beta-Lactam derivatives as enzyme inhibitors: 1-peptidyl derivatives of 4-phenylazetidin-2-one as inhibitors of elastase and papain

N-Peptidyl substituted azetidin-2-ones were synthesized and evaluated as inhibitors of the serine protease elastase, and the cysteine protease papain. All compounds were synthesized from 4-phenylazetidin-2-one, either from the racemate or from the pure enantiomers. The (S)-enantiomer was prepared by enantioselective synthesis from (S)-beta-phenyl-beta-alanine, while the (R)-enantiomer was obtained by enzymatic resolution with alpha-chymotrypsin. N-Alkylation with bromoacetates introduced a spacer group which, after hydrolysis to the free acid, was acylated with amino acid esters or di- or tripeptide esters. The enzymatic assays proved some derivatives to be effective inhibitors of PPE and/or papain. N-BOC protected amino acid derivatives without a spacer group inhibited PPE reversibly, while derivatives with spacer group showed either weak or no inhibitory properties. On the other hand, papain was inactivated irreversibly by ethyl (RS)-2-oxo-4-phenylazetidin-1-acetate. The highest inhibitory activity against papain was found for the diastereomers of N-(2-oxo-4-phenylazetidin-1-acetyl)-L-alanyl-L-valine benzyl ester, a compound with a spacer group.     

beta-Lactam derivatives as enzyme inhibitors: N-substituted derivatives of (S)-4-oxoazetidine-2-carboxylate as inhibitors of elastase and papain

N-Alkyl and N-acyl substituted 4-oxoazetidine-2-carboxylates are synthesized and evaluated as inhibitors of the proteases porcine pancreatic elastase (PPE) and papain. The compounds are obtained by alkylation or acylation at the nitrogen of benzyl (S)-4-oxoazetidine-2-carboxylate, which is synthesized by a modified literature procedure. The enzymatic assays prove some derivatives to be effective inhibitors of PPE and/or papain. The N-BOC protected amino acid derivatives 10 and 13 inhibit PPE reversibly with KI-values in the micromolar range. On the other hand, papain is inactivated irreversibly by benzyl (S)-2-(benzyloxycarbonyl)azetidin-1-acetate (6).     

Synthesis and biological evaluation of 1-(3-chloro-2-oxo-4-phenylazetidin-1-yl)-3-(2-oxo-2-(10H-phenothiazin-10-yl)ethyl)urea derivatives

A new series of eleven novel 1-(3-chloro-2-oxo-4-phenylazetidin-1yl)-3-(2-oxo-2-(10H-phenothiazin-10-yl)ethyl)urea derivatives were synthesized by cyclocondensation of various Schiff bases of phenothiazine with chloroacetyl chloride in the presence of triethylamine. Various Schiff bases of phenothiazine were synthesized by condensation of 4-(2-oxo-2-(10H-phenothiazin-10-yl)ethyl semicarbazide with various aryl aldehydes. The synthesized compounds were characterized by IR, MASS and ¹H NMR spectral data and evaluated for in vitro antimicrobial, antitubercular, antioxidant and anticancer activities by disc diffusion method, MIC method, REMA, DPPH, FRAP and MTT assay method, respectively. All synthesized compounds showed moderate-to-significant anti-bacterial and anti-fungal activity and compound 4d, 4g, 4h and 4k showed good antioxidant activity with EC₅₀ value of 55, 57, 56 and 47 μg/ml tested by DPPH method. The compounds 4j at a concentration of 10 μg/ml showed inhibition against the growth of Mycobacterium tuberculosis and 4f showed significant activity against human cervical cancer cell line with IC₅₀ values of 18.26 μM.     

Calcium-independent phosphodiesterase inhibitors as putative antidepressants: [3-(bicycloalkyloxy)-4-methoxyphenyl]-2-imidazolidinones

The synthesis and biological properties of a novel series of selective calcium-independent phosphodiesterase inhibitors are described. These compounds also inhibit the specific binding of [3H]rolipram to rat brain membranes and exhibit efficacy in preclinical models of antidepressant activity in mice, such as reducing immobility in the forced-swim test and reversing reserpine-induced hypothermia. Imidazolidinones 4 and 16 were found to be the most potent compounds studied.     

