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WO2004096779A1 - Quinazolinone-derivatives and their use for preparation of pharmaceutical compositions having parp enzyme inhibitory effect - Google Patents

Quinazolinone-derivatives and their use for preparation of pharmaceutical compositions having parp enzyme inhibitory effect Download PDF

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WO2004096779A1
WO2004096779A1 PCT/HU2004/000044 HU2004000044W WO2004096779A1 WO 2004096779 A1 WO2004096779 A1 WO 2004096779A1 HU 2004000044 W HU2004000044 W HU 2004000044W WO 2004096779 A1 WO2004096779 A1 WO 2004096779A1
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Balázs SÜMEGI
Kálmán HIDEG
Tamás KÁLAI
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Priority to US10/554,749 priority patent/US7994182B2/en
Priority to EP04730604A priority patent/EP1622881B1/en
Priority to SI200431482T priority patent/SI1622881T1/en
Priority to PL04730604T priority patent/PL1622881T3/en
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/04—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings directly linked by a ring-member-to-ring-member bond
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D239/00—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings
    • C07D239/70—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings condensed with carbocyclic rings or ring systems
    • C07D239/72—Quinazolines; Hydrogenated quinazolines
    • C07D239/95—Quinazolines; Hydrogenated quinazolines with hetero atoms directly attached in positions 2 and 4
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/06—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a carbon chain containing only aliphatic carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links

Definitions

  • the subject of the present invention are quinazolinone derivatives of general formula (I), that inhibit a DNA- repairing enzyme, poly(ADP-ribose) polymerase (PARP), enabling them to be used for the preparation of pharmaceutical compositions for preventing or treating illnesses where PARP-inhibition yields a beneficial effect.
  • PARP poly(ADP-ribose) polymerase
  • the PARP enzyme occurring in the eukaryotic nucleus is a DNA-binding protein, having a serious role in the genomical repair mechanism. 1,2
  • the PARP enzyme is activated directly by genotoxic agents (alkylating agents, reactive oxygen radicals, ionising radiation) or indirectly by DNA strand break occuring after the enzymatic excision of a mismatched DNA base.
  • PARP catalyses the biochemical transformation of nicotinamide adenine dinucleotide (NAD) to poly(ADP-ribose) and nicotinamide, hence the latter is one of the weak feedback inhibiting agents of the enzyme.
  • NAD nicotinamide adenine dinucleotide
  • PARP plays an important role in the genomical repair, its significant activation entails extensive ADP-ribosylation and the depletion of NAD depots leading to the decrease of the ATP-level due to resynthesising NAD, and finally, in the lack of ATP and NAD it causes failure in the mitochondrial function and ultimately cell death.
  • the present inventors have already synthesised a promising antiarrhythmic 4-quinazolinone derivative named H-2641, see formula (B) 8 . After this, its PARP inhibiting activity could be studied and compared to the same of basic quinazolinone compounds of formulae (l)-(3). We note that, according to earlier observations, substitution at position 3 of the quinazolinone ring increased the IC 50 value of the compounds, so further investigations in this direction did not seem to be promising. 6
  • the elaboration of the present invention was aimed at the preparation of new compounds containing the basic quinazoline-4(3H) ring bearing such sidechains in positions 2 or 3 that have an advantageous effect on the pharmacological properties of the molecule primarily by increasing their effect against oxidative stress due to the free radical trapping ability of the sidechains.
  • the effectiveness against oxidative stress can be an advantageous feature of the PARP inhibitors, because this way they can provide an increased protection against cell destruction caused by antiviral 11 and antitumour drugs.
  • the therapeutic application of antiviral and antitumour compounds unavoidably leads to the release of reactive oxygen free radicals.
  • the in stau nascen ⁇ capture of these radicals can moderate the side effects of these drugs more than presently used PARP inhibitors could do.
  • R 1 stands for hydrogen or a group of general formula (a) and k is 1, 2, 3 or 4, n is 0 or 1, Q stands for oxyl group (0-) or hydrogen, Ra and Re independently from each other stand for hydrogen or C w alkyl group, Rb and Rd independently from each other stand for alkyl group and the broken line stands for optional valence bond;
  • R 2 stands for a) hydrogen or C ⁇ . ⁇ alkyl group, if R 1 is other than hydrogen, and b) if R 1 is a hydrogen, then R 2 may stand for
  • Compounds of the present invention may contain one or two asymmetrical carbon atoms, when the meanings of the Ra-Rd groups are different. Present invention implicitly involves all the potential isomer forms as well.
  • the present invention includes also a process for the preparation of compounds of general formula (I) and their pharmaceutically acceptable salts - the meanings of the symbols applied in the formula are given above -, where a) in case of the synthesis of compounds of general formula (I) wherein R 1 stands for a group of general formula (a), a compound of general formula (II a ) is reacted with a compound of general formula (III a b) — the meanings of the symbols applied in the formulae are given above, furthermore, in general formula (II a ) R 2a is either hydrogen or d- ⁇ alkyl group and in general formula (I_I ab ) X is a leaving group, preferably a halogen atom — b) in case of the synthesis of compounds of general formula (I) wherein R 2 stands for a group of general formula (b), a compound of formula (I_ b _) is reacted with a compound of general formula (III ab ) — the meanings of the symbols applied in
  • reaction schemes A)-D The preparation processes of the compounds of present invention are grouped by the side chains to be built in [see reaction schemes A)-D)].
  • general synthetic procedures are given for the synthesis of each group of compounds.
  • the synthesis of compounds disclosed in the examples are shown in the reaction schemes A') - D').
  • the oxyl group of compounds prepared according to reaction schemes A), B), and D) can be reduced to hydrogen using known methods. (This transformation is shown in general example V.)
