Proton counts for aromatic protons are given only for unresolved ultiplets; the other aromatic signals are for single protons (with ortho coupling constants of J^" = 6-8 Hz where appropriate) . In addition to the peaks listed, the carboxamides of Form A had a common pattern for the side chain: δ 2.4 (s, 6 H, N(CH₃)₂), 2.7 (t, <J = 6 Hz, 2 H, CHN) , 3.75 (q, J^" = 6 Hz, 2 H, NHCH₂) . Electrospray mass spectra were recorded on a VG Bio-Q triple quadruple mass spectrometer, with water/MeOH/AcOH (50:50:1) as the mobile phase. Microanalyses were performed at the Campbell Microanalytical Laboratory, University of Otago, New Zealand.

7-Oxo-7iϊ-benz [de] anthracene-11-carboxylic acid (23) was prepared from methyl 2-iodobenzoate and methyl 8- bromo-1-naphthoate as reported (Rule et al , 1934) ( Rule and Smith, 1937) (Scheme 3c), mp 272-273 "C.

Ji NMR δ 7.71 (t), 7.79 (t) , 7.89-7.96 (m, 2 H) , 8.27. (d) , 8.40 (d), 8.46-8.53 (m, 2 H) , 8.65 (d) .

Example 1 : Preparation of Anthraquinone precursors according to the Reactions of Figure 2 1 -Methyl -4 -ni troanthraquinone (13b) .

To a cold solution of 1-methylanthraquinone (13a) (Fieser and Hey ann, 1942) (Scholl and Donat, 1931) (2.22 g, 1 mmol) in concentrated H₂S0₄ (15 mL) was added finely ground KNO₃ (1.0 g) at 0-5 °C over 30 min. The resultant mixture was stirred at 4 °C overnight, then poured on to ice, and the precipitate which separated was collected by filtration, washed thoroughly with water and dried to give a grey solid (2.40 g, 90%), mp 252-254 °C. [lit. (Fain and Plakidin, 1961) mp 261.1-261.5 °C) .

4 -Ni troanthraquinone-l -carboxylic acid (14b)

Manganese dioxide (Fatiadi, 1976) (3.0 g) was added in portions over 15 min to a stirring mixture of 13b (1.58 g, 5.91 mmol) and concentrated H₂Sθ₄ (15 mL) . The reaction mixture was stirred at room temperature for 15 min then at 60 °C overnight. After being cooled, it was poured on to ice and sodium sulfite (2.0 g) was added to consume unreacted manganese dioxide. The solid which remained was filtered off, washed with water, and then thoroughly extracted with 5% ammonia solution. Some insoluble material was filtered off and the filtrate was acidified with concentrated HCl . The precipitate which formed was filtered off and dried to give the product as a pale-brown solid (0.97 g, 54%), mp 308 °C (dec.) (lit. (Fain and Plakidin, 1961) mp 310-311 °C (dec). 4 -Aminoanthraquinone-1 -carboxylic acid (14c)

A mixture of 14b (0.95 g, 3.2 mmol) and sodium sulfide (5.0 g) in water (50 L) was heated under reflux for lh, then cooled and carefully acidified with concentrated HCl . The precipitate which formed was filtered off, washed with water and dried to give the product as a brown solid (0.82 g, 96%), mp 239-241 °C (lit. (Fain and Plakidin, 1961) mp 241.1-241.5 °C. ^XH NMR δ 7.20 (d) , 7.43 (d) , 7.85-7.93 ( , 2 H) , 8.07 (d) , 8.19 (d) , 12.68 (br s, C0₂H) . 5-Ni tro- (1 d) and 8-ni tro- (14eJ anthraquinone-1 - carboxylic acids

Anthraquinone-1-carboxylic acid (Perkin, 1920) (10 g) was nitrated and the isomers separated according to a literature procedure (Sakat et al, 1973). The less soluble 14d (4.8 g) was obtained pure, while 14e (2.1 g) still contained some 5-isomer and other impurities. The latter was stirred with boiling toluene (70 mL) and filtered while hot; the insoluble material (0.8 g) was pure 14e, mp 233-236 °C (lit. (Sakat et al, 1973) mp 288-295 °C) . 5 -Aminoanthraquinone-1 -carboxylic acid (14f)

A mixture of 14d (1.50 g, 5.22 mmol), sodium sulfide (10 g) and water (50 L) was refluxed for 1 h, then cooled on ice and filtered. The filtrate was acidified with concentrated HCl, and the precipitate which separated was collected by filtration, washed with water and dried to give the amine as a brown solid (1.30 g, 96%), mp 222-223 °C. !H NMR δ 7 . 21 ( d) , 7 . 34 ( d) , 7 . 54 ( t ) , 7 . 74 ( d) , 7 . 92 ( t ) , 8 . 27 ( d) .

5 -Ni tro- (15b) and 8-ni tro- (15c) ant hraquinone-1 -sulfonic acids (Ullmann and Kertesz, .1919;

To a solution of potassium anthraquinone-1-sulfonate (30 g) in concentrated H₂SO₄ (160 mL) at 50 °C, was added dropwise concentrated HNO₃ (7.0 mL) , and the mixture was then heated at 95-100 °C for 1 h before being kept at 4 °C overnight. The crude 5-nitroanthraquinone-l-sulfonic acid was filtered off. The filtrate was poured onto ice, kept at 4 °C for 24 h, and the crude 8-nitroanthraquinone-l- sulfonic acid was then filtered off. Each acid was dissolved in water (lOO L), saturated KC1 solution was added and the potassium salts of the acids separated on standing and were filtered off (5-nitro, 11.0 g; 8-nitro, 22.5 g) . 5-Chloro-l -ni troanthraquinone (16a)

A solution of potassium 5-nitroanthraquinone-l- sulfonate (19.0 g) in water (500 mL) and concentrated HCl (70 L) was heated to reflux with stirring under nitrogen for 1.5 h. A solution of sodium chlorate (17.0 g) in water (100 mL) was added over 2 h, and the resulting mixture was further refluxed for 1 h. After being cooled to 4 °C and kept overnight, the solid was collected by filtration, washed with cold water, and dried to give 16a (11.60 g, 79%), mp 310-312 °C (lit. (Maki and Nagai, 1930) mp 315 °C) .

^XH NMR δ 7.86 (t), 7.98 (d) , 8.07-8.14 (m, 2 H) , 8.19 (d) , 8.37 (d) .

8-chloro-l -ni troanthraquinone (16b) .

This was prepared as for 16a, in 85% yield, mp 199- 200 °C (lit. (Ullmann and Kertesz, .1919) mp 263 °C) . ^XH NMR δ 7 . 89 (t ) , 7 . 98 ( d) , 8 . 06 ( t , 8 . 18-8 .23 (m, 2 H) , 8 . 37 ( d) .

5-Chloro-l -aminoanthraquinone (16c) A mixture of 16a (5.75 g, 20 mmol), sodium sulfide

(24 g) and water (180 mL) was refluxed with stirring for 1 h. After being cooled to room temperature, 2% NaOH solution (180 mL) was added and the mixture was heated at 50 °C for 1 h. The precipitate was then filtered off, washed thoroughly with warm water and dried to give the product as a brick-red solid (4.6 g, 89%), mp 215-216 °C (lit. (Maki and Nagai, 1930) mp 219 °C) .

Ji NMR (CDCI₃) δ 6.79 (br s, 2 H, NH ) , 6.93 (d) , 7.46 (t) , 7.57-7.67 (m, 2 H) , 7.70 (d) , 8.28 (d) .

8 -Chi oro -1 -aminoan thraquinone (16d)

This was prepared as for 16c, in 88% yield, as a red solid, mp 227-228 °C (lit. (Wor ser et al, 1993) mp 230-232 °C. ^XH NMR (CDCI3) δ 6.78 (br s, NH₂ ) , 6.96 (d) , 7.43 (t) ,

7.56-7.62 ( , 2 H) , 7.75 (d) , 8.24 (d) .

8-Aminoanthraquinone-l -carboni trile (16e)

Copper (I) cyanide (1.8 g, 20 mmol) was added in one portion to a warm solution of 16d (2.6 g, 10 mmol) in dimethyl formamide (40 mL) and the mixture was stirred and refluxed for 3 h, then cooled and poured into water (250 mL) . The solid was filtered off, refluxed with 10% nitric acid (125 mL) for 2 h, then cooled on ice. The solid which remained was filtered off, washed with water, dried and recrystallized from acetic acid to give the nitrile (1.8 g, 72%), as a black solid, mp 263-265 °C .

8 -Aminoanthraquinone-1 - carboxylic acid ( 14g)

(a) Reduction of 14e as for the preparation of 14f gave 14g as a red solid mp 256-259 °C (after changing form >205 °C) . Ji NMR δ 7.21 (d) , 7.40 (d) , 7.54 (t), 7.77 (d) ,

7.86 (t) , 8.20 (d) . (b) Nitrile 16e (1.8 g) in sulfuric acid (40 mL, from concentrated sulfuric acid and water 5:1) was heated on an oil bath at 165 °C for 1.5 h, then cooled and poured onto ice/water. The solid which separated was filtered off, washed with water, then stirred with 4% sodium hydroxide solution and filtered. The filtrate was taken to pH 3-4 with concentrated hydrochloric acid and the solid was filtered off, washed with water and dried to give the acid 14g (1.1 g, 57%) .

