3,4-disubstituted azetidinones as selective inhibitors of the cysteine protease cathepsin K. Exploring P3 elements for potency and selectivity

The synthesis of a series of highly potent and selective inhibitors of cathepsin K based on the 3,4-disubstituted azetidin-2-one warhead is reported. A high degree of potency and selectivity was achieved by introducing a basic nitrogen into the distal part of the P3 element of the molecule. Data from kinetic and mass spectrometry experiments are consistent with the interpretation that compounds of this series transiently acylate the sulfhydrile of cathepsin K.     

Synthesis and antihypertensive activity of novel 3-benzyl-2-substituted-3H-[1,2,4]triazolo[5,1-b]quinazolin-9-ones

A series of 3-benzyl-2-substituted-3H-[1,2,4]triazolo[5,1-b]quinazolin-9-ones have been synthesized by the cyclocondensation of 3-amino-2-benzylamino-3H-quinazolin-4-one with a variety of one-carbon donors. The starting material 3-amino-2-benzylamino-3H-quinazolin-4-one was synthesized from methyl anthranilate by a novel innovative route. The title compounds were evaluated for their in vivo antihypertensive activity using spontaneously hypertensive rats (SHR). While all the test compounds exhibited significant antihypertensive activity, 3-benzyl-2-methyl-3H-[1,2,4]triazolo[5,1-b] quinazolin-9-one exhibited antihypertensive activity more than the reference standard prazocin.     

Design and structure-activity relationship of heterocyclic analogs of 4-amino-3-benzimidazol-2-ylhydroquinolin-2-ones as inhibitors of receptor tyrosine kinases

Herein are described a series of novel heterocyclic analogs of the 4-amino-3-benzimidazol-2-ylhydroquinolin-2-one scaffold. These compounds are potent inhibitors of receptor tyrosine kinases and exhibit favorable pharmacokinetic profiles. The synthesis and SAR of these compounds are described.     

Microsomal carbonyl reductase responsible for reduction of 4-phenyl-3-buten-2-one in rats

trans-4-Phenyl-3-buten-2-one (PBO), a flavoring additive, was transformed to the carbonyl-reduced product, trans-4-phenyl-3-buten-2-ol (PBOL) by rat liver microsomes, but not by liver cytosol, in the presence of NADH or NADPH. PBOL formed was identified by comparison with an authentic sample. The reductase activity was not inhibited by quercitrin, an inhibitor of cytosolic carbonyl reductase. The carbonyl reduction product of PBO by liver microsomes was identified as the R-enantiomer of PBOL by HPLC analysis. Rat blood also exhibited the carbonyl reductase activity in the presence of NADH or NADPH, but to a lesser extent.     

Solid-phase parallel synthesis and SAR of 4-amidofuran-3-one inhibitors of cathepsin S: Effect of sulfonamides P3 substituents on potency and selectivity

Highly potent and selective 4-amidofuran-3-one inhibitors of cathepsin S are described. The synthesis and structure–activity relationship of a series of inhibitors with a sulfonamide moiety in the P3 position is presented. Several members of the series show sub-nanomolar inhibition of the target enzyme as well as an excellent selectivity profile and good cellular potency. Molecular modeling of the most interesting inhibitors describes interactions in the extended S3 pocket and explains the observed selectivity towards cathepsin K.     

Inhibition of human chymase by 2-amino-3,1-benzoxazin-4-ones

A series of 2-sec.amino-4H-3,1-benzoxazin-4-ones was evaluated as acyl-enzyme inhibitors of human recombinant chymase. The compounds were also assayed for inhibition of human cathepsin G, bovine chymotrypsin, and human leukocyte elastase. Introduction of an aromatic moiety into the 2-substituent resulted in strong inhibition of chymase, cathepsin G, and chymotrypsin. Extension of the N(Me)CH2Ph substituent by one methylene unit was unfavourable to inhibit these proteases. Towards chymase, 2-(N-benzyl-N-methylamino)-4H-3,1-benzoxazin-4-one (32) and 2-(N-benzyl-N-methylamino)-6-methyl-4H-3,1-benzoxazin-4-one (33) were found to exhibit Ki values of 11 and 17 nM, respectively, and form stable acyl-enzymes with half-lives of 53 and 25 min, respectively. Benzoxazinone 33 also inhibited the human chymase-catalyzed formation of angiotensin 11 from angiotensin I.     