  • salts can be formed by known methods.
  • Pharmaceutically acceptable salts are obviously preferred, e.g. salts formed with hydrochloric acid, maleic acid, and ascorbic acid. If the synthesis provides not the desired salt, then the free compounds can be obtained, or it can be transformed to other type of salt by known procedures.
  • optically active compounds of general formula (I) can be prepared from the corresponding optically active starting compounds, or the end product, being in mixture form, can be separated to optically active isomers by the usual resolving procedures.
  • the present invention also includes the pharmaceutical compositions containing any of the new compounds described above and the use of the compounds for the preparation of drugs having PARP enzyme- inhibitor effects.
  • compositions embraced by the present invention preferably contain one or more quinazolinone derivatives of general formula (I) or their pharmaceutically accepted salts in 0.1 to 95 % by weight as active ingredients, beside the usual auxiliary ingredients used in pharmaceutical compositions.
  • composition embraced by present invention can occur in either solid or liquid forms, and can be used for peroral, parenteral or rectal administrations, or for local treatment.
  • Solid pharmaceutical compositions for peroral use may be powders, capsules, pills, film-coated pills, microcapsules etc, and they may contain carriers, like gelatin, sorbitol, poly(vinyl-pyrrolidone); filling materials like lactose, glucose, starch, calcium phosphate; excipients, like magnesium stearate, talcum, poly(ethylene- glycol), silicon dioxide; moisturizers, like sodium lauryl sulphate.
  • carriers like gelatin, sorbitol, poly(vinyl-pyrrolidone)
  • filling materials like lactose, glucose, starch, calcium phosphate
  • excipients like magnesium stearate, talcum, poly(ethylene- glycol), silicon dioxide
  • moisturizers like sodium lauryl sulphate.
  • Liquid pharmaceutical compositions for peroral use can be solutions, suspensions or emulsions, containing carriers like suspending agents, such as gelatin, carboxymethylcellulose; emulgeators, like sorbitane-monooleate; solvents, like water, oils, glycerol, propylene glycol, ethanol; preservatives, such as methyl or propyl esters of p- hydroxy-benzoic acid.
  • suspending agents such as gelatin, carboxymethylcellulose
  • emulgeators like sorbitane-monooleate
  • solvents like water, oils, glycerol, propylene glycol, ethanol
  • preservatives such as methyl or propyl esters of p- hydroxy-benzoic acid.
  • compositions for parenteral use usually consist of the sterile solution of the active ingredient.
  • composition embraced by the present invention is prepared by mixing one or more compounds of general formula (I) or its pharmaceutically accepted salt as active ingredient with one or more excipients, them the mixture is transformed to a pharmaceutical composition by known method.
  • the applicable methods can be obtained from scientific literature, for example the above mentioned Remington s Pharmaceutical Sciences manual.
  • Pharmaceutical compositions embraced by the present invention contain mostly one dosage unit.
  • the usual daily dose for adults is 0.1 to 5000 mg of the compound of general formula (I), or from its pharmaceutically acceptable salts, which can be administered in one or more portions.
  • the effective dosage can depend on several factors, it is prescribed and determined by the physician on the base of usual parameters (weight, age, extent and stage of the illness, etc.).
  • the resulting raw amide is then dissolved in a mixture of dioxane (40 ml) and water (40 ml). The mixture is heated to boiling, NaB0 3 x 4H 2 0 (4,6 g, 30,0 mmol) is added in small portions under 2 hours, then the reaction mixture is refluxed until all the amide is reacted (5 hours). After cooling the solvents are evaporated under reduced pressure, the residue is dissolved in CHC1 3 (20 ml), then washed with brine (10 ml). The organic phase is separated, dried (MgS0 4 ), filtered, evaporated, then the sediment is purified by flash column chromatography (CHCl 3 /Et 2 ⁇ ).
  • the protecting effect of 4-quinazolinone derivatives against cell death caused by hydrogen peroxide in WRL-68 human liver cell line was obtained from American Type Culture Collection (Rockville, MD). The one layer thick cell lines were cultured in Dulbecco's modified Eagle medium containing 1% antibiotic-antimycotic solution and 10% fetal calf serum under humid 37°C air containing 5% C0 2 . The cells were separated every three days.
  • the cells were transformed to a 96-well plate with 2.5 x 10 4 cells/well initial concentration and were cultured overnight in humid 37°C air containing 5% C0 2 .
  • the following day 0.3 mM hydrogen peroxide was added to the medium either by itself or in the presence of one of the protecting agents.
  • the medium was removed and 0.5% water-soluble mitochondria stain, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazoliurn bromide (Mi l 1 ) was added to the cells.
  • the incubation was carried on for another three hours, the medium was removed and the metabolically reduced insoluble blue formasan dye was redissolved in acidic isopropanol.
  • optical density was determined at 550 run by Anthos Labtech 2010 ELISA Reader (Wien, Austria). In every experiment at least 6 parallel samples were treated and the experiments were repeated three times. Table 1 presents theconcentrations (given in ⁇ M) where the hydrogen peroxide induced cell death was inhibited by 50%.
  • the poly(ADP-ribose) polymerase enzyme was isolated from rat liver as described earlier [22,23]. The putative inhibitory effect of 4-quinazolinone derivatives was examined in this experiment.
  • the activity of the PARP enzyme was determined in 130 ⁇ l solution containing 100 mM Tris-HCl buffer (pH 8.0), 10 mM MgCl 2 , 10% glycerol, 1.5 mM DTT, 1 mM [Adenin-2,8- 3 H]-NAD + (4,500 cpm/nmol), 10 ⁇ g activated DNA and 10 ⁇ g histone.
  • the compounds of the present invention greatly improve the status of the multiple organ failure syndrome present in sepsis (and its most severe form, septic shock) or in other diseases. Furthermore, they could be used in the treatment of renal failure, hepatic failure, neurodegenerative and inflammatory diseases, and diseases related to ischemia and reperfusion.
  • the compounds of the present invention decrease the severity of infarction, type 2 diabetes and insulin resistance as well as the metabolic X syndrome, the rheumatoid arthritis and also decrease the tumour formation in chronic inflammations.