2-Acetylamino-3-[4-(2-acetylamino-2-carboxyethylsulfanylcarbonylamino)- phenylcarbamoylsulfanyl]propionic acid and its derivatives as a novel class of glutathione reductase inhibitors

Glutathione reductase (GR) catalyzes the reduction of oxidized glutathione to reduced glutathione. The enzyme is an attractive target for the development of antimalarial agents, agents to decrease malarial drug resistance and anticancer agents. In addition, inhibition of the enzyme has been employed as a tool in research for various purposes. In this paper, we present a rational design of 2-acetylamino-3-[4-(2-acetylamino-2-carboxyethylsulfanylcarbonylamino)phenylcarbamoylsulfanyl]propionic acid and its derivatives as irreversible GR inhibitors. The K(i) and k(inact) values of 2-acetylamino-3-[4-(2-acetylamino-2-carboxyethylsulfanylcarbonylamino)phenylcarbamoylsulfanyl]propionic acid, the most potent derivative of the series, are 88 muM and 0.1 min(-1), respectively. Although the K(i) value of the inhibitor is in the micromolar range, it is more potent than N,N-bis(2-chloroethyl)-N-nitrosourea, which is currently the most commonly employed irreversible GR inhibitor with a reported IC(50) value of 646 microM. Additional attractive features of the inhibitor include its ready availability through a one-step synthesis and good solubility in both organic and aqueous solutions.     

Synthesis and anti-HIV activity of 4-[(1,2-dihydro-2-oxo-3H-indol-3-ylidene) amino]-N(4,6-dimethyl-2-pyrimidinyl)-benzene sulfonamide and its derivatives.

4-[(1,2-Dihydro-2-oxo-3H-indol-3-ylidene) amino]-N(4,6-dimethyl-2-pyrimidinyl)-benzene sulphonamide and its derivatives were synthesized by reaction of isatin and its derivatives with sulphadimidine. Their chemical structures have been confirmed by IR, (1)H NMR data and elemental analysis. Investigation of anti-HIV activity of compounds were tested against replication of HIV-1 (IIIB) and HIV-2 (ROD) strains in acutely infected MT-4 cells and the activity compared with standard azidothymidine. Among the compounds tested, 4-[(1,2-dihydro-2 oxo-3H-indol-3-ylidene)amino]-N(4,6-dimethyl-2-pyrimidinyl)-benzene sulphonamide and its N-acetyl derivative were the most active compounds.     

Synthesis and anti-HIV activity of 4-[(1,2-dihydro-2-oxo-3H-indol-3-ylidene) amino]-N(4,6-dimethyl-2-pyrimidinyl)-benzene sulfonamide and its derivatives

4-[(1,2-Dihydro-2-oxo-3H-indol-3-ylidene) amino]-N(4,6-dimethyl-2-pyrimidinyl)-benzene sulphonamide and its derivatives were synthesized by reaction of isatin and its derivatives with sulphadimidine. Their chemical structures have been confirmed by IR, ¹H NMR data and elemental analysis. Investigation of anti-HIV activity of compounds were tested against replication of HIV-1 (IIIB) and HIV-2 (ROD) strains in acutely infected MT-4 cells and the activity compared with standard azidothymidine. Among the compounds tested, 4-[(1,2-dihydro-2 oxo-3H-indol-3-ylidene)amino]-N(4,6-dimethyl-2-pyrimidinyl)-benzene sulphonamide and its N-acetyl derivative were the most active compounds.     