Hydroxylated 2-amino-3H-phenoxazin-3-one derivatives as products of 2-hydroxy-1,4-benzoxazin-3-one (HBOA) biotransformation by Chaetosphaeria sp., an endophytic fungus from Aphelandra tetragona

The biotransformation of the phytoanticipin HBOA and its major degradation metabolites 2-hydroxy-N-(2-hydroxyphenyl)acetamide (7) and N-(2-hydroxyphenyl)acetamide (8) by Chaetosphaeria sp., an endophytic fungus isolated from Aphelandra tetragona, was studied. Three new metabolites could be identified as 2-amino-7-hydroxy-3H-phenoxazin-3-one (12), 2-acetylamino-7-hydroxy-3H-phenoxazin-3-one (13) and 7-hydroxy-2-(2-hydroxyacetyl)-amino-3H-phenoxazin-3-one (14). Structure elucidation of 12 and 13 was performed by MS, 1H, 13C NMR and 2D NMR techniques and confirmed by chemical transformation.     

Oxidative condensation of 2-amino-4-methylphenol to dihydrophenoxazinone compound by human hemoglobin.

2-Amino-4-methylphenol was converted to a brownish yellow material by the lysates of human erythrocytes or purified human hemoglobin. The reaction proceeded oxidatively, coupled with the oxidation of hemoglobin. The major component of the brownish yellow material produced by oxidative condensation of 2-amino-4-methylphenol was identified as 3-amino-1,4 alpha-dihydro-4 alpha, 8-dimethyl-2H-phenoxazin-2-one on the basis of its spectral data including NMR spectra, IR spectra, EI mass spectra, and absorption spectra. The changes in 3-amino-1,4 alpha-dihydro-4 alpha,8-dimethyl-2H-phenoxazin-2-one during incubation of purified human hemoglobin and 2-amino-4-methylphenol were analyzed spectrophotometrically and by using HPLC. The reaction mechanism involved may be similar to that of actinomycin synthase, which oxidizes 2-amino-5-methylphenol to the dihydrophenoxazinone.     

N-Arylated pyrrolidin-2-ones and morpholin-3-ones as potassium channel openers

Based on the most stable conformation of ZD6169, a series of N-arylated derivatives of oxazolidindione (2), morpholin-3-one (3-5), piperidin-2-one (6), and pyrrolidin-2-one (7-13) was synthesized and evaluated for potassium channel opening activity. In the in-vitro assays, N-(4-benzoylphenyl)-piperidin-2-on (6) and N-(4-benzoylphenyl)-3,3-dimethyl-pyrrolidin-2-one (9) demonstrated potent and selective relaxant activity at the bladder detrusor muscle [IC50, bladder)=7.4 and 6.7 microM, respectively; IC50 ratio (portal vein/bladder)=41 and 51, respectively].     

3-Acylamino-azetidin-2-one as a novel class of cysteine proteases inhibitors

A new class of inhibitors for cysteine proteases cathepsin B, L, K and S is described. These inhibitors are based on the beta-lactam ring designed to interact with the nucleophilic thiol of the cysteine in the active site of cysteine proteases. Some 3-acylamino-azetidin-2-one derivatives showed very potent inhibition activities for cathepsins L, K and S at the nanomolar or subnanomolar IC(50) values.     

Phenoxazinone synthesis by human hemoglobin.

We found that 2-amino-5-methylphenol was converted to the dihydrophenoxazinone with a reddish brown color by purified human hemoglobin, lysates of human erythrocytes, and human erythrocytes. The reddish brown compound was identified as 2-amino-4,4 alpha-dihydro-4 alpha,7-dimethyl-3H-phenoxazin-3-one by the measurement of NMR spectra, IR spectra, EI mass spectra, and absorption spectra. The changes in this phenoxazinone were studied under various conditions after mixing 2-amino-5-methylphenol with purified oxy- or methemoglobin, or with human erythrocytes. The production of 2-amino-4,4 alpha-dihydro-4 alpha,7-dimethyl-3H-phenoxazine-3-one from 2-amino-5-methylphenol was found to be tightly coupled with the oxidation of ferrous hemoglobin and reduction of ferric hemoglobin under aerobic conditions. By studying the production rates of the dihydrophenoxazinone and the oxido-reduction rates of ferrous and ferric hemoglobins during the reactions of ferrous or ferric hemoglobin with 2-amino-5-methylphenol under aerobic and anaerobic conditions, the reaction mechanism was extensively proposed.     