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Abstract

The subject of the present invention are quinazoline derivatives and their pharmaceutically acceptable salts of general formula (I), that inhibit a DNA-repairing enzyme, poly(ADP-ribose) polymerase (PARP), enabling them to be used for the preparation of pharmaceutical compositions for preventing or treating illnesses where PARP-inhibition yields a beneficial effect. In general formula (I) R1 stands for either hydrogen or a group of general formula (a); R2 stands for a) hydrogen or C1-6 alkyl group, if R1 is other than hydrogen, and b) if R1 is hydrogen, then R2 may be a group of general formula (b), (c) or (d). The subject of the present invention also embraces the preparation processes of the compounds described above.

Description

Quinazolinone-derivatives and their use for preparation of pharmaceutical compositions having PARP enzyme inhibitory effect
The subject of the present invention are quinazolinone derivatives of general formula (I), that inhibit a DNA- repairing enzyme, poly(ADP-ribose) polymerase (PARP), enabling them to be used for the preparation of pharmaceutical compositions for preventing or treating illnesses where PARP-inhibition yields a beneficial effect.
The PARP enzyme occurring in the eukaryotic nucleus is a DNA-binding protein, having a serious role in the genomical repair mechanism.1,2 The PARP enzyme is activated directly by genotoxic agents (alkylating agents, reactive oxygen radicals, ionising radiation) or indirectly by DNA strand break occuring after the enzymatic excision of a mismatched DNA base. PARP catalyses the biochemical transformation of nicotinamide adenine dinucleotide (NAD) to poly(ADP-ribose) and nicotinamide, hence the latter is one of the weak feedback inhibiting agents of the enzyme. Although PARP plays an important role in the genomical repair, its significant activation entails extensive ADP-ribosylation and the depletion of NAD depots leading to the decrease of the ATP-level due to resynthesising NAD, and finally, in the lack of ATP and NAD it causes failure in the mitochondrial function and ultimately cell death.
The structural formulas of the first PARP inhibitors showed homology to nicotinamide and benzamide analogues.3 Recent research was aimed at the synthesis of more effective and more selective PARP inhibitors fitted to the crystal structure of PARP's catalytic domain, leading to the discovery of polycyclic amides and lactams.4'5 Presently, the most effective compound is 8-hydroxy-2-methyl-quinazoline-4(3H)-one [NU-1025, see formula (A), IC50=400 nM].6'7 Later, new tricyclic inhibitors were prepared starting from this compound.
The present inventors have already synthesised a promising antiarrhythmic 4-quinazolinone derivative named H-2641, see formula (B)8. After this, its PARP inhibiting activity could be studied and compared to the same of basic quinazolinone compounds of formulae (l)-(3). We note that, according to earlier observations, substitution at position 3 of the quinazolinone ring increased the IC50 value of the compounds, so further investigations in this direction did not seem to be promising.6
The elaboration of the present invention was aimed at the preparation of new compounds containing the basic quinazoline-4(3H) ring bearing such sidechains in positions 2 or 3 that have an advantageous effect on the pharmacological properties of the molecule primarily by increasing their effect against oxidative stress due to the free radical trapping ability of the sidechains.
The effectiveness against oxidative stress can be an advantageous feature of the PARP inhibitors, because this way they can provide an increased protection against cell destruction caused by antiviral11 and antitumour drugs. The therapeutic application of antiviral and antitumour compounds unavoidably leads to the release of reactive oxygen free radicals. The in stau nascenώ capture of these radicals can moderate the side effects of these drugs more than presently used PARP inhibitors could do.
The present invention provides the compounds of general formula (I) and their pharmaceutically acceptable salts, wherein R1 stands for hydrogen or a group of general formula (a) and k is 1, 2, 3 or 4, n is 0 or 1, Q stands for oxyl group (0-) or hydrogen, Ra and Re independently from each other stand for hydrogen or Cw alkyl group, Rb and Rd independently from each other stand for alkyl group and the broken line stands for optional valence bond; R2 stands for a) hydrogen or Cι.β alkyl group, if R1 is other than hydrogen, and b) if R1 is a hydrogen, then R2 may stand for
(i) a group of general formula (b), wherein the meanings of k, n, Q, Ra, Rb, Re, Rd and the broken line are as given above, or
(ii) a group of general formula (c), wherein k is 1, 2 or 3; R3 and R4 independently from each other stand for Cι.6 alkyl group, or together with the attached nitrogen form a group of general formula (e), wherein p is 0 or 1 and R'a, R'b, R'c es R'd independently from each other stand for hydrogen or Cι.6 alkyl group, or iii) a group of general formula (d), wherein the meanings of n, Q, Ra, Rb, Re, Rd the broken line are as given above and m is 0, 1, 2 or 3. The advantageous groups of compounds are determined in the subclaims attached.
The following compounds seemed to be extremely advantageous: (15), (17a), (17b), (17c), (17d), (23) and (24) [compounds can be assigned to these codes in the examples and in the reaction schemes A')-Dλ) ]. The compounds (17a) and (17d) proved to be the most advantageous ones.
Compounds of the present invention may contain one or two asymmetrical carbon atoms, when the meanings of the Ra-Rd groups are different. Present invention implicitly involves all the potential isomer forms as well.
The present invention includes also a process for the preparation of compounds of general formula (I) and their pharmaceutically acceptable salts - the meanings of the symbols applied in the formula are given above -, where a) in case of the synthesis of compounds of general formula (I) wherein R1 stands for a group of general formula (a), a compound of general formula (IIa) is reacted with a compound of general formula (IIIab) — the meanings of the symbols applied in the formulae are given above, furthermore, in general formula (IIa) R2a is either hydrogen or d-β alkyl group and in general formula (I_Iab) X is a leaving group, preferably a halogen atom — b) in case of the synthesis of compounds of general formula (I) wherein R2 stands for a group of general formula (b), a compound of formula (I_b_) is reacted with a compound of general formula (IIIab) — the meanings of the symbols applied in formula (IIIab) are given above, and X is a leaving group, preferably a halogen atom -, c) in case of the synthesis of compounds of general formula (I) wherein R2 stands for a group of general formula (c), a compound of formula (II _) is reacted with a compound of general formula (III-) — the meanings of the symbols applied in formula (iπo) are given above, and X is a leaving group, preferably a halogen — d) in case of the synthesis of compounds of general formula (I) wherein R2 stands for a group of general formula (d), a compound of formula (IId) is reacted with a compound of general formula (Ilia) — the meanings of the symbols applied in formula (Hid) are given above, and X is a leaving group, preferably a halogen — and in case of the synthesis of compounds containing hydrogen in the position of group Q, the compound of general formula (I) containing oxylgroup (0-) in the position of group Q obtained by any of the processes given above is reduced, and, if desired, the compound obtained by any of the processes given above is transformed to a pharmaceutically acceptable salt, or the compound obtained in salt form is transformed into another salt, or the free compound is released from its salt form.