Methyl 8-Aminoanthraquinone-l -carboxylate (14h)

A mixture of 8-aminoanthraquinone-l-carboxylic acid (14g) (1.71 g, 6.4 mmol), absolute methanol (50 mL) and concentrated sulphuric acid (0.3 g) was refluxed for 16 h, then cooled and filtered. The filtrate was concentrated to a small volume, water was added and the solid which separated was filtered off and air-dried to give the red- brown ester (1.40 g, 78%), mp 149-151 °C . Ji-NMR (CDC1₃) δ 4.02 (s, OCH₃), 6.95 (d) , 7.45 (t), 7.61-7.66 ( , 2H) , 7.74 (t) , 8.34 (d) .

Example 2: Preparation of 2-Methyl-7-oxo-7H- dibenz [f , ij ] isoquinoline-8-carboxylic acid (19c) . An example of the general preparation of the tetracyclic system of Formula II (Y=V=CH, X=N) from aminoanthraquinones according to step 1 of Scheme 3b. A mixture of 14f (2.67 g, 10.0 mmol), acetone (5.8 g) and 4% NaOH solution (75 L) was refluxed for lh, with stirring, under nitrogen. The mixture was then cooled, acidified with concentrated HCl, and the precipitate which formed was collected by filtration, washed with water and dried to give the product as a brown solid (2.78 g, 96%), mp >314 °C (from EtOH/H₂0) . ^XH NMR δ 2 . 83 ( s , 3 H, CH₃ ) , 7 . 66 (d) , 7 . 96 ( t ) , 8 . 04 (t ) , 8 .39 ( d) , 8 . 47 ( d) , 8 . 60 ( s ) , 8 . 78 ( d) .

¹³C NMR δ 25.3 (CH₃), 119.3 (CH) , 121.1 (C) , 125.5 (CH) , 5 127.9 (CH) , 128.1 (C) , 129.0 (CH) , 130.3 (CH) , 133.6 (C) , 134.0 (CH) , 134.3 (C) , 135.6 (CH) . 137.2 (C) , 146.8 (C) , 160.8 (C) , 170.9 (C) , 181.2 (C) . ESMS : m/z 290 (M+l) .

The following were prepared in this manner: 10 2 -Methyl -7H-dibenz [f, ij] isoquinolin-7 -one (19a). This was prepared from 17 (reflux for 24 h) . The precipitate which formed was filtered from the hot mixture, dried and recrystallized from toluene to give the product as a 'light brown solid (72%), mp 244-246 °C (lit. (Schweizer et al, 15 1977) mp 190 °C) .

^XH NMR (CDCI₃) δ 2.79 (s, 3 H, CH₃), 7.60 (t, H-9), 7.71 (t, H-10), 7.86 (t, H-5), 7.96 (s, H-l) , 8.23 (d, H-ll), 8.28 (d, H-4), 8.43 (d, H-8) , 8.56 (d, H-6) . 20

¹³C NMR (CDCI₃) δ 25.7 (CH₃), 117.4 (CH-1) , 121.9 (C-llc) , 123.4 (CH-11), 128.3 (C-6a, CH-6,8), 129.7 (CH-5), 130.3 (CH-9), 132.1 (C-7a) , 133.5 (CH-10), 133.7 (C-lla) , 134.6 (C-llb) , 135.4 (CH-4), 147.3 (C-3a) , 159.9 (C-2), 182.5 ^•25 (C-7) .

The same compound was isolated when this reaction was carried out with 4-aminoanthraquinone-l-carboxylic acid (14c) (reflux under nitrogen for 1 h) . 30 2 -Methyl -7 -oxo-7H-dibenz [£, ij ^'] isoquinoline- 4 - carboxylic acid (19b) . This was prepared from 1- aminoanthraquinone-2-carboxylic acid (12) (Bennett et al , 1982), as a dark solid (95%), mp 264-265 °C (some sublimation > 227 °C) .

35

Hϊ NMR δ 2.59 (s, 3 H, CH₃ ) , 7.57 (t) , 7.67 (t) , 7.91 (d) , 8.01 (d) , 8.06 (s), 8.12 (d) , 8.24 (d) . ¹³C NMR δ 24.6 (CH₃), 119.0 (CH) , 125.0 (CH) , 127.1 (CH) , 127.4 (CH) , 131.6 (CH) , 133.1 (CH) , 134.4 (CH) , 142.9 (C) , 160.0 (C), 165.3 (C) , 180.5 (C) (Low solubility prevented assignment of C at d <140 ppm) . ESMS : m/z 290 (M+l) .

2~Methyl-7-oxo-7H-dibenz [f, ij ] isoquinoline-11- carboxylic acid (19d) . This was prepared from 14g, as a brown solid (0.84 g from 0.80 g) which contained ca 15% impurities (from NMR analysis) and was used in this state in the amidation reaction below.

Ji NMR δ 2.76 (s, 3 H, CH₃ ) , 7.81 (t) , 7.96 (d) , 8.03 (t), 8.07 (s), 8.38 (d) , 8.47 (d) , 8.52 (d) .

8 -Chloro-2 -methyl -7H-dibenz [ f , ij ] isoquinolin-7 -one (19e) . This was prepared from 16c (3 h reflux), as a yellow solid (98%), mp 208-209 °C. (some sublimation > 186 °C) (lit. (Nabar et al, 1983) mp 156-157 °C) .

^XH NMR (CDC1₃) δ 2.85 (s, 3 H, CH₃ ) , 7.58-7.65 (m, 2 H) , 7.90 (t), 8.00 (s), 8.27-8.31 (m, 2 H) , 8.54 (d) .

¹³C NMR (CDCI3) d 25.8 (CH3), 117.8 (CH) , 121.2 (C) , 122.7

(CH) , 128.6 (CH) , 129.2 (C) , 130.1 (CH) , 132.8 (CH) , 134.0 (C), 134.2 (CH) , 134.9 (CH) , 136.7 (C) , 147.1 (C) , 160.0

(C), 181.6 (C) .ESMS: m/z 280 (100%), 282 (36%) (both M+l) .

11 -Chi oro -2 -methyl - 7H-dibenz [f, ij] i soqulnol i -7 -one (19f ) . This was prepared from 16d (3 h reflux) , as a brown-red solid (96%) , mp 160-162 °C [from light petroleum (bp 90-120 °C) ] .

Ji NMR (CDCI3) δ 2.88 (s, 3 H, CH3) , 7.54 (t) , 7.82 (dd, J = 7.9, 1.4 Hz), 7.91 (t) , 8.38 (dd, J = 8.2, 1.1 Hz), 8.54 (dd, J =7.7, 1.4 Hz) , 8.62 (dd, J = 7.3, 1.1 Hz) , 9.31 (s) . ¹³C NMR (CDCI₃) δ 26.1 (CH₃), 122.8 (CH) , 128.0 (CH) , 128.4 (CH) , 129.3 (CH) , 129.9 (CH) , 136.2 (CH) , 137.5 (CH) , 147.2 (C) , 160.1 (C) , 181.6 (C) (Impurities prevented assignment of C at δ <140 ppm) . ESMS: m/z 280 (100%), 282 (36%) (both M+l) .

Example 3 Preparation of 2-Methyl-7H-benzo [e]perimidin- 7-one (19g) .

An exampl e of the general preparation of the tetracycli c system of Formula II (V=CH,

X=Y=N) from aminoanthraquinones according to Scheme 3a . To a solution of N, N-dimethylacetamide (2.18 g, 25 mmol) in dry acetonitrile (30 mL) at 5-10 °C was added, dropwise, phosphoryl chloride (1.86 mL, 20 mmol) during 10 min. The resulting mixture was stirred at room temperature for 1 h, 1-aminoanthraquinone (2.23 g, 10 mmol) was added in one portion and the mixture was stirred at room temperature for 1 h, then at 50 °C for 8 h. After being cooled, the mixture was poured onto ice, basified with 10% ΝaOH, and the precipitate which formed was collected. by filtration, washed with a little water and air-dried to give the red-brown intermediate amidine 18 (2.74 g, 94%), mp 163 °C.

^XH ΝMR (CDCI₃) δ 1.80 (s, 3 H, CH3 ) , 3.13 (s, 6 H, Ν(CH₃)₂), 7.07 (d), 7.55 (t), 7.67-7.71 (m, 2 H) , 7.93 (d) , 8.18- 8.24 (m, 2 H) .

¹³C NMR (CDCI₃) δ 16.0 (CH₃), 38.2 (NCH₃), 121.2 (CH) , 122.8 (C), 126.4 (CH) , 126.9 (CH) , 131.9 (CH) , 132.9 (CH) , 133.6 (CH) , 133.8 (CH) , 135.0 (C) , 135.3 (C) , 154.2 (C) , 156.9 (C), 183.3 (C), 184.1 (C).ESMS: m/z 293 (M+l).

A mixture of amidine 18 (2.20 g, 7.5 mmol), ammonium acetate (2.89 g, 37.5 mmol) and ethanol (40 mL) was ■ refluxed for 2 h, cooled to room temperature, and the precipitate was collected by filtration, washed with ethanol and water, then air-dried. The product was obtained as a pale yellow solid (1.67 g, 90%), mp 211-212 °C (lit. (Weidinger et al, 1963) mp 201-203 °C) .