Angiotensin converting enzyme inhibitors as antihypertensive agents: 1-[(2-mercaptocycloalkyl)carbonyl]-L-prolines

The synthesis of 1-[(2-mercaptocyclopentyl)carbonyl]-L-prolines, 1-[(2-mercaptocyclobutyl)carbonyl]-L-prolines and related benzoyl derivatives as pure isomers is described. The abilities of all the compounds to inhibit angiotensin converting enzyme (ACE) in vitro and in vivo and to lower the systolic blood pressure in renal hypertensive dogs were determined. Three of them, namely 1-[[2-(benzoylthio)cyclopentyl]carbonyl]-L-proline (10f(R,S], 1-[(2-mercaptocyclopentyl)carbonyl]-L-proline (10g(R,S], and 1-[[2-(benzoylthio)cyclobutyl]carbonyl]-L-proline (16f(R,S], were found to be as potent as captopril in reducing blood pressure. The influence of chirality and ring size on the ACE inhibition is described.     

Behaviors of crystalline waters of (RS)-7-(2-aminomethylmorpholino)-1- cyclopropyl-6,8-difluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid]

Physicochemical characteristics of crystalline water of (RS)-7-(2-aminomethylmorpholino)-1-cyclopropyl-6,8-difluoro-1,4-di hydro-4-oxo-3-quinoline-carboxylic acid were studied by thermal analyses and powder X-ray diffractometry. The dihydrate of the compound was stable under ambient or humidified conditions. The dihydrate converted to a monohydrate on drying under mild conditions. The mono- and dihydrates transformed into alpha-type anhydrate at 110 degrees C. These three kinds of crystals were convertible each other under appropriate conditions. At 165 degrees C, the hydrates and the alpha-anhydrate converted to beta-type anhydrate. The compressed effects on the crystalline waters were also discussed for the dihydrate by kinetic analyses.     

6-Halogen derivatives of 2-(3-benzoylphenyl)propionic acid

The synthesis of the 6-fluoro (I a), 6-chloro (I b) and 6-bromo (I c) derivatives of the known antiinflammatory agent 2-(3-benzoylphenyl)propionic acid (ketoprofen) is reported. The procedure employed involves the direct phase-transfer methylation of the appropriate arylacetonitriles and the subsequent hydrolysis of the alpha-methyl arylacetonitriles (V) thus obtained. It is noteworthy that (V b) and (V c) were not accompanied by alpha, alpha-dimethylated contaminants, owing to the steric hindrance of the chloro and bromine substituent. The pharmacological evaluation of (I a-c) showed that the presence of the halogen unexpectedly abolished the antiinflammatory and antipyretic activities of the model drug. The 6-ethoxy derivative (I d), isolated as by-product of the synthesis of (I a) was also found inactive. A hypothesis has been formulated on the loss of activity of these compounds.     

Inhibitor of angiotensin I converting enzyme: (4R)-3-[(2S)-3-mercapto-2-methylpropanoyl]-4-thiazolidinecarboxylic acid (YS-980)

Pharmacological studies on a new angiotensin I converting enzyme (ACE) inhibitor (4R)-3-[(2S)-3-mercapto-2-methylpropanoyl]-4-thiazolidinecarboxylic acid (YS-980), were carried out in vitro and in vivo. The activity of ACE prepared from rabbit lung by DEAE-Sepharose chromatography was determined using angiotensin I (Ang I) or hippuryl-l-histidyl-l-leucine (HHL) as a substrate. The concentrations of YS-980 producing 50 per cent inhibition of ACE activity (ID₅₀) when HHL or Ang I was used as a substrate were 9.5 and 13 nM, respectively. The mode of inhibition was competitive, with a K_i of 6 nM. Renin activity was not affected by YS-980. Arterial blood pressure was decreased slightly by intravenous injection of YS-980 (0.1 or 1.0mg/kg) to anesthetized rats with or without pretreatment with pentolinium. Pretreatment with YS-980 suppressed the pressor response to Ang I but not to norepinephrine. Intravenous injection of YS-980 (1mg/kg) caused a 21 per cent increase in the pressor response to angiotensin II. These results indicate that YS-980 is a potent and specific inhibitor of ACE.     