The preparation processes of the compounds of present invention are grouped by the side chains to be built in [see reaction schemes A)-D)]. In the examples general synthetic procedures are given for the synthesis of each group of compounds. The synthesis of compounds disclosed in the examples are shown in the reaction schemes A') - D'). Optionally, the oxyl group of compounds prepared according to reaction schemes A), B), and D) can be reduced to hydrogen using known methods. (This transformation is shown in general example V.)
Here we mention that in the reactions shown in schemes A) - D) the presence of such a starting compound is compulsory, that contains a leaving group marked by X. This can be a halogen atom, preferably bromine or chlorine, or a group of general formula -OSO2R, where R stands for e.g. methyl- ethyl- or 4-methyl-phenyl group. Of course, other usual protecting groups can be utilised. (See for example T.W. Greene, P.G.M. Wuts: Protective Groups in Organic Synthesis, 3rd edition, Hardcover 1999.)
From the compounds of the present invention salts can be formed by known methods. Pharmaceutically acceptable salts are obviously preferred, e.g. salts formed with hydrochloric acid, maleic acid, and ascorbic acid. If the synthesis provides not the desired salt, then the free compounds can be obtained, or it can be transformed to other type of salt by known procedures.
The optically active compounds of general formula (I) can be prepared from the corresponding optically active starting compounds, or the end product, being in mixture form, can be separated to optically active isomers by the usual resolving procedures.
The present invention also includes the pharmaceutical compositions containing any of the new compounds described above and the use of the compounds for the preparation of drugs having PARP enzyme- inhibitor effects.
Pharmaceutical compositions embraced by the present invention preferably contain one or more quinazolinone derivatives of general formula (I) or their pharmaceutically accepted salts in 0.1 to 95 % by weight as active ingredients, beside the usual auxiliary ingredients used in pharmaceutical compositions.
The pharmaceutical composition embraced by present invention can occur in either solid or liquid forms, and can be used for peroral, parenteral or rectal administrations, or for local treatment.
Solid pharmaceutical compositions for peroral use may be powders, capsules, pills, film-coated pills, microcapsules etc, and they may contain carriers, like gelatin, sorbitol, poly(vinyl-pyrrolidone); filling materials like lactose, glucose, starch, calcium phosphate; excipients, like magnesium stearate, talcum, poly(ethylene- glycol), silicon dioxide; moisturizers, like sodium lauryl sulphate.
Liquid pharmaceutical compositions for peroral use can be solutions, suspensions or emulsions, containing carriers like suspending agents, such as gelatin, carboxymethylcellulose; emulgeators, like sorbitane-monooleate; solvents, like water, oils, glycerol, propylene glycol, ethanol; preservatives, such as methyl or propyl esters of p- hydroxy-benzoic acid.
Pharmaceutical compositions for parenteral use usually consist of the sterile solution of the active ingredient.
The forms of administration mentioned above along with other foπns are well known, see manual: Remington's Pharmaceutical Sciences 18th edition, Mack Publishing Co., Easton, USA, (1990). Pharmaceutical composition embraced by the present invention is prepared by mixing one or more compounds of general formula (I) or its pharmaceutically accepted salt as active ingredient with one or more excipients, them the mixture is transformed to a pharmaceutical composition by known method. The applicable methods can be obtained from scientific literature, for example the above mentioned Remington s Pharmaceutical Sciences manual. Pharmaceutical compositions embraced by the present invention contain mostly one dosage unit. The usual daily dose for adults is 0.1 to 5000 mg of the compound of general formula (I), or from its pharmaceutically acceptable salts, which can be administered in one or more portions. The effective dosage can depend on several factors, it is prescribed and determined by the physician on the base of usual parameters (weight, age, extent and stage of the illness, etc.).
SYNTHETIC EXAMPLES
Materials and Methods
Melting points were determined with a Boetius Micro Melting Point apparatus and are uncorrected. Elemental analyses (C, H, N, S) were performed on Fisons EA 1110 CHNS elemental analyser. The IR spectra were consistent with the assigned structure in each case. Mass spectra were recorded on a VG TRIO-2 instrument in the El mode. !H NMR spectra were recorded with Varian Unity Inova 400 WB spectrometer; chemical shifts were referenced to TMS. Flash column chromatography was performed on Merck Kieselgel 60 (0.040-0.063 mm). The HPLC analyses were performed on an HPllOO instrument using a Hypersyl BDS-C18 column with UV detection at 250 nm. Qualitative TLC was carried out on commercially available plates (20 x 20 x 0.02 cm) coated with Merck Kieselgel GF254. Compounds with general formulae (1), (2) and (3), and 2-dimethylamino-l- ethylchloride, 3-dimethylamino-l-propylchloride, l-(2-chloroethyl)piperidine and 2-aminobenzonitrile were purchased from Aldrich, the compound H-26418 with structural formula (B), and compounds with formula (4)12, (5)13, (6)14, (7)16, (16a-d)17, (19)19 and (20)19 were prepared according to published procedures.
We note here that the compound of formula (1) and the compound of formula (1') are tautomers, thus the two structural formulas describe the same compound. In the present description the oxo form is used uniformly in all formulas containing this structural unit without any theoretical consideration.
I. General procedure according to reaction scheme A) [the structure of the referred compounds can be identified in reaction scheme A')]
A solution of 2-methylquinazolin-4(3H)one (2) (1,60 g, 10,0 mmol), the corresponding compound of formula (6) or (7) carrying an allyl bromide part (10,0 mmol) and K2C03 (1,38 g, 10,0 mmol) in DMF (15 ml) is stirred for 6 hours at 90°C. The solvent is evaporated, the residue is dissolved in CHC13 (30 ml) and washed with brine (10 ml). The organic phase is separated, dried (MgS04), filtered, evaporated, then the solid residue is purified by flash column chromatography (hexane EtOAc).
Example 1/1.
3-[(l-Oxyl-2,2,5,5-tetiamethyl-2,5-dihydro-lH-pyrrol-3-yl)methyl]-2-metlιylquinazolin-4(3H)-one [compound of formula (8)]:
Yield: 1.77 g, 57% (yellow substance).
Melting point: 110-112°C.
Calculated composition: CΛNaOz: C 68.97; H 7.40; N 13.41.
Found composition: C 68.71; H 7.39; N 13.59.
IR (nujol) v: 1620, 1585, 1570 cm"1. MS (m z, %): 312 (M", 6), 282 (21), 138 (75), 122 (100).
II. General procedure according to reaction scheme B) [the structure of the referred compounds can be identified in reaction scheme B')]
A solution of 2-mercaptoquinazolin-4(3H)one (3) (1,78 g, 10,0 mmol), the corresponding compound of formula (6) or (7) carrying an allyl bromide part (10,0 mmol) and K2C03 (1,38 g, 10,0 mmol) in DMF (25 ml) is stirred for 6 hours at 90°C. The solvent is evaporated, the residue is dissolved in CHC13 (30 ml), and washed with brine (10 ml). The organic phase is separated, dried (MgS04), filtered, evaporated, then the solid residue is purified by flash column chromatography (CHC13 Et20).
Example II/l.
2-{[(l-Oxyl-2,2,5,5-teframemyl-2,5-d y(ko-m-pyrrol-3-yl)methyl]thio}quinazolin-4(3H)-one (12)
Yield: 1.91 g (58%).
Melting point: 138-140°C.
Calculated composition: C,7H2oN302S: C 61.79; H 6.11; N 12.73; S 9.68.