*H NMR (CDCl₃)δ 3.00 (s, 3 H, CH₃ ) , 7.74 (t), 7.83 (t) , 8.02 (t), 8.23 (d), 8.41(d), 8.51 (d) , 8.90 (d) .

¹³C NMR (CDC1₃) δ 26.8 (CH3), 117.8 (C) , 125.6 (CH) , 127.9 (CH) , 128.4 (CH) , 128.8 (C) , 132.4 (CH) , 133.7 (C) , 133.9

(CH) , 134.1 (CH) , 134.3 (CH) , 134.8 (C) , 149.9 (C) , 156.7 (C) , 165.8 (C), 182.2 (C) . ESMS: m/z 247 (M+l).

7 -Oxo-7Ε-benzo [e] per imidine-11 -carboxylic acid (28b) was prepared in a similar manner, as follows (Scheme 3c) : Methyl 7 -Oxo-7_-benzo [e] per imidine-11 -carboxy late (28a) _ To a stirring solution of methyl 8- aminoanthraquinone-1-carboxylate (14h) (0.10 g, 0.35 mmol) in dry acetonitrile (5 ml) , at room temperature under nitrogen, was added dimethylformamide dimethylacetal (0.27 ml, 1.87 mmol) . The solution was heated under reflux for 3 h then cooled to room temperature. The solvent was removed at reduced pressure to leave the dark brown amidine intermediate (27). Ammonium acetate (0.11 g, 1.19 mmol) and dry ethanol (3 mL) were added and the mixture was heated under reflux for 1 h, then cooled to room temperature and the precipitate that formed was collected by filtration to give 28a as a light tan solid (0.06 g, 58%) . Ji NMR (CDCI3) δ 4.04 (s, OCH₃) , 7.77-7.81 (m, 2H) , 8.12

(t), 8.39 (d), 8.54 (dd, J = 7.1, 1.5 Hz), 8.63 (d) , 9.47 (s) .

7 -Oxo-7E-benzo [e]perimidine-ll -carboxylic acid (28b) . A mixture of ester 28a ( 0 .37 g, 1 .25 mmol ) in trifluoroacetic acid ( 15 mL ) and water ( 15 mL) was heated under reflux for 7 days, then cooled and water was added. The solid which separated was filtered and washed with water to yield the acid as a black solid (0.31 g, 89%) . *H NMR δ 7.83-7.94 (m, 2H) , 8.29 (t) , 8.40-8.45 (m, 2H) , 8.57 (d) , 9.47 (s) .

Example 4 Preparation of 2-Formyl-7ff- dibenz [f , ij] isoquinolin-7-one (20a) . An example of the general oxidation of 2-methyl groups according to Scheme 3b step 2.

To a hot suspension of selenium dioxide (6.66 g, 60 mmol) in 1,4-dioxane (100 mL) , was added 19a (2.45 g, lOmmol) in one portion, and the resulting mixture was heated under reflux for 3 h, then filtered while hot. The filtrate was concentrated to a small volume and the solid which separated was filtered off and recrystallized from dioxane to give the aldehyde as a pale yellow solid (2.30 g, 88%) , mp 261-263 °C.

Ji NMR (CDC1₃) δ 7.69 (t) , 7.81 (t), 8.03 (t) , 8.42 (d) , 8.48 (d), 8.55 (d) , 8.73 (s) , 8.77 (d) , 10.26 (s, CHO) .

¹³C NMR (CDCI3) δ 112.7 (CH) , 124.2 (CH) , 124.9 (C) , 128.5 (CH) , 128.8 (C) , 130.7 (CH) , 131.1 (CH) , 131.8 (CH) , 132.0 (C) , 133.3 (C) , 134.0 (CH) , 136.4 (C) , 137.1 (CH) , 147.4

(C), 153.3 (C), 181.9 (C) , 193.4 (C) . ESMS: m/z 260 (M+l).

The following were prepared in a similar manner:

8-Chloro-2-formyl-7H-dibenz [f, ij] isoquinolin-7 -one (20e) . The filtrate from the hot filtration was evaporated to dryness. The residue was extracted with hot CHCI₃, filtered, and the filtrate was washed twice with water, then brine, and dried (MgSO_ι) . The solvent was removed to give the aldehyde as a yellow solid (2.69 g, 92%), mp 254- 255 °C (some sublimation > 210 °C) . --E NMR (CDCI3) δ 7.66-7.71 (m, 2 H) , 8.04 (t) , 8.41 (dd, J = 6.4, 2.8 Hz, IH) , 8.53 (d) , 8.72 (s) , 8.73 (d) , 10.26 (s, CHO) .

¹³C NMR (CDCI₃) δ 113.2 (CH) , 123.5 (CH) , 124.4 (C) , 128.6 (C) , 129.6 (C) , 131.1 (CH) , 132.1 (CH) , 133.4 (CH) , 135.0 (CH) , 135.9 (C) , 136.3 (C) , 136.6 (CH) , 137.0 (C) , 147.3 (C) , 153.4 (C) , 181.0 (C) , 193.3 (C) . ESMS: m/z 294 (100%) , 296 (35%) (both M+l) .

ll-Chloro-2~formyl-7 -dibenz[f, ij ] isoquinolin-7 -one (20f ) . This was prepared as for 20e, as a grey solid (87%) , mp 224-226 °C.

*H NMR (CDCI₃) δ 7.58 (t) , 7.86 (dd, J = 7.8, 1.3Hz) , 8.04 (t) , 8.52 (dd, J = 7.7, 1.3Hz) , 8.60 (dd, J = 8.3, 1.3Hz) , 8.78 (dd, J = 7.3, 1.3Hz) , 9.97 (s) , 10.28 (s, CHO) .

¹³C NMR (CDCI3) δ 117.9 (CH) , 125.3 (C) , 127.6 (C) , 128.0 (CH) , 130.2 (CH) , 130.4 (C) , 130.6 (CH) , 131.8 (CH) , 133.7 (C), 134.6 (C) , 134.8 (C) , 137.9 (2 x CH) , 147.3 (C) , 153.3 (C) , 181.0 (C), 193.0 (CH) . ESMS: m/z 294 (100%), 296 (37%) (both M+l) .

2-Formyl-7H-benzo[e]perimidin-7-one (20g) . This was prepared as for 20a, and recrystallized from toluene to give a pale yellow solid (89%) , mp 237-238 °C.

^XH NMR .(CDCI3) δ 7.80 (t) , 7.88 (t), 8.19 (t) , 8.44 (d) , ^' 8.53 (d) , 8.72 (d) , 9.05 (d) , 10.36 (s, CHO) .

^:H NMR (CDCI3) δ 120.2 (C) , 126.4 (CH) , 128.2 (CH) , 129.1 (C) , 131.7 (CH) , 131.8 (C) , 133.4 (CH) , 133.6 (C) , 135.0 (CH) , 136.0 (CH) , 149.6 (C) , 156.6 (C) , 158.4 (C) , 181.5 (C), 191.7 (C) . ESMS: m/z 293 (M+l) . Example 5 Preparation of 7-Oxo-7J- dibenz [ f, ij] isoquinoline-2-carboxylic acid (21a) . An example of the general oxidation of an aldehyde group according to Scheme 3b step 3 .

A solution of sodium chlorite (4.0 g) and sodium dihydrogen phosphate (4.0 g) in water (40 mL) was added over 30 min. to a mixture of aldehyde 20a (1.04 g, 4 mmol) in dioxane (80 mL) and 2-methylbut-2-ene (20 mL) , and the resulting mixture was stirred at room temperature for 16 h. Most of the organic solvent was evaporated at reduced pressure, water (20 mL) was added to the residue, the pH was adjusted to 2 with concentrated HCl, and the precipitate which separated was collected by filtration, washed with water and dried to give the acid as a yellow solid (0.98 g, 89%), mp 224-225 °C (from CHCl3).

H NMR (CDC1₃) δ 7.73 (t) , 7.86 (t), 8.07 (t), 8.48-8.54 (m, 3 H) , 8.82 (d) , 9.05 (s) .

¹³C NMR (CDCI₃) δ 114.8 (CH) , 124.5 (CH) , 124.9 (C) , 128.7 (CH) , 128.8 (C), 131.2 (CH) , 131.6 (CH) , 131.8 (CH) , 132.1 (C), 133.0 (C), 134.3 (CH) , 136.0 (CH) , 138.1 (C) , 145.4 (C) , 146.9 (C) , 163.9 (C) , 181.9 (C) . ESMS: m/z 276 (M+l).

The following acids were prepared in similar manner: 8 -Chloro-7 -oxo-7H-dibenz [ f , ij ] isoquinoline-2 - carboxylic acid (21e) . This was prepared from 20e, as an orange solid (98%), mp 245-247 °C. H NMR δ 7.76-7.82 (m, 2 H) , 8.10 (t, 8.50-8.53 (m, 2 H) , 8.69 (d) , 8.92 (s). ESMS: m/z 310 (100%), 312 (42%) (both M+l) . ll -Chloro- ' -oxo-7H-dibenz [f , ij ] isoquinoline-2 - carboxylic acid (21f) . This was prepared from 20f, as a brown solid (99%), mp 230-231 °C (some sublimation > 196 °C) . H NMR δ 7 .72 ( t ) , 8 . 01 ( d) , 8 . 11 ( t ) , 8 .36 ( d) , 8 . 56-8 . 62 (m, 2 H) , 9 . 94 ( s ) .