New 4-[(1-benzyl-1H-indol-3-yl)carbonyl]-3-hydroxyfuran-2(5H)-ones, beta-diketo acid analogs as HIV-1 integrase inhibitors

In addition to our recent report on a series of rationally designed benzylindolyldiketo acids acting as potent HIV-1 integrase strand transfer inhibitors, we disclose the results obtained with novel compounds chemically modified on the diketo acid moiety in order to investigate its influence on the biological activity and cytotoxicity. The activity of designed and synthesized 4-[(1-benzyl-1H-indol-3-yl)carbonyl]-3-hydroxyfuran-2(5H)-one derivatives lies in the micromolar range with regard to HIV IN enzymatic activity. The microwave-assisted synthesis was employed in some steps of the chemical procedures.     

Synthesis and Ca2+ antagonistic activity of 2-[2-[(aminoalkyl)oxy]-5-methoxyphenyl]-3,4-dihydro-4-methyl-3-oxo-2H- 1,4-benzothiazines

As an extension of the previous investigation (J. Med. Chem. 1988, 31, 919), we synthesized a series of 2-[2-[(aminoalkyl)oxy]-5-methoxyphenyl]-3,4-dihydro-4-methyl-3-oxo-2H- 1,4-benzothiazines (3) and evaluated their Ca2+ antagonistic activities. Ca2+ antagonistic activity was measured with isolated depolarized guinea pig taenia cecum. On the basis of their potent Ca2+ antagonistic activity, six benzothiazines were selected and further evaluated for their vasocardioselectivity. Among these six compounds, the key compound 15 [3,4-dihydro-2-[5-methoxy-2-[3-[N-methyl-N-[2-[3,4- (methylenedioxy)phenoxy]ethyl]amino]propoxy]phenyl]-4-methyl-3-oxo- 2H-1,4-benzothiazine hydrogen fumarate] was recognized as having the lowest cardioselectivity. Following optical resolution, the absolute configuration of the compound's optically active enantiomer was determined by means of X-ray crystallography of a synthetic precursor (+)-4a. The Ca2+ antagonistic activity of 15 was found to reside primarily in (+)-15 (which was about 7 times more potent than (-)-15). The in vitro study showed that (+)-15 had a low cardioselectivity compared to verapamil and diltiazem. This result suggests that (+)-15 would exhibit less adverse effects due to cardiac inhibition than diltiazem and verapamil in therapeutic use.     

(E)-3-(4-羟基-2-甲氧基苯基)丙烯酸甲酯的合成

4-羟基-2-甲氧基苯甲醛和丙二酸在吡啶和苯胺催化下,经Knoevenagel缩合反应制得(E)-3-(4-羟基-2-甲氧基苯基)丙烯酸,再在1-(3-二甲胺基丙基)-3-乙基碳二亚胺盐酸盐和4-二甲胺基吡啶作用下与甲醇成酯制得(E)-3-(4-羟基-2-甲氧基苯基)丙烯酸甲酯,总收率为58.5％.     

对氯苯氧异丁酸甲氧基苯丙烯酸酯的小鼠急性毒性和大鼠长期毒性

目的评价对氯苯氧异丁酸甲氧基苯丙烯酸酯(AZ)的安全性。方法 小鼠分别ig给予AZ3000～1001mg.kg-1或ip给予AZ1600～983mg.kg-1,观察14d,测定小鼠的半数致死量(LD50)。大鼠ig给予AZ500,291.7和166.7mg.kg-1(为临床拟用剂量的60倍、35倍和20倍),连续13周,停药2周后,观察大鼠一般状况、体质量增长、血液学、血液生化学和病理组织学等指标的变化。结果小鼠ig及ip给AZ的LD50分别为2053和1254mg.kg-1。大鼠给药13周后,AZ500mg.kg-1组血清转氨酶、尿素氮和肝系数升高(P<0.05),AZ291.7mg.kg-1组血清天冬氨酸转氨酶(AST)和肝系数升高(P<0.05),其他各组各指标与溶媒对照组相比均无显著性差异。病理组织学检查显示,AZ500mg.kg-1有1例肝组织炎细胞浸润,其余组大鼠未见与药物毒性相关的明显病变。停药2周后各异常指标均恢复正常。结论 AZ在规定剂量下使用应有较好的安全性。     