Found composition: C 61.92; H 6.30; N 12.79; S 9.60.
IR (nujol) v: 1675, 1605, 1585, 1560 cm"1.
MS (m/z, %): 330 (M", 5), 300 (20), 178 (54), 122 (100).
Example II/2
2-{[(l-Oxyl-2,2,6,6-teframethyl-l,2,3,6-tefrahydropyridin-4-yl)methyl]thio}quinazolin-4(3H)-one (14)
Yield: 1.51 g (44%).
Melting point: 79-82°C.
Calculated composition: Cι8H22N302S: C 62.76; H 6.44; N 12.21; S 9.29.
Found composition: C 62.72; H 6.31; N 12.34; S 9.44.
IR (nujol) v: 1670, 1610, 1585, 1560 cm"1.
MS (m/z, %): 344 (M+, 6), 314 (35), 136 (61), 121 (100).
HI. General procedure according to reaction scheme C) [the structure of the referred compounds can be identified in reaction scheme C')]
A solution of 2-mercaptoquinazolin-4(3H)-one (3) (1,78 g, 10,0 mmol), the corresponding compound of formula (16a), (16b), (16c) or (16d) carrying an alkyl chloride part (10,0 mmol) and K2C03 (2,26 g, 10,0 mmol) in DMF (25 ml) is stirred for 6 hours at 90°C. The solvent is evaporated, the solid residue is dissolved in CHC13 (30 ml), and washed with brine (10 ml). The organic phase is separated, dried (MgS0 ), filtered, evaporated, then the solid residue is purified by flash column chromatography (CHC13/Et20).
Example HI/1.
2-{ [2-(Dimethylamino)etlιyl]thio}quinazolin-4(3H)-one (17a)
Yield: 1.29 g (52%).
Melting point: 155-157°C.
Calculated composition: Cι2H15N3OS: C 57.81; H 6.06; N 16.85; S 12.86.
Found composition: C 57.92; H 6.11; N 16.80; S 12.70.
IR (nujol) v: 1665, 1630, 1570 cm"1. 'H NMR (CDC13) δH: 8.17 (1H, d), 7.65 (1H, td), 7.55 (1H, d), 7.35 (1H, t), 3.14 (2H, m), 2.92 (2H, m) and 2.48 (6H, s).
MS (m/z, %): 249 ( *, 2), 234 (16), 162 (44), 71 (100).
Example HI/2.
2-{[3-(Dimethylamino)propyl]thio}quinazolin-4(3H)-one (17b)
Yield: 1.18 g (45%).
Melting point: 93-95°C.
Calculated composition: Cι3HI7N3OS: C 59.29; H 6.51; N 15.96; S 12.17.
Found composition: C 59.70; H 6.51; N 16.02; S 12.02.
IR (nujol) v: 1665, 1630, 1570 cm"1.
"H NMR (CDC13) δH: 8.16 (1H, d), 7.65 (1H, t), 7.54 (1H, d), 7.34 (1H, t), 3.23 (2H, t), 2.68 (2H, t), 2.38 (6H, s) and 1.96 (2H, ).
MS (m/z, %): 263 (M", 1), 205 (2), 85 (100), 58 (65).
Example III/3.
2-[(2-Piperidin-l-ylethyl)tlιio]quinazolin-4(3H)-one (17c)
Yield: 2.02 g (70%).
Melting point: 131-133°C.
Calculated composition: Cι5H,9N3OS: C 62.26; H 6.62; N 14.53; S 11.06.
Found composition: C 62.74; H 6.51; N 14.59; S 10.99.
IR (nujol) v: 1660, 1640, 1575, 1550 cm"1.
•H NMR (DMSO-de) δH: 12.82 (1H, bs), 8.02 (1H, d), 7.74 (1H, t), 7.49 (1H, d) 7.40 (1H, t), 3.34 (2H, t), 2.64 (2H, t), 2.46 (4H, m), 1.53 (4H, m) and 1.39 (2H, m).
MS (m/z, %): 289 (M", 1), 178 (3), 111 (100), 98 (79).
Example III/4.
2-{[2-(2,2,6,6-Tettame lpiperidin-l-yl)ethyl]t o}quinazolin-4(3H)-one (17d)
Yield: 1.34 g (39%).
Melting point: 221°C.
Calculated composition: C]9H27N3OS: C 66.05; H 7.88; N 12.17; S 9.26.
Found composition: C 66.15; H 7.85; N 12.29; S 9.10.
IR (nujol) v: 1670, 1640, 1575, 1560 cm"1.
:H NMR (DMSO-d6) δH: 12.54 (1H, bs), 8.01 (1H, d), 7.74 (1H, t), 7.41 (1H, d), 7.38 (1H, t), 3.07 (2H, m), 2.77 (2H, m), 1.49 (2H, bs), 1.39 (4H, bs) and 1.10 (12H, s).
MS (m/z, %): 345 (M1", <1), 330 (1), 205 (16), 154 (100).
IV. General procedure according to reaction scheme D) [the structure of the referred compounds can be identified in reaction scheme D')]
Compound (19) or (20) (10.0 mmol) dissolved in freshly prepared CH2C12 (10 ml) is added dropwise at 0°C to the stirred solution of 2-amino-benzonitrile (1,18 g, 10,0 mmol) and Et3N (1,11 g, 11,0 mmol) dissolved in CH2C12 (20 ml), then the mixed solution is stirred for another hour. The organic phase is washed with 10% aqueous K2C03 (15 ml), 5% H2S04 (10 ml) in water and brine (15 ml), then the organic phase is separated, dried (MgS04), filtered and evaporated. The resulting raw amide is then dissolved in a mixture of dioxane (40 ml) and water (40 ml). The mixture is heated to boiling, NaB03 x 4H20 (4,6 g, 30,0 mmol) is added in small portions under 2 hours, then the reaction mixture is refluxed until all the amide is reacted (5 hours). After cooling the solvents are evaporated under reduced pressure, the residue is dissolved in CHC13 (20 ml), then washed with brine (10 ml). The organic phase is separated, dried (MgS04), filtered, evaporated, then the sediment is purified by flash column chromatography (CHCl3/Et2θ).
Example IV/1.
2-(l-Oxyl-2,2,5,5-tetramethyl-2,5-dihydro-lH-pyrrol-3-yl)quinazolin-4(3H)-one (21)
Yield: 1.22 g (43%) (yellow substance).
Melting point: 224-226°C.
Calculated composition: Ci6Hι8N302: C 67.57; H 6.38; N 14.78.
Found composition: C 67.41; H 6.51; N 14.66.
IR (nujol) v: 1650, 1600, 1570 cm"1.
MS (m z, %): 284 (M1", 6), 270 (15), 254 (18), 41 (100).
Example IV/2.
2-(l-Oxyl-2,2,6,6-tetramethyl-l,2,3,6-tetraliydropyridin-4-yl)quinazolin-4(3H)-one (23)
Yield: 1.04 g (35%) (orange colour).
Melting point: 226-228°C.
Calculated composition: C17H20N3O2: C 68.42; H 6.76; N 14.09.
Found composition: C 68.41; H 6.83; N 13.94.
IR (nujol) v: 1670, 1600, 1580 cm"1. MS (m/z, %): 298 (W , 10), 160 (23), 138 (48), 122 (100).
V. General synthetic procedure for reducing N-oxyl radical to a secondary amine Iron powder (1.40 g, 25 mmol) is added to the solution of the compound of general formula (I) carrying N-oxyl group in position of Q (5.0 mmol) in glacial acetic acid (8 ml), and the mixture is heated to 70°C, until the reaction begins. The mixture is stirred for an hour, then diluted with water (20 ml), the solution is decanted from the remaining iron powder, the decanted aqueous solution is alkalified with solid K C03. The mixture was extracted with CHC13 (3 x 15 mL), dried (MgS04), filtered, evaporated and after chromatographic purification (CHCl3/MeOH) the resulting amide is obtained as a white or off-white solid material. Example V/l.
3-[(2,2,5,5-Tetramethyl-2,5-dihydro-lH-pyrrol-3-yl)methyl]-2-methylquinazolin-4(3H)-one (9) Yield: 623 mg (42%). Melting point: 95-97°C.
Calculated composition: Cι8H23N30: C 72.68; H 7.80; N 14.14. Found composition: C 72.57; H 7.82; N 14.21. IR (nujol) v: 3260, 1670, 1630, 1570, 1560 cm"1.
*H NMR PMSO-dβ) δH: 8.11 (1H, d), 7.90 (1H, t), 7.82 (1H, d), 7.61 (1H, t), 5.79 (1H, s), 5.11 (2H, s), 2.62 (3H, s), 1.29 (6H, s) and 1.18 (6H, s). MS (m z, %): 297 (M , 1), 282 (26), 160 (3), 122 (100). Example V/2. 2-{[(2,2,5,5-Tetrametlιyl-2,5-dihydro-lH-pyrrol-3-yl)methyl]tlιio}quinazolin-4(3H)-one (13) Yield: 582 mg (37%).
Melting point: 150-151°C.
Calculated composition: C H2!N3OS: C 64.73; H 6.72; N 13.33; S 10.15.
Found composition: C 64.76; H 6.63; N 13.23; S 10.02.
IR (nujol) v: 3250, 1675, 1605, 1585, 1560 cm"1.
'H NMR (CDCl3) δH: 8.16 (IH, d), 7.65 (IH, t), 7.52 (IH, d), 7.33 (IH, t), 5.60 (IH, s), 3.91 (2H, s), 1.29 (6H, s) and 1.15 (6H, s). MS (m z, %): 315 (M", 1), 300 (26), 191 (14), 122 (100).
Example V/3.
2-{[(2,2,6,6-Teframetiιyl ,2,3,6-tefrahydropyridin-4-yl)methyl]thio}quinazolin-4(3H)-one (15)
Yield: 789 mg (48%).
Melting point: 180-182°C.
Calculated composition: C18H23N3OS: C 65.62; H 7.04; N 12.76; S 9.71.
Found composition: C 65.72; H 7.03; N 12.59; S 9.81.
IR (nujol) v: 3260, 1670, 1610, 1585, 1570 cm"1.
'H NMR (CDC13) δH: 8.21 (IH, dd), 7.70 (IH, td), 7.56 (IH, d), 7.38 (IH, t), 5.74 (IH, s), 3.96 (2H, s), 2.03 (2H, s), 1.21 (6H, s) and 1.19 (6H, s). MS (m/z, %): 329 (M÷, 2), 314 (23), 136 (100), 121 (92). Example V/4.
2-(2,2,5,5-Tetramethyl-2,5-dihydro-lH-pyrrol-3-yl)quinazolin-4(3H)-one (22) Yield: 914 mg (68%). Melting point: 254-256°C.
Calculated composition: C16Hι9N30: C 71.33; H 7.11; N 15.61. Found composition: C 71.38; H 7.06; N 15.60. IR (nujol) v: 3240, 1655, 1600, 1570 cm"1.
]H NMR (DMSO-d6) δH: 12.01 (IH, bs), 8.10 (IH, dd), 7.79 (IH, td), 7.62 (IH, d), 7.49 (IH, td), 6.96 (IH, s), 1.51 (6H, s) and 1.24 (6H, s). MS (m/z, %): 269 (M1", 1), 254 (24), 137 (40), 108 (100). Example V/5.