¹³C NMR δ 120.5 (CH) , 123.4 (C) , 126.6 (C) , 127.5 (CH) , 129.5 (C) , 130.5 (CH) , 130.7 (CH) , 131.1 (CH) , 132.3 (C) , 133.1 (C) , 134.1 (C), 137.7 (CH) , 137.9 (CH) , 146.2 (C) , 149.6 (C) , 165.9 (C) , 179.8 (C) . ESMS: m/z 310 (100%), 312 (40%) (both M+l) .

^' -Oxo-7H-benzo [ e] perimidine-2 -carboxyli c acid (21g) . This was prepared from 20g, as a yellow solid (0.25 g, 90%) , mp 266-267 °C .

Ji NMR δ 7.88 (t) , 7.98 (t) , 8.25-8.31 (m, 2 H) , 8.49 (d) , 8.56 (d) , 8.84 (d) . ¹³C NMR δ 119.2 (C) , 125.7 (CH) , 127.6 (CH), 128.5 (C), 130.4 (CH) , 133.3 (CH) , 134.1 (C) , 134.8 (CH) , 135.5 (CH) , 149.0 (C) , 156.0 (C) , 157.1 (C) , 165.6 (C), 181.2 (C) . ESMS: m/z 277 (M+l).

Example 6 Preparation of 7-oxo-lH- dibenz [ f, ij] isoquinoline-2, 8-dicarboxylic acid (21c) . A. ixture of 19c (2.32 g, 8.0 mmol) and Seθ₂ (4.0 g) in dry dioxane (lOOmL) was heated to reflux, with stirring, for 4 h., then filtered while hot, and the filtrate was evaporated to dryness at reduced pressure. The residue was extracted with 5% NaOH and the extract was acidified with concentrated HCl. The precipitate which formed was collected by filtration, washed with water and dried at 80 °C to give the diacid as a brown solid (2.12 g, 83%) , mp. >316 °C.

*H NMR δ 7.68 (d) , 7.95 (t) , 8.16 (t) , 8.58-8.64 (m, 2 H) , 8.84 (d) , 9.04 (s) . ESMS: m/z 320 (M+l). Example 7 Preparation of N- [2- (Dimethylamino) ethyl] -7- oxo-7H-dibenz [f, ij] isoquinolin-2-carboxamide (24a) . An example of the general method A for ami de form tion.

To a solution of acid 21a (0.22 g, 0.8 mmol) in dry tetrahydrofuran (15 mL) was added 1, 1 ' -carbonyldiimidazole (0.20 g, 1.2 mmol) and the mixture was refluxed for 3 h. The solvent was removed at reduced pressure and the residue was dissolved in CH₂CI₂ (20 mL) , washed with 10%

Νa₂C0₃ solution, warm water and dried (Na₂Sθ ) . The solvent was evaporated to give the intermediate imidazolide (22) as a pale brown solid (0.23 g, 88%), mp 210-211 °C, which was used in the next step without further purification.

^αH NMR (CDCI₃) δ 7.20 (s, ImH) , 7.70 (t) , 7.81 (t) , 8.05

(t) , 8.06 (s, ImH), 8.42-8.53 (m, 3 H) , 8.79 (d) , 8.97 (s) , 9.15 (s, ImH) .

¹³C NMR (CDCI3) δ 117.3 (CH) , 118.2 (CH) , 124.2 (CH) , 124.4 (C) , 128.6 (CH) , 130.5 (CH) , 131.1 (CH) , 131.4 (CH) , 132.1 (CH) , 133.0 (C) , 134.1 (CH) , 137.0 (CH) , 140.0^' (CH) , 146.2 (C) , 150.0 (C) , 163.1 (C) , 181.9 (C)-3 (C) were not observed.

To a solution of imidazolide 22 (0.17 g, 0.52 mmol) in dry CH₂C1₂ (15 mL) was added a solution of N,N- dimethylethylenediamine (55 mg, 0.62 mmol) in CH₂C1₂ (5mL) .

The solution was stirred at room temperature for 24 h, then washed with 10% ₂C0₃ solution, warm water (10 L x 2), dried (Na Sθ₄) and the solvent was removed to give the amide 24a as a brown solid (0.15 g, 83%), mp 143-144. °C [from toluene/light petroleum (bp 60-90 °C) ] .

Ji NMR (CDCI₃) δ side chain + 7.63 (t) , 7.76 (t) , 7.91 (t) , 8.38 (d), 8.41 (2 x d, 2 H) , 8.49 (br s, NH) , 8.63 (d) , 8.97 (s) . ¹³C NMR (CDCI₃) δ 37.4 (CH₂) , 45.4 (CH₃), 58.2 (CH₂) , 114.5 (CH) , 124.1 (C), 124.3 (CH) , 128.3 (CH) , 128.5 (C) , 130.2 (CH) , 130.5 (CH) , 130.8 (CH) , 131.9 (C) , 133.6 (C) , 133.9 (CH) , 136.2 (C), 136.4 (CH) , 146.0 (C) , 150.9 (C) , 164.1 (C) , 182.2 (C) . ESMS: m/z 346 (M+l).

For microanalysis, a sample was converted to the ono-perchlorate salt, mp 252-253 °C (from EtOH) . Anal. Calc for C ιHι₉N₃0 .HCIO4. C, 56.6; H, 4.3; N, 9.4. Found: C, 56.2; H, 4.4; N, 9.1.

The following imidazolides and amides were made in a similar manner:

N- [2- (Dimethylamino) ethyl] - 2 -methyl -7 -oxo -7 E- dibenz [f, ij ] isoquinoline-4-carboxamide (24f) . The intermediate imidazolide was a brown-red solid (85%), mp 163-165 °C.

Ji NMR (CDCI₃) δ 2.71 (s, 3 H, CH₃ ) , 7.11 (s, ImH), 7.48 (s, ImH), 7.69 (t) , 7.75 (s, ImH), 7.81 (t), 8.05 (d) , 8.09 (s), 8.34 (d) , 8.50 (d) , 8.67(d).

¹³C NMR (CDCI3) δ 25.9 (CH3) , 117.1 (CH) , 118.4 (CH) , 122.1 (C) , 123.6 (CH) , 127.2 (CH) , 128.7 (CH) , 129.0 (CH) , 130.6 (C) , 130.9 (2 x CH) , 131.9 (C) , 133.5 (C) , 134.1 (CH) , 134.9 (C) , 137.3 (C) , 138.3 (CH) , 144.7 (C) , 161.8 (C) , 165.6 (C) , 181.9 (C) .^{' ■}

The amide 24f was obtained as a brick-red solid (63%) , mp 207-208 °C [from benzene/light petroleum (bp 60- 90 °C) ] .

^αH NMR ^"(CDCI3) δ side chain + 2.86 (s, 3 H, CH₃) , 7.63 (t) , 7.75 (t) , 8.01 (s) , 8.25 (d) , 8.43 (d) , 8.62 (d) , 8.94 (d) , 11.63 (br s, NH) .

¹³C NMR (CDCI3) δ 25.4 (CH₃), 37.7 (CH₂) , 45.2 (CH3), .57.9 (CH₂) , 116.9 (CH) , 121.9 (C) , 123.5 (CH) , 127.7 (CH) , 128.. (CH) , 130.0 (C) , 130.7 (CH) , 131.7 (C) , 133.0 (CH) , 133.3 (C) , 133.5 (C), 133.7 (CH) , 135.7 (C) , 144.8 (C) , 159.2 (C), 164.8 (C) , 182.1 (C) . ESMS: m/z 360 (M+l) . Anal. Calc. for C₂₂H₂iN₃0₂: C, 73.5; H, 5.9; N, 11.7. Found: C, 73.2; H, 5.9; N, 11.6.

N- [2- (Dimethylamino) ethyl] - 2 -methyl -7 -oxo -7 E- dibenz [f , ij] isoquinoline- 8 -carboxamide (24g) . The intermediate imidazolide was a dark solid (83%), mp >310 °C.

^XH NMR (CDC1₃) δ 2.90 (s, 3 H, CH₃ ) , 7.09 (S, ImH), 7.46 (s, ImH), 7.64 (d) , 7.77 (s, ImH), 7.88-7.95 (m, 2 H) , 8.14 (s), 8.36 (d), 8.46 (d) , 8.57 (d) .

The imidazolide was reacted as in the preparation of 24a, but in tetrahydrofuran as solvent and with reflux for 3 days. The solvent was removed and the residue was dissolved in CHC1₂ and treated as for 24a to give the amide 24g as a brick-red solid (67%), mp 174-176 °C [from benzene/light petroleum (bp 60-90 °C) ] .

Ji NMR (CDCI₃) δ side chain + 2.85 (s, 3 H, CH₃), 6.38 (br s, NH) , 7.54 (d), 7.73 (t) , 7.87 (t) , 8.02 (s), 8.30 (d) , 8.32 (d) , 8.52 (d) .