The nature of the collagenolytic cathepsin of rat liver and its distribution in other rat tissues

1. An enzyme present in rat liver extracts degraded insoluble collagen maximally at pH3.5. Collagenolytic activity was more abundant in kidney, spleen and bone marrow and was also present in decreasing concentrations in ileum, lung, heart, skin and muscle. 2. The crude collagenolytic cathepsin was activated by cysteine and dithiothreitol, but not by 2-mercaptoethanol. Iodoacetamide, p-chloromercuribenzoate and 7-amino-1-chloro-3-l-tosylamidoheptan-2-one hydrochloride inhibited the enzyme. Zn(2+), Fe(3+) and Hg(2+) ions were strongly inhibitory, but Ca(2+), Co(2+), Mg(2+) and Fe(2+) ions had little or no effect. EDTA was an activator of the enzyme. Inhibitors of cathepsin B were found to enhance collagenolysis, but phenylpyruvic acid, a cathepsin D inhibitor, inhibited the enzyme. Di-isopropyl phosphorofluoridate had no effect. 3. Collagenolysis at pH3.5 and 28 degrees C was restricted to cleavage of the telopeptide region in insoluble collagen, and the material that was solubilized consisted mostly of alpha-chains. 4. The collagenolytic cathepsin was separated from cathepsins B2 and D by fractionation on Sephadex G-100 and a partial separation from cathepsin B1 was obtained by chromatography on DEAE-Sephadex. 5. The function of the collagenolytic cathepsin in the catabolism of collagen is discussed in relation to the action of the other lysosomal proteinases and the neutral collagenase.     

Identification of 1,4-Benzoxazin-3-ones in Maize Extracts by Gas-Liquid Chromatography and Mass Spectrometry

Gas-liquid chromatography-mass spectrometry (GLC-MS) has been used for the separation, detection, and identification of 1,4-benzoxazin-3-ones (hydroxamic acids and lactams) and benzoxazolinones found in maize (Zea mays L.) extracts. Compounds of interest were partitioned into ethyl acetate from aqueous maize seedling extracts. For analysis by GLC-MS, trimethylsilyl derivatives were prepared, chromatographed on a column of 3% OV-1, and detected in the mass spectrometer. Mass spectra were obtained for all peaks present in extracts of four maize lines. A data comparison system was developed for relating unidentified spectra to the spectra of the reference compounds. Based on spectral comparisons, three hydroxamic acids (2,4-dihydroxy-2H-1, 4-benzoxazin-3(4H)-one; 2,4-dihydroxy-7-methoxy-2H-1,4-benzoxazin-3(4H)-one; and 2,4-dihydroxy-7,8-dimethoxy-2H-1,4-benzoxazin-3(4H)-one), three lactams (2-hydroxy-2H-1,4-benzoxazin-3(4H)-one; 2,7-dihydroxy-2H-1,4-benzoxazin-3(4H)-one; and 2-hydroxy-7-methoxy-2H-1,4-benzoxazin-3(4H)-one), one benzoxazolinone (6-methoxybenzoxazolinone), and two organic acids (malic and aconitic) were identified in the extracts. In addition, one other hydroxamic acid and one other related compound were tentatively identified based on mass spectral evidence.     

Highly potent and selective alphaVbeta3-receptor antagonists: solid-phase synthesis and SAR of 1-substituted 4-amino-1H-pyrimidin-2-ones

Solid-phase synthesis and SAR of alpha(V)beta(3)-receptor antagonists based on a N(1)-substituted 4-amino-1H-pyrimidin-2-one scaffold are described. The most potent compounds exhibited IC(50) values towards alpha(V)beta(3) in the nano- to subnanomolar range and high selectivity versus related integrins like alpha(IIb)beta(3). For selected examples efficacy in functional cellular assays was demonstrated.     