2-(2,2,6,6-Teframethyl-l,2,3,6-tetrahydropyridin-4-yl)quinazolin-4(3H)-one (24) Yield: 816 mg (55%). Melting point: 265-268°C.
Calculated composition: CπH21N30: C 72.04; H 7.47; N 14.84. Found composition: C 72.21; H 7.53; N 14.64. IR (nujol) v: 3260, 1670, 1600, 1580 cm"1.
!H NMR (DMSO-d6) δH: 12.01 (IH, bs), 8.10 (IH, d), 7.78 (IH, t), 7.64 (IH, d), 7.47 (IH, t), 6.89 (IH, s), 2.35 (2H, s), 1.23 (6H, s) and 1.13 (6H, s). MS (m/z, %): 283 (M+, 8), 268 (75), 138 (52), 122 (100). Pharmacological studies
The protecting effect of 4-quinazolinone derivatives against cell death caused by hydrogen peroxide in WRL-68 human liver cell line. Cell culture The human WRL-68 cell line was obtained from American Type Culture Collection (Rockville, MD). The one layer thick cell lines were cultured in Dulbecco's modified Eagle medium containing 1% antibiotic-antimycotic solution and 10% fetal calf serum under humid 37°C air containing 5% C02. The cells were separated every three days.
The detection of cell survival
The cells were transformed to a 96-well plate with 2.5 x 104 cells/well initial concentration and were cultured overnight in humid 37°C air containing 5% C02. The following day 0.3 mM hydrogen peroxide was added to the medium either by itself or in the presence of one of the protecting agents. Three hours later the medium was removed and 0.5% water-soluble mitochondria stain, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazoliurn bromide (Mi l 1) was added to the cells. The incubation was carried on for another three hours, the medium was removed and the metabolically reduced insoluble blue formasan dye was redissolved in acidic isopropanol. The optical density was determined at 550 run by Anthos Labtech 2010 ELISA Reader (Wien, Austria). In every experiment at least 6 parallel samples were treated and the experiments were repeated three times. Table 1 presents theconcentrations (given in μM) where the hydrogen peroxide induced cell death was inhibited by 50%.
The in vitro inhibitory effect of 4-quinazolinone derivatives on PARP enzyme
The poly(ADP-ribose) polymerase enzyme was isolated from rat liver as described earlier [22,23]. The putative inhibitory effect of 4-quinazolinone derivatives was examined in this experiment. The activity of the PARP enzyme was determined in 130 μl solution containing 100 mM Tris-HCl buffer (pH 8.0), 10 mM MgCl2, 10% glycerol, 1.5 mM DTT, 1 mM [Adenin-2,8-3H]-NAD+ (4,500 cpm/nmol), 10 μg activated DNA and 10 μg histone. With or without the presence of 4-quinazolinone derivatives, the samples were incubated for 15 minutes, then the reactions were brought to stop by the addition of 8% trichloroacetic acid. After the addition of 0.5 mg albumin the samples were allowed to precipitate for 20 minutes on ice, then the insoluble substances were filtered on a glass filter, collected and washed five times with 5% perchloric acid. The protein-bound radioactivity was measured by LS-200 Beckman scintillation counter, the data is presented in μM in Table 2.
Table 1. The inhibitory effect of the 4-quinazolinone derivatives (H-2641, 1-24) against hydrogen peroxide induced cell death in WRL-68 human liver cell line.
Figure imgf000010_0001
Table 2. The in vitro inhibitory effect of the substances 17a-d on PARP enzyme
Figure imgf000011_0001
In summary, a series of 4(3H)-quinazolinone derivatives were synthesised by alkylating the original cyclic compounds and by nitrile acylation of antranilic acid, which was followed by the closure of the quinazoline ring. 4-(3H)-quinazolinone derivatives containing S-ethyl sidechains substituted with tertiary amines on the carbon of position 2 proved to be the most effective PARP inhibitors among the synthesised and examined compounds. The effect of nitroxide or amino precursor forms was smaller than that of the tertiary amines, but similar to each other, nevertheless having free radical scavenger ability they might be favourable according to previous results.9'10'21
These data show that the compounds of the present invention significantly decrease the effects of oxidative stress and inhibit poly(ADP-ribose) polymerase. The protecting effect of the above mentioned compounds mainly concern their effect on poly(ADP-ribose)-polymerase (PARP) inhibition, thus liaving a protecting role in every kind of pathological processes, where the protecting effect of PARP inhibition has a great importance.
Consequently, the compounds of the present invention greatly improve the status of the multiple organ failure syndrome present in sepsis (and its most severe form, septic shock) or in other diseases. Furthermore, they could be used in the treatment of renal failure, hepatic failure, neurodegenerative and inflammatory diseases, and diseases related to ischemia and reperfusion.
Thus, the compounds of the present invention decrease the severity of infarction, type 2 diabetes and insulin resistance as well as the metabolic X syndrome, the rheumatoid arthritis and also decrease the tumour formation in chronic inflammations.
References
1. Szabό, Cs. (Ed.) Cell Death, CRC Press, Boca Raton, 2000.
2. de Murcia, G. (Ed.) From DNA Damage and Stress Signalling to Cell Death - Poly ADP- Ribosylating Reactions, Oxford University Press, Oxford, 2000.
3. Halmosi, R.; Berente, Z.; Osz, E.; Tόth, K.; Literati-Nagy, P.; Sϋmegi, B. Mol. Phannacol. 2001, 59, 1497.
4. Steinliagen, H; Gerisch, M.; Mittendorf, J.; Schlemmer, K-H.; Albrecht, B. Bioorg. Med. Chem. Lett. 2002, 12, 3187.
5. Li, H-J; Zhang, J. IDrugs 2001, 4, 804.
6. Griffin, J.; Srinivasan, S.; Bowman, K.; Kalvert, H. A.; Curtin, N. J.; Newel, D. R; Pemberton, L. C; Golding, B. T. J. Med. Chem. 1998, 41, 5247.
7. Tentori, L.; Leonetti, C; Scarsella, M.; d'Amati, G.; Portarena, I.; Zupi, G; Bonmassar, E.; Graziani, G. Blood 2002, 99, 2241.
8. Hankovszky, H. O.; Hideg, K.; Bόdi, L; Frank, L. J. Med. Chem. 1986, 29, 1138.
9. Twomey, P.; Taira, J.; DeGraff, W.; Mitchell, J. B.; Russo, A.; Krishna, M. C; Hankovszky, H. O.; Frank, L.; Hideg, K. Free Rad. Biol. & Med. 1997, 22, 909.
10. Krishna, M. C; Degraff, W.; Hankovszky, H. O.; Sar, P. C; Kalai, T.; Jekδ, J.; Russo, A.; Mitchell, J. B.; Hideg, K. J. Med. Chem. 1998, 41, 3477. L 0. Ut). ZUU4
11
11. Szabados, E.; Fischer, G. M.; Tόth, K.; Csete, B.; Nemeti, B.; Trombitas, K.; Habon, T.; Endrei, D.; Sϋmegi, B. Free Rad. Biol. & Med. 1999, 26, 309.
12. Shakhadoyatov, K. M.; Yangibaev, S.; Yun, L. M.; Kadirov, C. S. Chem. Natl. Comp. 1982, 18, 106.
13. Liu, K. C; Hsu, L. Y. Arch. Pharm. 1985, 318, 502.
14. Hankovszky, H. O.; Hideg, K.; Lex, L.; Kulcsar, G. Synthesis 1980, 914.
15. Sar, P. C; Kalai, T.; Baracz, M. N.; Jerkovich, Gy.; Hideg, K. Synth. Commun. 1995, 25, 2929.
16. Hideg, K.; Csekδ, J.; Hankovszky, H. O.; Sohar, P. Can. J. Chem. 1986, 64, 1482.
17. Robertson, J.; Biel, J. H.; DiPerro, F. J. Med. Chem. 1963, 6, 381.
18. Huber, I.; Szabό, A.; Fiilop, F.; Bernath, G; Sohar, P. Tetrahedron 1992, 48, 4949.
19. Rozantsev E. G. Free Nitroxyl Radicals, Plenum Press, New York, 1970.
20. Baudoin, B.; Ribeill, B.; Vicker, N. Synth. Commun. 1993, 23, 2833.
21. Samuni, A.; Goldstein, S.; Russo, A.; Mitchell, J. B.; Krishna, M. C; Neta, P. J. Am. Chem. Soc. 2002, 124, 8719.
22. Shah, G. M.; Poirier, D; Duchaine, C; Brochu, G.; Desnoyers, S; Lagueux, J.; Verreult, A.; Hoflack, J. C; Kirkland, J. B.; Poirier, G. G. Anal. Biochem. 1995, 227, 1.
23. Szabados, E.; Literati-Nagy, P.; Farkas, B.; Sϋmegi, B. Biochem. Pharmacol. 2000, 59, 937.
24. Skahtzky, D. J.; Marakovits, J. T.; Maegley, K. A.; Ekker, A.; Yu, X-H.; Hostomsky, Z.; Webber, S. E.; Eastman, B. W.; Almassy, R.; Li, J.; Cuirtin, N. J.; Newell, D. R.; Calvert, A. H.; Griffm, R. J.; Golding, B. T. J. Med. Chem, 2003, 46, 210-213.
Summary of drawings
Figure imgf000012_0001
x PTi λ mm