¹³C NMR (CDCI₃) δ 25.8 (CH₃), 37.4 (CH₂) , 45.0 (CH₃), 57.5 (CH₂), 117.9 (CH) , 121.5 (C) , 124.4 (CH) , 128.5 (C) , 128.6 (CH) , 129.2 (C), 130.0 (2 x CH) , 133.0 (CH) , 134.3 (C) , 134.7 (C), 135.3 (CH) , 139.9 (C) , 147.3 (C) , 160.1 (C) ,

171.1 (C) , 181.8 (C) . ESMS: m/z 360 (M+l). Anal. Calc. for C₂₂H₂ιN₃0₂ H₂0: C, 70.0; H, 6.1; N, 11.13. Found: C, 70.0, H, 5.7; N, 11.3.

N- [2- (Dimethylamino) ethyl] -2 -methyl -7- oxo -7 E- dibenz [f, ij ] isoquinoline-11-carboxamide (24h) . The intermediate imidazolide was prepared in dioxane (4 h reflux) and obtained as an impure brown solid (1.45 g from 1.45 g of impure 19d) .

This was reacted with N,N-dimethylethylenediamine as for 24g to give a brown solid (1.3 g) which was washed through a short alumina column with CHCI₃ to give the crude product (1.0 g) .

This was chromatographed repeatedly [neutral alumina, ethyl acetate/diethylamine (50:1), 65 °C] to give sufficient of the target compound free of an amide impurity which eluted only slightly ahead. Amide 24h was obtained as an orange solid (0.33 g) , mp 183-185 °C (from ethyl acetate) .

^XH NMR (CDCI₃) δ side chain + 2.85 (s, 3 H, CH₃ ) , 6.97 (br s, NH) , 7.56 (t), 7.68 (dd, J =7.4, 1.5 Hz), 7.80 (tj , 8.07 (s), 8.26 (dd, J = 8.2, 0.9 Hz), 8.43 (dd, J = 7.7, 1.5 Hz), 8.48 (dd, J = 7.3, 0.9 Hz).

¹³C NMR (CDCI₃) δ 25.8 (CH₃), 37.7 (CH₂) , 45.0 (CH₃), 57.2 (CH₂) , 121.4 (CH) , 122.0 (C) , 127.5 (C) , 128.4 (CH) , 129.4 (2 x CH), 129.7 (CH) , 130.6 (C) , 133.0 (C) , 133.1 (C) , 133.6 (CH) , 135.9 (CH) , 136.5 (C) , 147.2 (C) , 159.7 (C) , 170.9 (C) , 182.0 (C) . ESMS: m/z 360 (M+l). Anal. Calc. for C₂₂H2_lN₃O₂.0.25H₂O: C, 72.6; H, 5.9; N, 11.6. Found: C, 72.2, H, 5.8; N, 11.4.

N- [2- (Dimethylamino) ethyl] -2, 4 -dimethyl -7 -oxo-7H- dibenz [f, ij] isoquinoline-11 -carboxamide (24i) . Alternative purification of the above crude 24h (550 mg) by preparative HPLC (on C-18 reverse-phase silica gel, eluting with a mixture of 65% of 4:1 MeCN/water and 35% of 0.045 M ammonium formate, pH 4.5) gave two peaks, eluting at 15 and 21 min. Further purification of these products, by filtration through columns of silica gel in a gradient of 0-4% MeOH in CH₂C1₂, gave respectively 24h (212 mg of 97% purity) and 24i (80 mg of 97% purity. The latter compound had H NMR (CDCI3) δ side chain + 2.88 (s, 3 H, CH₃) , 2.97 (s, 3 H, CH₃), 7.53 (t, J = 7.6 Hz, 1 H, H-9), 7.62 (d, J = 7.9 Hz, 1 H, H-5), 7.67 (dd, J = 7.5, 1.5 Hz, 1 H, H-10), 8.05 (s, 1 H, H-l), 8.34 (d, J = 7.5 Hz, 1 H, H-6), 8.37 (dd, J = 8.0, 1.5 Hz, H-8) . HRMS Calculated for C₂₃H₂₃N₃0₂; 373.1790. Found; 373.1782.

N- [2- (Dimethylamino) ethyl] -8-chloro-7-oxo-7H- dibenz [f, ij] isoquinolin-2-carboxamide (24c). The intermediate imidazolide separated when the THF solution was cooled, as a yellow solid (90%), mp 242 °C (dec).

^αH NMR (CDCI₃) δ 7.20 (s, ImH), 7.65-7.72 (m, 2 H) , 8.06- 8.10 (m, 2 H) , 8.44 (d) , 8.52 (d) , 8.75 (d) , 8.98 (s), 9.15 (s, ImH) .

The amide 24c was obtained as a yellow solid (98%) , mp 117-119 °C [from benzene/light petroleum (bp 60-90 °C)] .

Ji NMR (CDCI₃) δ side chain + 7.67-7.70 (m, 2 H) , 7.99 (t) , 8.45 (dd, J = 8.4, 1.2 Hz, 1 H) , 8.50 (dd, J = 5.7, 3.6 Hz, 1 H) , 8.53 (br s, NH) , 8.68 (dd, J = 7.3, 1.2 Hz, 1 H) , 9.07 (s) .

¹³C NMR (CDCI₃) δ 37.3 (CH₂) , 45.4 (CH₃), 58.2 (CH₂) , 114.9 (CH) , 123.7 (CH) , 128.5 (C) , 129.4 (C) , 130.6 (CH) , 130.8 (CH) , 133.3 (CH) , 134.7 (CH) , 135.8 (CH) , 136.0 (C) , 136.6 (C), 136.8 (C), 145.9 (C) , 150.9 (C) , 161.1 (C) , 181.3 (C) . ESMS: m/z 380 (100%), 382 (36%), (both M+l) . Anal. Calc. for C₂ιH₁₈ClN₃0₂.H₂0: C, 63.4; H, 5.1; N, 10.6; Found:

C, 63.3; H, 4.6; N, 10.2.

N- [2- (Dimethylamino) ethyl] -ll-chloro-7 -oxo-7H- dibenz [f, ij ] isoquinolin-2 -carboxamide (24d) . The intermediate imidazolide was obtained as a yellow solid (97%) , mp 227 °C (dec.) . ^!H NMR (CDCI₃) δ 7.20 (s, ImH), 7.61(t), 7.88 (d) , 8.02 (s, ImH), 8.07 (t), 8.55 (d) , 8.59 (d) , 8.82 (d) , 9.13 (s, ImH) , 10.25 (s) .

The amide 24d was obtained as a viscous oil (83%), which gradually solidified at room temperature. Further purification was achieved by column chromatography [alumina, CHCl₃/benzene (1:1)], followed by recrystallization from CHCl₃/hexane to give a yellow solid, mp 109-110 °C .

^XH NMR (CDCI₃) δ side chain + 7.56 (t) , 7.87 (d) , 7.97 (t), 8.47-8.55 (m, 3 H (incl. NH) ) , 8.74 (d) , 10.32 (s) .

¹³C NMR (CDCI₃) δ 37.5 (CH₂) , 45.5 (CH₃) , 58.3 (CH₂) , 119.7 (CH), 124.7 (C), 127.6 (C) , 127.9 (CH) , 129.8 (CH) , 130.3 (CH) , 130.6 (CH), 131.0 (C) , 133.8 (C) , 134.6 (C) , 134.9 (C), 137.4 (CH), 137.9 (CH) , 146.4 (C) , 151.1 (C) , 164.0 (C) , 181.4 (C) . ESMS: m/z 380 (100%), 382 (36%), (both M+l). Anal. Calc. for C₂ιHι₈ClN₃θ₂.0.5H₂0: C, 64.9; H, 4.9;

N, 10.8. Found: C, 65.2; H, 4.7; N, 10.8.

N- [2- (Dimethylamino) ethyl] -7-oxo-7H- benzo[e] per imidine-2 -carboxamide (24b) . The intermediate imidazolide was prepared in dioxane, and obtained as a pale yellow solid (81%), mp 205 °C (dec).

Ji NMR (CDCI₃) δ 7.22 (s, ImH), 7.81-7.92 (m, 2 H) , 7.94 (s, ImH), 8.24 (t) , 8.47 (d) , 8.52 (d) , 8.74 (s, ImH), 8.76 (d) , 8.94 (d) .

The amide 24b was obtained as a yellow solid (82%), mp 141-142 °C [from benzene/light petroleum (bp 60-90 °C)] . ^XH NMR (CDCI3) δ side chain + 7.78 (t) , 7.87 (t) , 8.13 (t), 8.42 (d), 8.53 (d) , 8.65 (d) , 8.95 (br s, NH) , 9.04 (d) .

¹³C NMR (CDCI3) δ 37.2 (CH₂) , 45.2 (CH₃) , 58.1 (CH₂), 119.8 (C) , 126.4 (CH) , 128.1 (CH) , 129.0 (C) , 130.7 (CH) , 133.1 (CH) , 133.7 (C) , 134.0 (C) , 134.4 (CH) , 134.7 (CH) , 136.0 (CH) , 149.7 (C) , 154.9 (C) , 157.5 (C) , 162.9 (C) , 181.7 (C) . ESMS: m/z 347 (M+l) . Anal. Calc. for C₂₀H₁₈N4θ₂.H₂0: C, 65.9; H, 5.5; N, 15.4. Found: C, 66.3; H, 5.1; N, 15.3.