Novel, potent aldose reductase inhibitors: 3,4-dihydro-4-oxo-3-[[5-(trifluoromethyl)-2-benzothiazolyl] methyl]-1-phthalazineacetic acid (zopolrestat) and congeners

A new working hypothesis that there is a hitherto unrecognized binding site on the aldose reductase (AR) enzyme with strong affinity for benzothiazoles was pursued for the design of novel, potent aldose reductase inhibitors (ARIs). The first application of this hypothesis led to a novel series of 3,4-dihydro-4-oxo-3-(benzothiazolylmethyl)-1-phthalazineacetic+ + + acids. The parent of this series (207) was a potent inhibitor of AR from human placenta (IC50 = 1.9 x 10(-8) M) and was orally active in preventing sorbitol accumulation in rat sciatic nerve, in an acute test of diabetic complications (ED50 = 18.5 mg/kg). Optimization of this lead through medicinal chemical rationale, including analogy from other drug series, led to more potent congeners of 207 and culminated in the design of 3,4-dihydro-4-oxo-3-[[5-(trifluoromethyl)-2-benzothiazolyl] methyl]-1-phthalazineacetic acid (216, CP-73,850, zopolrestat). Zopolrestat was found to be more potent than 207, both in vitro and in vivo. Its IC50 against AR and ED50 in the acute test were 3.1 x 10(-9)M and 3.6 mg/kg, respectively. Its ED50s in reversing already elevated sorbitol accumulation in rat sciatic nerve, retina, and lens in a chronic test were 1.9, 17.6, and 18.4 mg/kg, respectively. It was well absorbed in diabetic patients, resulting in high blood level, showed a highly favorable plasma half-life (27.5 h), and is undergoing further clinical evaluation. An assortment of synthetic methods used for the construction of benzothiazoles, including an efficient synthesis of zopolrestat, is described. Structure-activity relationships in the new series are discussed.     

beta-lactam derivatives as enzyme inhibitors: carboxy peptidyl derivatives of (S)-4-oxoazetidine-2-carboxylate as peptidomimetics.

4-Oxoazetidine-2-carboxylic acid, protected at the nitrogen by silyl groups, was coupled with amino acid and oligopeptide esters. Desilylation and deprotection of the amino acid residues yielded the free beta-lactam peptides. Structure and properties were elucidated by spectroscopic methods and discussed. Some selected compounds were tested as fibrinogen inhibitors and for thrombocyte aggregation. None of the compounds showed any activity up to a concentration of 10(-5) Mol/l. Some other compounds exhibited a weak inhibitory activity against elastase (PPE).     

Synthesis of hexaprofen [2-(4-cyclohexylphenyl) propionic acid]

A novel preparative method for hexaprofen, which is a potent antiinflammatory agent, is described. Friedel-Crafts reaction of cyclohexylbenzene with ethyl α-chloro-α-(methylthio) acetate1 and α-chloro-α-(methylthio) acetonitrile2 afforded ethyl 2-(methylthio)-2-(4-cyclohexylphenyl) acetate7 and 2-methylthio-2-(4-cyclohexylphenyl) acetonitrile8, respectively. Compounds7 and8 were converted into the corresponding ethyl 2-methylthio-2-(4-cyclohexylphenyl) propionate9 and 2-methylthio-2-(4-cyclohexylphenyl) propionitrile10 by methylation with sodium hydride and methyl iodide. Hexaprofen13 was prepared by hydrolysis of ethyl 2-(4-cyclohexylphenyl) propionate11 and of 2-(4-cyclohexylphenyl) propionitrile12 followed by desulfurization of compounds9 and10.