Claims

ru ι/πu<-uu '. / u u u u •* *. 12Claims
1. Compounds of general formula (I) and their pharmaceutically acceptable salts thereof, wherein R1 stands for hydrogen or a group of general formula (a) and k is 1, 2, 3 or 4, n is 0 or 1,
Q stands for oxyl group (0 ) or hydrogen,
Ra and Re independently from each other stand for hydrogen or G..6 alkyl group, Rb and Rd independently from each other stand for
Figure imgf000013_0001
alkyl group and the broken line stands for optional valence bond; R2 stands for a) hydrogen or Cι-6 alkyl group, if R1 is other than hydrogen, and b) if R1 is a hydrogen, then R2 may stand for
(i) a group of general formula (b), wherein the meanings of k, n, Q, Ra, Rb, Re, Rd and the broken line are as given above, or
(ii) a group of general formula (c), wherein k is 1, 2 or 3; R3 and R4 independently from each other stand for Cι.6 alkyl group, or together with the attached nitrogen form a group of general formula (e), wherein p is 0 or 1 and R'a, R'b, R'c es R'd independently from each other stand for hydrogen or Ci-e alkyl group, or iii) a group of general formula (d), wherein the meanings of n, Q, Ra, Rb, Re, Rd the broken line are as given above and m is 0, 1, 2 or 3.
2. The compounds of general formula (I) according to claim 1, where R2 stands for a group of general formula (a), wherein k=l, n=0, Q=hydrogen, furtliermore Ra, Rb, Re and Rd stand for C]-3 alkyl group, preferably methyl group, and the broken line stands for a valence bond.
3. The compounds of general formula (I) according to claim 1, where R2 stands for a group of general formula (b), where k=l, n=0 or 1, furthermore Ra, Rb, Re and Rd stand for Cι.3 alkyl group, preferably methyl group. and the broken line stands for a valence bond.
4. The compounds of general formula (I) according to claim 1, where R2 stands for a group of general formula (b), where k=l, n=l, Q=hydrogen, furthermore Ra, Rb, Re and Rd stand for Cι-3 alkyl group, preferably methyl group, and the broken line stands for a valence bond.
5. The compounds of general formula (I) according to claim 1, where R2 stands for a group of general formula (c), where k=2 or 3, furthermore R3 and R4 stand for C]-3 alkyl group, preferably methyl group.
6. The compounds of general formula (I) according to claim 1, where R2 stands for a group of general formula (c), where k=2 or 3, furthermore R3 and R4 together stand for a group with structural formula (e), where p=l and R'a, R'b, R'c and R'd stand for a hydrogen atom.
7. The compounds of general formula (I) according to claim 1, where R2 stands for a group of general formula (c), where k=2 or 3, furthermore R3 and R4 together stand for a group with structural formula (e), where p=l and R'a, R'b, R'c and R'd stand for Cι-3 alkyl group, preferably methyl group.
8. The compounds of general formula (I) according to claim 1, where R2 stands for a group of general formula (d), where k=0, n=0 or 1, furthermore Ra, Rb, Re and Rd stand for C]-3 alkyl group, preferably methyl group, and the broken line stands for a valence bond.
9. The compounds of general formula (I) according to claim 1, where R2 stands for a group of general formula (d), where k=0, n=l, furthermore Ra, Rb, Re and Rd stand for Cι.3 alkyl group, preferably methyl group, and the broken line stands for a valence bond.
10. A process for the preparation of compounds of general formula (I) and pharmaceutically acceptable salts thereof- the meanings of the symbols applied in the formula are given in claim 1 — , characterised by that a) in case of the synthesis of compounds of general formula (I) wherein R1 stands for a group of general formula (a), a compound of general formula (IIa) is reacted with a compound of general formula (IIIa ) — the meanings of the symbols applied in the formulae are given in claim 1, furthermore, in general formula (IIa) R2a is either hydrogen or Cι-6 alkyl group and in general formula (IIIab) X is a leaving group, preferably a halogen atom — , b) in case of the synthesis of compounds of general formula (I) wherein R2 stands for a group of general formula (b), a compound of formula (IIbo) is reacted with a compound of general formula (IIIab) — the meanings of the symbols applied in formula (IIIab) are given in claim 1, and X is a leaving group, preferably a halogen atom -, c) in case of the synthesis of compounds of general formula (I) wherein R2 stands for a group of general formula (c), a compound of formula (IIbc) is reacted with a compound of general formula (III-) — the meanings of the symbols applied in formula (III-) are given in claim 1, and X is a leaving group, preferably a halogen — , d) in case of the synthesis of compounds of general formula (I) wherein R2 stands for a group of general formula (d), a compound of formula (IIj) is reacted with a compound of general formula (Hid) — the meanings of the symbols applied in formula (iπd) are given above, and X is a leaving group, preferably a halogen — and in case of the synthesis of compounds containing hydrogen in the position of group Q, the compound of general formula (I) containing oxyl group (0-) in the position of group Q obtained by any of the processes given above is reduced, and, if desired, the compound obtained by any of the processes given above is transformed to a pharmaceutically acceptable salt, or the compound obtained in salt form is transformed into another salt, or the free compound is released from its salt form.
11. A pharmaceutical composition containing one or more compounds of claims 1 to 9 as active ingredients together with pharmaceutically acceptable auxiliary ingredients.
12. Use of compounds of claims 1 to 9 for the preparation a pharmaceutical composition having PARP- inhibiting effect.
13. The use of claim 12, where active ingredient is used for the preparation of a pharmaceutical composition for the treatment or prevention of illnesses selected from the following group: sepsis/septic shock, multiple organ failure syndrome, renal failure, hepatic failure, neurodegenerative and inflammatory diseases, and diseases related to ischemia and reperfusion.
14. The use of claim 13, where active ingredient is used for the preparation of a pharmaceutical composition for the treatment or prevention of illnesses selected from the following group: infarction, diabetes, insulin resistance, metabolic X syndrome, rheumatoid arthritis, and tumour formation in chronic inflammations.
15. Use of compounds of claims 1 to 9 for the treatment of an illness associated with PARP-activity. rw/HU_.004 / 0 0-0 0 4
14
16. The use of claim 15, where the illness is selected from the following group: sepsis/septic shock, multiple organ failure syndrome, renal failure, hepatic failure, neurodegenerative and inflammatory diseases, and diseases related to ischemia and reperfusion.
17. The use of claim 15, where the illness is selected from the following group: infarction, diabetes, insulin resistance, metabolic X syndrome, rheumatoid arthritis, and tumour formation in chronic inflammations.
PCT/HU2004/000044 2003-04-30 2004-04-30 Quinazolinone-derivatives and their use for preparation of pharmaceutical compositions having parp enzyme inhibitory effect Ceased WO2004096779A1 (en)

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Cited By (10)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2011058367A2 (en) 2009-11-13 2011-05-19 Astrazeneca Ab Diagnostic test for predicting responsiveness to treatment with poly(adp-ribose) polymerase (parp) inhibitor
WO2012028578A1 (en) * 2010-09-03 2012-03-08 Bayer Cropscience Ag Substituted fused pyrimidinones and dihydropyrimidinones
WO2013078771A1 (en) 2011-11-30 2013-06-06 成都地奥制药集团有限公司 Poly (adp-ribose) polymerase inhibitor
WO2018162439A1 (en) 2017-03-08 2018-09-13 Onxeo New predictive biomarker for the sensitivity to a treatment of cancer with a dbait molecule
WO2018197461A1 (en) 2017-04-28 2018-11-01 Akribes Biomedical Gmbh A parp inhibitor in combination with a glucocorticoid and/or ascorbic acid and/or a protein growth factor for the treatment of impaired wound healing
WO2019175132A1 (en) 2018-03-13 2019-09-19 Onxeo A dbait molecule against acquired resistance in the treatment of cancer
EP3594343A1 (en) 2015-07-23 2020-01-15 Institut Curie Use of a combination of dbait molecule and parp inhibitors to treat cancer
US10799501B2 (en) 2015-11-05 2020-10-13 King's College Hospital Nhs Foundation Trust Combination of an inhibitor of PARP with an inhibitor of GSK-3 or DOT1L
WO2021148581A1 (en) 2020-01-22 2021-07-29 Onxeo Novel dbait molecule and its use
WO2024261243A1 (en) 2023-06-21 2024-12-26 Hemispherian As Combination comprising a deoxycytidine derivative and a parp inhibitor for use in a method of treating hr proficient cancer

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN106083741A (en) * 2016-02-22 2016-11-09 南京中医药大学 2 sulfur are for quinazolinedione derivatives