N- [3- ( dimethylamino ) propyl ] -2 -methyl-7 -oxo-7H- naphtho [1, 2 ,3-de] quinoline-11-carboxamide (24j). Reaction of 19d (500 mg) with CDI and direct reaction of the crude imidazolide with excess 3- (dimethylamino) propane-1, 3- diamine, followed by purification of the product on HPLC, gave 24j as a solid (100 mg) ; ^ NMR (CDC1₃) δ 2.60 (m, 2 H) , 2.12 (s, 6H) , 2.27 (m, 2H) , 2.50 (m, 2H) , 2.76 (s, 3H) , 7.78 (t), 7.80 (dd, J =7.4, 1.5 Hz), 7.84 (t), 8.14 (s), 8.41 (dd, J = 8.2, 0.9 Hz), 8.46 (dd, J = 7.7, 1.5 Hz), 8.56 (dd, J = 7.3, 0.9 Hz), 8.83 (br s, NH) .

2 -Methyl-7 -oxo-N- [2- (1 -piperidinyl) ethyl] -7H- naphtho[l, 2, 3 -de] quinoline-11 -carboxamide (24k). Reaction of 19d (500 mg) with CDI and direct reaction of the crude imidazolide with excess (1-piperidinyl) ethane-1, 2-diamine, followed by purification of the product on HPLC, gave 24k as a solid (90 mg) ; Ji NMR (CDC1₃) δ 1.38 (m, 2H) , 1.50 (DDDm, 4H) , 2.40 (m, 4H) , 2.76 (s, 3H) , 3.46 (m, 4H) , 7.78 (t), 7.80 (dd, J =7.4, 1.5 Hz), 7.84 (t) , 8.14 (s), 8.41 (dd, J = 8.2, 0.9 Hz), 8.46 (dd, J = 7.7, 1.5 Hz), 8.56 (dd, J = 7.3, 0.9 Hz), 8.79 (br s, NH) .

N-{2- [ (2 -hydroxy ethyl ) amino] ethyl } -2 -methyl -7 -oxo - 7H- naphtho [1,2 ,3-de] quinoline-11-carboxamide (241). Reaction of 19d (500 mg) with CDI and direct reaction of the crude imidazolide with excess (2-hydroxyethyl) amino] ethane-1, 2- diamine, followed by purification of the product on HPLC, gave 241 as a solid (100 mg) ; H NMR (CDC1₃) δ 2.61 (m, 2H) , 2.76, (s, 3H) , 2.78 (m, 2H) , 3.44 (m, 4H) , 4.45 (br s, IH) , 7.78 (t), 7.80 (dd, J =7.4, 1.5 Hz), 7.84 (t), 8.14 (s), 8.41 (dd, J = 8.2, 0.9 Hz), 8.46 (dd, J = 7.7, 1.5 Hz), 8.56 (dd, J = 7.3, 0.9 Hz), 8.85 (br s, NH) . Anal or mass spec

N- [ (IS) -2- (dimethylamino) -1 -methyl ethyl] -2 -methyl-7 -oxo- 7H-naphtho [1 , 2 , 3 -de] quinoline-11 -carboxamide (24m) . Reaction of 19d (500 mg) with CDI and direct reaction of the crude imidazolide with excess (15) -2- (dimethylamino) - 1-methylethane-l, 2-diamine, followed by purification of the product on HPLC, gave 24m as a solid (25 mg) ; ¹H NMR

(CDCI3) δ 1.47 (d, J = 6.3 Hz, 3H) , 2.29 (s, 6H) , 2.37 (m, IH) , 2.54 (m, IH) , 2.78 (s, 3H) , 4.31 (m, IH) , 6.96 (br s, NH) , 7.51 (t), 7.65 (dd, J =7.4, 1.5 Hz), 7.79 (t) , 8.19

(s), 8.26 (dd, J = 8.2, 0.9 Hz), 8.37 (dd, J = 7.7, 1.5 Hz), 8.41 (dd, J = 7.3, 0.9 Hz).

N- [ (IR) -2- (dimethylamino ) -1 -methyl ethyl] -2 -methyl -7 -oxo- 7H-naphtho [1 , 2, 3 -de] quinoline- 11 -carboxamide (24n) . Reaction of 19d (500 mg) with CDI and direct reaction of the crude imidazolide with excess (1-R) -2- (dimethylamino) - 1-methylethane-l, 2-diamine, followed by purification of the product on HPLC, gave 24n as a solid (53 mg) ; ¹H NMR

(CDCI3) δ 1.47 (d, J = 6.3 Hz, 3H) , 2.29 (s, 6H) , 2.37 (m, IH) , 2.54 (m, IH) , 2.78 (s, 3H) , 4.31 (m, IH) , 6.96 (br s, NH) , 7.51 (t), 7.65 (dd, J =7.4, 1.5 Hz), 7.79 (t), 8.19

(s), 8.26 (dd, J = 8.2, 0.9 Hz), 8.37 (dd, J = 7.7, 1.5 Hz), 8.41 (dd, J = 7.3, 0.9 Hz).

N, N' - [ [ ( 2 -Aminoethyl) methyl imino] di -3 , 1 - propanediyl] bis- [7-oxo-7H-dibenz [f, ij ] isoquinoline-2 - carboxamide] (26) . Imidazolide 22 (1 mmol) was treated with N, N'- [bis (2-aminoethyl) ] -I\T, AT'-dimethyl-l, 3- propanediamine (0.5 mmol) as for the preparation of 24a, and column chromatography of the crude product (alumina, CHCI3) gave the bisamide as a fawn solid (0.11 g, 31%), mp

155-156 °C.

²H NMR (CDCI₃) δ 1.81 (m, 2 H, CH₂) , 2.34 (s, 6 H, NCH₃ ) , 2.64 (m, 4 H, CH₂) , 2.70 (m, 4 H,CH ), 3.60 (m, 4 H, CH₂), 7.54 (t, 2 H) , 7.68 (t, 2 H) , 7.87 (t, 2 H) , 8.20-8.26 (m, 6 H) , 8.46 (br s, 2 H, NH) , 8.55 (d, 2 H) , 8.71 (s, 2 H) .

¹³C NMR (CDCI3) δ 25.3 (CH₂), 37.2 (CH₂) , 42.1 (CH₃), -55.6 (CH₂), 56.2 (CH₂), 114.2 (CH) , 123.8 (C) , 124.1 (CH) , 128.1 (CH) , 128.4 (C), 130.1 (CH) , 130.4 (CH) , 130.6 (CH) , 131.7 (C), 133.3 (C), 133.8 (CH) , 135.9 (C) , 136.2 (CH) , 145.7 (C), 150.6 (C), 163.6 (C) , 181.8 (C) . ESMS: m/z 703 (M+l). Anal. Calc. for C₄₃H_{38 6}O₄.H₂0: C, 71.7; H, 5.6; N, 11.7. Found: C, 71.6; H, 5.4; N, 11.6.

Example 8 Preparation ofN,N'-bis [2-

(dimethylamino) ethyl] -4-chloro-7-oxo-7ff- dibenz [ f , ij] isoquinolin-2 , 8-dicarboxamide

(24e) . An example of the general method B for ami de forma tion. A mixture of diacid 21c (0.40 g, 1.25 mmol) and thionyl chloride (10 L) was heated at 80 °C for 1 h. After being cooled, the excess of thionyl chloride was removed at reduced pressure and finally by azotropic distillation with benzene. To the residue, dissolved in CHCl₂ (25 mL) and cooled on ice, was added dropwise Ν,Ν- dimethylethylenediamine (0.44 g, 5 mmol) in CH₂CI₂. The resulting solution was stirred at 0 °C for 30 min, room temperature for 1 h., then washed with 10% NaHC0₃ solution, water, brine and dried (MgSθ₄) . The solvent was removed and the residue was chromatographed [alumina- (neutral); CHCI₃ (containing 1% Et₂NH) ] to give the diamide as a brown solid (0.17 g, 27%), mp. 152-154 °C .

Jl NMR (CDCI3) δ 2.29 [s, N(CH₃)₂], 2.34 [s, N(CH₃)₂], 2.61 (t, J = 6.2 Hz, CH₂) , 2.67 (t, J = 6.0 Hz, CH₂) , 3.58-3.68 ( , 4 H, CH₂ x 2), 6.69 (br t, NH) , 7.55 (d, H-9), 7.72 (d, H-10), 7.92 (d, H-5), 8.34 (d, H-11), 8.37 (d, H-6), 8.54 (br t, NH) , 8.81 (s, H-l) .