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3073826A (en) * 1960-10-20 1963-01-15 Mead Johnson & Co 3-pyrrolidylmethyl-4-quinazolones
WO2002048117A1 (en) 2000-12-11 2002-06-20 Fujisawa Pharmaceutical Co., Ltd. Quinazolinone derivatives
WO2002094790A1 (en) 2001-05-23 2002-11-28 Mitsubishi Pharma Corporation Fused heterocyclic compound and medicinal use thereof

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH07107056B2 (en) * 1987-01-30 1995-11-15 日清製粉株式会社 4 (3H) -quinazolinone derivative, method for producing the same and antiulcer agent containing the same
US20020022636A1 (en) 1997-09-03 2002-02-21 Jia-He Li Oxo-substituted compounds, process of making, and compositions and methods for inhibiting parp activity

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3073826A (en) * 1960-10-20 1963-01-15 Mead Johnson & Co 3-pyrrolidylmethyl-4-quinazolones
WO2002048117A1 (en) 2000-12-11 2002-06-20 Fujisawa Pharmaceutical Co., Ltd. Quinazolinone derivatives
WO2002094790A1 (en) 2001-05-23 2002-11-28 Mitsubishi Pharma Corporation Fused heterocyclic compound and medicinal use thereof
EP1396488A1 (en) * 2001-05-23 2004-03-10 Mitsubishi Pharma Corporation Fused heterocyclic compound and medicinal use thereof

Non-Patent Citations (9)

* Cited by examiner, † Cited by third party
Title
DATABASE CHEMABS [online] CHEMICAL ABSTRACTS SERVICE, COLUMBUS, OHIO, US; MURAV'EVA, K. M. ET AL: "Derivatives of 2-mercapto-4-quinazolone as compounds having potential antitubercular activity", XP002291368, retrieved from STN Database accession no. 1969:96748 *
GRIFFIN R J ET AL: "RESISTANCE MODIFYING AGENTS.5.SYNTHESIS AND BIOLOGICAL PROPERTIES OF QUINAZOLINONE INHIBITORS OF THE DNA REPAIR ENZYME POLY(ADP-RIBOSE) POLYMERASE (PARP)", JOURNAL OF MEDICINAL AND PHARMACEUTICAL CHEMISTRY, AMERICAN CHEMICAL SOCIETY. EASTON, US, vol. 41, 1998, pages 5247 - 5256, XP002208913 *
HANKOVSZKY O H ET AL: "NEW ANTIARRHITHMIC AGENTS. 2,2,5,5-TETRAMETHYL-3-PYRROLINE-3-CARBOXA MIDES AND 2,2,5,5-TETRAMETHYLPYRROLIDINE-3-CARBOXAMIDES", JOURNAL OF MEDICINAL CHEMISTRY, AMERICAN CHEMICAL SOCIETY. WASHINGTON, US, vol. 29, no. 7, 1986, pages 1138 - 1152, XP002068557, ISSN: 0022-2623 *
KHIM. GETEROTSIKL. SOEDIN., SB. 1: AZOTSODERZHASHCHIE GETEROTSIKLY , 411-14. EDITOR(S): HILLERS, S. PUBLISHER: IZD. "ZINATNE", RIGA, USSR. CODEN: 20NNA2, 1967 *
KRISHNA M C ET AL: "STUDIES OF STRUCTURE-ACTIVITY RELATIONSHIP OF NITROXIDE FREE RADICALS AND THEIR PRECURSORS AS MODIFIERS AGAINST OXIDATIVE DAMAGE", JOURNAL OF MEDICINAL CHEMISTRY, AMERICAN CHEMICAL SOCIETY. WASHINGTON, US, vol. 41, no. 8, 1998, pages 3477 - 3492, XP001146049, ISSN: 0022-2623 *
KULCSAR, GYOZO ET AL: "Synthesis and study of new 4-quinazolinone inhibitors of the DNA repair enzyme poly(ADP-ribose) polymerase ( PARP )", ARKIVOC (GAINESVILLE, FL, UNITED STATES) , (5), 121-131 CODEN: AGFUAR URL: HTTP://WWW.ARKAT-USA.ORG/ARK/JOURNAL/2003/BERNATH/GB-733J/GB- 733J.PDF, 11 June 2003 (2003-06-11), XP002291367 *
MANABU HORI ET AL.: "Nootropic agents, 4-Alkoxy-2-(1-piperazinyl)quinazoline derivatives", CHEMICAL AND PHARMACEUTICAL BULLETIN, PHARMACEUTICAL SOCIETY OF JAPAN, vol. 39, no. 2, February 1991 (1991-02-01), pages 367 - 371
MANABU HORI ET AL: "Nootropic agents, 4-Alkoxy-2-(1-piperazinyl)quinazoline Derivatives", CHEMICAL AND PHARMACEUTICAL BULLETIN, PHARMACEUTICAL SOCIETY OF JAPAN. TOKYO, JP, vol. 39, no. 2, February 1991 (1991-02-01), pages 367 - 371, XP002128281, ISSN: 0009-2363 *
MURAEVA ET AL.: "Derivetives of 2-mercapto-4-quinazolone as compounds having potential antitubercular activity", CHEMICAL ABSTRACT SERVICE

Cited By (16)

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Publication number Priority date Publication date Assignee Title
WO2011058367A2 (en) 2009-11-13 2011-05-19 Astrazeneca Ab Diagnostic test for predicting responsiveness to treatment with poly(adp-ribose) polymerase (parp) inhibitor
CN103228141B (en) * 2010-09-03 2016-04-20 拜耳知识产权有限责任公司 Substituted fused pyrimidinones and dihydropyrimidinones
WO2012028578A1 (en) * 2010-09-03 2012-03-08 Bayer Cropscience Ag Substituted fused pyrimidinones and dihydropyrimidinones
CN103228141A (en) * 2010-09-03 2013-07-31 拜耳知识产权有限责任公司 Substituted fused pyrimidinones and dihydropyrimidinones
US9006265B2 (en) 2010-09-03 2015-04-14 Bayer Intellectual Property Gmbh Substituted fused pyrimidinones and dihydropyrimidinones
AU2011298423B2 (en) * 2010-09-03 2015-11-05 Bayer Intellectual Property Gmbh Substituted fused pyrimidinones and dihydropyrimidinones
US9718787B2 (en) 2011-11-30 2017-08-01 Chengdu Di'ao Pharmaceutical Group Co., Ltd. Poly (ADP-ribose) polymerase inhibitor
US9187430B2 (en) 2011-11-30 2015-11-17 Chengdu Di'ao Pharmaceutical Group Co., Ltd. Poly (ADP-ribose) polymerase inhibitor
WO2013078771A1 (en) 2011-11-30 2013-06-06 成都地奥制药集团有限公司 Poly (adp-ribose) polymerase inhibitor
EP3594343A1 (en) 2015-07-23 2020-01-15 Institut Curie Use of a combination of dbait molecule and parp inhibitors to treat cancer
US10799501B2 (en) 2015-11-05 2020-10-13 King's College Hospital Nhs Foundation Trust Combination of an inhibitor of PARP with an inhibitor of GSK-3 or DOT1L
WO2018162439A1 (en) 2017-03-08 2018-09-13 Onxeo New predictive biomarker for the sensitivity to a treatment of cancer with a dbait molecule
WO2018197461A1 (en) 2017-04-28 2018-11-01 Akribes Biomedical Gmbh A parp inhibitor in combination with a glucocorticoid and/or ascorbic acid and/or a protein growth factor for the treatment of impaired wound healing
WO2019175132A1 (en) 2018-03-13 2019-09-19 Onxeo A dbait molecule against acquired resistance in the treatment of cancer
WO2021148581A1 (en) 2020-01-22 2021-07-29 Onxeo Novel dbait molecule and its use
WO2024261243A1 (en) 2023-06-21 2024-12-26 Hemispherian As Combination comprising a deoxycytidine derivative and a parp inhibitor for use in a method of treating hr proficient cancer

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ATE469133T1 (en) 2010-06-15
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HUP0301173A3 (en) 2008-08-28
US7994182B2 (en) 2011-08-09

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