¹³C NMR (CDCI3) δ 37.4 (2 x CH₂) , 45.1 (CH3) , 45.4 ( CH3 ) , 57.6 (CH₂) , 58.0 (CH₂) , 115.3 (CH-1) , 124.4 (C-llc) , 125.2 (CH-11) , 127.1 (C-6a) , 128.4 (C-7a) , 130.5 (CH-6) , 130.7 (CH-9) , 130.8 (CH-5) , 133.5 (CH-10) , 133.9 (C-lla) , 136.4 (C-llb) , 139.9 (C-8) , 141.6 (C-4) , 141.9 (C-3a) , 150.7 (C- 2) , 163.3 (CONH-2), 170.6 (CONH-8), 180.5 (C-7) . ESMS: m/z 494 (100%) , 496 (40%) (both M+l) . Anal. Calc. for C26H28C1N₅0₃.H₂0: C, 61.0; H, 5.9; N, 13.7. Found: C, 61.3; H, 5.8; N, 14.1. The following amides were made in a similar manner: N- [2- (Dimethylamino) ethyl] -7-oxo-7H- benz [de] anthracene-11 -carboxamide (25) . Thionyl chloride (0.07 g, 0.59 mmol) was added to a suspension of 23 (0.14 g, 0.51 mmol) in dry benzene (20 mL) and the whole was heated under reflux for 2 h, then cooled to room temperature. N,N-Dimethylethylenediamine (0.11 g, 1.25 mmol) was added over 10 min. and the resulting mixture was stirred for 1 h, then heated under reflux for 1 h. Volatile material were evaporated at reduced pressure and the residue was dissolved in chloroform (30 mL) , washed with water (2 x 10 mL) , and dried. (Νa₂S0₄) . The solvent was removed to give 25 as a pale yellow solid (0.16g, 91%) , mp 163-165 °C [from benzene/light petroleum (bp 60- 90 °C) ] .

*H NMR (CDCI₃) δ side-chain + 6.57 (br t, CONH), 7.51 (t), 7.68 (t), 7.69-7.75 (m, 2 H) , 7.96 (d) , 8.18 (d) , 8.47 (d) , 8.52 (d) , 8.68 (d) .

¹³C NMR (CDCI₃) δ 37.6 (CH₂), 45.0 (CH₃), 57.3 (CH₂) , 125.4 (C), 126.27 (CH) , 126.34 (CH) , 17.7 (CH) , 128.2 (C) , 128.6 (CH) , 129.3 (CH) , 129.8 (CH) , 130.6 (CH) , 132.3 (C) , 132.8 (C), 133.3 (C), 133.7 (CH) , 135.5 (CH) , 135.8 (C) , 171.8 (C) , 183.3 (C) . Anal. Calc. for C₂₂H₂₀N₂O₂ • 0.75H₂0: C, 73.8; H, 6.05; N, 7.8. Found: C, 74.0; H, 5.6; N, 7.9.

N- [2- (Dimethylamino) ethyl] -7 -oxo-1 H-benzo [e]perimidine-ll- carboxamide (24o) . This was prepared from acid 28b, as for 25, in 73% yield as a brick red solid, mp 90-91 °C [from chloroform/light petroleum (bp 60-90 °C) (1:2)]. H NMR (CDCI3) δ side chain + 6.62 (br s, NH) , 7.75-7.77 (m, 2H) , 8.09 (t) , 8.35 (d) , 8.51 (t), 8.61 (d) , 9.47 (s). Example 9 Anti-Tumour Activity In Vi tro

The series of tetracyclic carboxamides identified in Table 1 was evaluated for growth inhibitory properties, measured as IC50 values, against murine P388 leukemia cells, Lewis lung carcinoma cells (LLTC), and human Jurkat leukemia cells (JLQ) . together with their amsacrine- and doxorubicin-resistant derivatives (JL/_. and JLβ respectively) , which were obtained and cultured as previously described (Finlay et al , 1990; Finlay et al , 1994) . Growth inhibition assays were performed by culturing cells in microculture plates (150 μl per well) as follows: P388: 4.5 X 10³ cells/well; 3 days LLTC: 1 X 10³ cells/well; 4 days Jurkat lines: 3.75 X 10³ cells/well; 4 days

Cell growth was determined by [ 3H] -thymidine uptake (P388) (Marshall et al , 1992) or by the sulphorhodamine assay (Skehan et al , 1990) . Independent assays were performed in duplicate, using doxorubicin, etoposide, camptothecin and DACA as reference compounds. The results are summarized in Table 2.

Table 2

Anti-Tumour Activity In Vitro of Tetracyclic Carboxamides

Footnotes a

IC50; concentration of drug to reduce cell number to

50% of control cultures.

R¹=(CH₂)₂NMe₂, R²= (CH₂) ₃NMe₂, R³= (CH₂) ₂N-piperazine,

R⁴= (CH₂)₂NH(CH₂)₂OH, R⁵=CH (S-Me) CH₂NMe₂ , R⁶=CE{R-

Me)CH₂NMe₂.

R²-R do not relate to the corresponding references in formulae II and III

Murine P388 leukemia. ^d Murine Lewis lung carcinoma. ^e Human Jurkat leukemia.

The L line is resistant to the DNA intercalator amsacrine and similar agents because of a reduced level of topo II enzyme. The JLjj line is resistant to doxorubicin, primarily by virtue of altered levels of topo II, but probably also via additional mechanisms . The ratios of the IC50 values of a drug in the parent line compared with one of the sublines (IC50 [ g /IC50 [JLc] and

(IC50 [JLD] /IC50 [J c] ) therefore provide some indication of the mechanism of cytotoxicity. Classical topo II inhibitors such as amsacrine, doxorubicin and etoposide have large ratios (10-90 fold) , whereas topo I inhibitors such as camptothecin and mixed topo I/II inhibitors such as DACA (4) have ratios of only about 2-fold. Values of these ratios of less than about 1.5-2 therefore suggest cytotoxicity by a non-topo II mediated mechanism.

Example 10: Anti-Tumour Activity In Vivo

Compounds 24h, 24f and 24i were evaluated against murine colon 38 tumours implanted subcutaneously in

C57BL/6 mice. This advanced colon 38 tumour model is fairly refractory to standard clinical topo II agents, as well as to anti-metabolites and alkylating agents. It therefore represents a good test for in vivo activity.

₃ Colon 38 tumours were grown subcutaneously from 1 mm fragments implanted in one flank of C57/B1 mice (anaesthetised with pentobarbitone 90 mg/kg) . When tumours reached a diameter of approximately 4 mm (7-8 days) , mice were divided into control and drug treatment groups (5 mice/group) , with similar average tumour volumes in each group. Drugs were administered as solutions of the hydrochloride salts in distilled water, and were injected in a volume of 0.01 mL/g body weight in two equal injections administered 1 h apart. Mice treated with the prior art compounds Doxorubicin, Daunorubicin, Amsacrine, Mitoxantrone, DACA, Etoposide or Irinotecan were used as positive controls. The mice were monitored closely, and tumour diameters were measured with callipers three times a week. Tumour volumes were calculated as 0.52xa 2Xb, where a and b are the minor and major tumour axes, and data plotted on a semilogarithmic plot as mean tumour volumes

(+ SEM) versus time after treatment . The results are shown in Figures 5, 6 and 7, and summarized in Table 3. The growth delay was calculated as the time .taken for tumours to reach a mean volume four-fold higher than their pre- treatment volume .

Table 3

It is evident from Table 3 that of the prior art compounds tested, only Doxorubicin, Irinotecan, and DACA resulted in a significant delay in growth of the Colon 38 tumour, and none resulted in long term survival. Of the three compounds of the invention, 24f and 24i resulted in a growth delay comparable to that induced by Doxorubicin and Irinotecan, and did not result in long term survival. However, compound 24 resulted in a growth delay of at least 30 days, and four of the five mice treated were long term survivors, with no tumour present at 60 days after the end of treatment. Theoretically this is long enough for a single surviving cell to repopulate the tumour, so these mice would almost certainly have had normal life spans if held for a sufficiently long time.

DISCUSSION

The compounds tested did not show exceptional activity in an in vi tro cell line panel, which included the topoisomerase-II resistant derivatives of the Jurkat leukaemia which indicated that the compounds targetted both topoisomerase I and II enzymes.

However, in subcutaneous Colon 38 tumours in vivo , compound 24h achieved 80-100% cures from various dosing regimes. Colon 38 is relatively refractory to clinical antimetabolites, alkylating agents and topoisomerase-directed agents. While most of the prior art compounds in Table 3 produced growth delays, none induced cures of this tumour.

Our results show that the novel compounds described here have cytotoxic activity against animal and human tumour cell lines, and are active against transplanted tumours in mice. Thus they have potential utility as anticancer drugs .

It will be apparent to the person skilled in the art that while the invention has been described in some detail for the purposes of clarity and understanding, various modifications and alterations to the embodiments and methods described herein may be made without departing from the scope of the inventive concept disclosed in this specification.

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Claims

CLAIMS :

1. A compound of formula II:

II

in which positional numbering, where mentioned, refers to the system illustrated above, and one or more W and one or more U are attached to a ring carbon or carbons at any of positions 1-11 or to a ring nitrogen at position 7 if present, and in which:

W is C(=Q)NRCHJ(CH₂)_nR¹ where is 0 or S, R is hydrogen or a C-^ alkyl group which is optionally substituted with one or more OH or NH₂ groups,

J is H or a Ci-Cg alkyl group optionally substituted with OH, OMe, NH2, NHMe or Me2 functions, R¹ is C(=NR²)NH₂, NHC(=NR³)NH₂ or NR⁴R⁵, where each of R² and R³ are independently hydrogen or a C_]__ alkyl group which is optionally substituted with one or more OH or NH₂ groups, and

R⁴ and R⁵ are independently hydrogen or a C_1-4 alkyl group which is optionally substituted with one or more OH or NH₂ groups, or R⁴ and R⁵ together with the nitrogen atom to which they are attached form an optionally substituted saturated or unsaturated heterocyclic group; and n is an integer from 0 to 5;

X, Y, V are independently CH or N;

Z is CH₂, CH-C₁_₄ alkyl, CO, 0, S, SO, S0₂, N-C^ alkyl or NH; and 6 7 U is H, halo, OH, C0₂H, NR R , nitro, cyano, C₁_₆ alkyl, C₁_₆ haloalkyl, C₁__Q alkoxy, C₁_₆ haloalkoxy, Cι-6 aminoalkyl or ₁_₅ aminoalkoxy in which R⁶ and R⁷ have the same definitions as R⁴ and R⁵ above; or a pharmaceutically-acceptable salt, N-oxide, hydrate, solvate, pharmaceutically acceptable derivative, pro-drug, tautomer or isomer thereof, with the proviso that W at position 4 cannot be C0NH(CH2)2NMe2, CONH (CH2) 2NEt2, CONH (CH2 ) 3 Me2 , CONH(CH2)3NEt2, CONH (CH2) 2Npiperidyl , or

CONH(CH2) 2Nmorpholinyl, when X and Y are N, V is CH, U is H and Z is CO.

2. The compound as claimed in claim 1, wherein X is N.

3. The compound as claimed in claim 1 or claim 2, wherein Z is CO, CH₂, 0 or S .

4. The compound as claimed in claim 3 wherein Z is CO.

5. The compound as claimed in any one of the preceding claims, wherein V and Y are CH, N or C-W.

6. The compound as claimed in claim 5, wherein W is CONH(CH₂)₂N(CH₃)₂.

7. The compound as claimed in claim 5 wherein V and Y are CH.

8. The compound as claimed in any one of the preceding claims, wherein the compound includes one U substituent only, and the substitutent U is H, Cl, methyl or methoxy.

9. The compound as claimed in any one of the preceding claims, wherein n is 2.

8. The compound as claimed in claim 1, wherein X is N, V and Y are CH, U is 2-methyl, and Z is CO.

9. The compound as claimed in claim 1, wherein the compound is N- [2- (dimethylamino) ethyl] -2-methyl-7-oxo-7IJ- dibenz [ f, ij] isoquinoline-11-carboxamide or a pharmaceutically acceptable salt or N-oxide thereof .

10. A compound of the formula III

(III)

in which : one or more W' and U' may be present at any of positions 1-11 of the ring system illustrated above;

W' has the same definition as W in formula. II as defined in claim 1, or W' is H; ϋ', V, X¹, Y' and Z' have the same definitions as U, V, X, Y and Z in formula II as defined in claim 1;

L is a direct bond or a linker group of valency x,

L¹ is a direct bond or a linker group attached to the ring system at one or two positions; and x is an integer from 2 to 4, or a pharmaceutically-acceptable salt, N-oxide, hydrate, solvate, pharmaceutically acceptable derivative, pro-drug, tautomer or isomer thereof, with the proviso that L and L' are not both direct bonds .

11. The compound as claimed in claim 10, wherein: L' is -C(=Q' )NR⁸CHJ' (CH₂)_m-, in which: Q' has the same definition as Q in formula II as defined in claim 1;

R⁸ has the same definition as R in formula II as defined in claim 1, or R⁸ is -C ( =0 ) - or -C(=S)- in which the carbon atom of this group is attached to the ring structure at a second carbon atom on the ring;

J' has the same definition as J in the compound of formula II as defined in claim 1, and m has the same definition as n in the compound of formula II as defined in claim 1.

12. The compound as claimed in claim 11, wherein m is 1 or 2.

13. The compound as claimed in claim 11 or claim 12, wherein L is a direct bond or a nitrogen-containing linker group .

14. The compound as claimed in claim 13, wherein L is a direct bond, N or -NR⁹- [ (CHR¹⁰)_rNR^1:L] _p- wherein : p is 0 or 1; r is an integer from 0 to 5; R⁹ and R¹¹ have the same definition as R in formula II as defined in claim 1, or when p is 1, R⁹ and R' ' may together form an optionally substituted branched or straight chained alkylene; and

R¹⁰ has the same definition as J in formula II as defined in claim 1.

15. The compound as claimed in claim 14, wherein R⁹ and R¹¹ are each independently H, CH₃, or C₂H₅.

16. The compound as claimed in claim 15, wherein p is • 1, and R⁹ and R¹¹ together are -CH₂CH₂-.

17. The compound as claimed in any one of claims 14 to 16, wherein p is 1, and r is 2 or 3.

18. The compound as claimed in claim 10, wherein L is O, S, an optionally substituted Cι-__o alkylene, alkenylene or alkynylene chain, which may optionally be interspersed with one or more aryl or heterocyclic groups (which may also be substituted) and/or one or more 0, S or N atoms; or L is an optionally substituted saturated or unsaturated aryl or heterocyclic group.

19. The compound as claimed in claim 18, wherein the optional substituents in L are selected from halogen; oxy; hydroxy; alkoxy; alkyl hio; cyano; azido; acyloxy; alkyl sulphonyl ; aryl and heteroaryl .

20. The compound as claimed in any one of claims 10 to 19, wherein x is 2.

21. A pharmaceutical or veterinary composition comprising a compound of formula II or III as defined in any one of claims 1 to 20, together with a pharmaceutically or veterinarily acceptable carrier, diluent, adjuvant and/or excipient.

22. A pharmaceutical or veterinary composition as claimed in claim 21, wherein the composition comprises one or more other anti-neoplastic agents.

23. A pharmaceutical or veterinary composition .as claimed in claim 22, wherein said other anti-neoplastic agent or agents are selected from the group consisting of anti-mitotic agents, anti-metabolites, hormonal regulators, DNA-reactive agents, or biological agents or antibodies .

24. A pharmaceutical or veterinary composition as claimed in claim 23, wherein the other anti-neoplastic agent or agents are selected from taxol, 5-fluorouracil, tamoxifen, cisplatin and interleukin- 2 (IL-2).

25. A pharmaceutical or veterinary composition as claimed in any one of claims 21 to 24 comprising a second DNA-binding anti-cancer therapeutic agent for reducing toxicity to the recipient of either or both of the compound of formula II or formula III and the other anti- neoplastic agent.

26. A method of treatment of a neoplastic condition, comprising the step of administering a therapeutically effective dose of a compound of formula II or formula III as defined in any one of claims 1 to 20, or a pharmaceutically acceptable derivative thereof, pro-drug thereof, tautomer and/or isomer thereof, or a composition as claimed claim 25, to a subject in need of such treatment .

27. The method as claimed in claim 22, wherein the compound of formula II or formula III is administered separately, sequentially or in conjunction with one or more other anti-neoplastic agents.

28. The method as claimed in claim 27, wherein said other anti-neoplastic agent or agents are selected from the group consisting of anti-mitotic agents, anti- metabolites, hormonal regulators, DNA-reactive agents, or biological agents or antibodies.

29. The method as claimed in claim 28, wherein the other anti-neoplastic agent or agents are selected from taxol, 5-fluorouracil, tamoxifen, cisplatin and interleukin- 2 (IL-2).

30. The method as claimed in any one of claims 26 to 29, wherein the compound of formula II or formula III is administered separately, sequentially or in conjunction with a second DNA-binding anti-cancer therapeutic agent in order to reduce toxicity to the recipient of either or both of the compounds of formula II or formula III, or the other anti-neoplastic agent.

31. The method as claimed in any one of claims 26 to 30, wherein the compound of formula II or formula III, or the composition comprising the compound of formula II or formula III is administered separately, sequentially or in combination with one or more agents that relieve side effects caused by drug treatment.

32. The method as claimed in claim 31, wherein the agent that releives side effects is granulocyte- macrophage-colony stimulating factor (GM-CSF) , or an anti- emetic.

33. Use of a compound of formula II or formula III as defined in any one of claims 1 to 20 in the manufacture of a medicament for the treatment or prophylaxis of a neoplastic condition.

34. A compound of formula IV:

(IV) in which : the compound includes one or more U' ' groups and one or more carboxylic acid groups are attached to a ring carbon or carbons or nitrogen when present in the ring structureat positions 1 - 11; and

U", V', X'', Y" and Z" have the same definitions as U, V, X, Y and Z, respectively, in formula II as defined in claim 1.

35. A method of synthesising a compound of formula II or formula III as defined in any one of claims 1 to 20, the method comprising the step of converting the carboxylic acid of formula IV as defined in claim 34 into the target amide of formula II or formula III .

36. The method as claimed in claim 35, wherein the method involves converting the compound of formula IV into an imidazolide, and reacting the imidazolide intermediate with the appropriate amine to obtain the target amide of formula II or formula III.

37. The method as claimed in claim 35, wherein the method involves converting the carboxylic acid of formula IV into an acid halide, and reacting the acid halide with an amine to obtain the target amide of formula II or formula III.

38. The method as claimed in claim 37, wherein the carboxylic acid is reacted with thionyl chloride to be converted into the corresponding acid chloride.