鲍氏层孔菌子实体的化学成分研究

采用硅胶和Sephadex LH20柱层析方法,从鲍氏层孔菌子实体提取物中分离得到8个化合物。运用NMR和MS等波谱法分析和鉴定为7(8),22(23)-二烯-3-酮-麦角甾烷、4,6,8(14),22(23)-四烯-3-酮-麦角甾烷、麦角甾醇、过氧化麦角甾醇、三十烷酸对羟基苯乙酯、4-(3,4-二羟苯基)-3-丁烯-2-酮、hispolon、hispidin。     

Synthesis and characterization of nickel(II) and copper(II) complexes with tetradentate Schiff base ligands

The 1:1 condensation of 1,2-diaminopropane and 1-phenylbutane-1,3-dione at high dilution gives a mixture of two positional isomers of terdentate mono-condensed Schiff bases 6-amino-3-methyl-1-phenyl-4-aza-2-hepten-1-one (HAMPAH) and 6-amino-3,5-dimethyl-1-phenyl-4-aza-2-hexen-1-one (HADPAH). The mixture of the terdentate ligands has been used for further condensation with pyridine-2-carboxaldehyde or 2-acetylpyridine to obtain the unsymmetrical tetradentate Schiff base ligands. The tetradentate Schiff bases are then allowed to react with the methanol solution of copper(II) and nickel(II) perchlorate separately. The X-ray diffraction confirms the structures of two of the complexes and shows that the condensation site of the diamine with 1-phenylbutane-1,3-dione is the same.     

Optimization of 1,4-diazepan-2-one containing dipeptidyl peptidase IV inhibitors for the treatment of type 2 diabetes

Following the discovery of N-acyl-1,4-diazepan-2-one as a novel pharmacophore for potent and selective DPP-4 inhibitors, optimization of this new lead with different substitution on the seven-membered ring resulted in several highly potent and selective, orally bioavailable, and efficacious DPP-4 inhibitors, such as 3R-methyl-1-cyclopropyl-1,4-diazepan-2-one derivative 9i (DPP-4 IC(50)=8.0 nM) and 3R,6R-dimethyl-1,4-diazepan-2-one derivative 14a (DPP-4 IC(50)=9.7 nM).     

Reaction of serine proteases with substituted 3-alkoxy-4-chloroisocoumarins and 3-alkoxy-7-amino-4-chloroisocoumarins: new reactive mechanism-based inhibitors

The time-dependent inactivation of several serine proteases including human leukocyte elastase, cathepsin G, rat mast cell proteases I and II, and human skin chymase by a number of 3-alkoxy-4-chloroisocoumarins, 3-alkoxy-4-chloro-7-nitroisocoumarins, and 3-alkoxy-7-amino-4-chloroisocoumarins at pH 7.5 and the inactivation of several trypsin-like enzymes including human thrombin and factor XIIa by 7-amino-4-chloro-3-ethoxyisocoumarin and 4-chloro-3-ethoxyisocoumarin are reported. The 3-alkoxy substituent of the isocoumarin is likely interacting with the S1 subsite of the enzyme since the most reactive inhibitor for a particular enzyme had a 3-substituent complementary to the enzyme's primary substrate specificity site (S1). Inactivation of several enzymes including human leukocyte elastase by the 3-alkoxy-7-amino-4-chlorisocoumarins is irreversible, and less than 3% activity is regained upon extensive dialysis of the inactivated enzyme. Addition of hydroxylamine to enzymes inactivated by the 3-alkoxy-7-amino-4-chloroisocoumarins results in a slow (t1/2 greater than 6.7 h) and incomplete (32-57%) regain in enzymatic activity at pH 7.5. Inactivation by the 3-alkoxy-4-chloroisocoumarins and 3-alkoxy-4-chloro-7-nitroisocoumarins on the other hand is transient, and full enzyme activity is regained rapidly either upon standing, after dialysis, or upon the addition of buffered hydroxylamine. The rate of inactivation by the substituted isocoumarins is decreased when substrates or reversible inhibitors are present in the incubation mixture, which indicates active site involvement. The inactivation rates are dependent upon the pH of the reaction mixture, the isocoumarin ring system is opened concurrently with inactivation, and the reaction of 3-alkoxy-7-amino-4-chloroisocoumarins with porcine pancreatic elastase is shown to be stoichiometric. The results are consistent with a scheme where 3-alkoxy-7-amino-4-chloroisocoumarins react with the active site serine of a serine protease to give an acyl enzyme in which a reactive quinone imine methide can be released. Irreversible inactivation could then occur upon alkylation of an active site nucleophile (probably histidine-57) by the acyl quinone imine methide. The finding that hydroxylamine slowly catalyzes partial reactivation indicates that several inactivated enzyme species may exist. The 3-alkoxy-substituted 4-chloroisocoumarins and 4-chloro-7-nitroisocoumarins are simple acylating agents and do not give stable inactivated enzyme structures.(ABSTRACT TRUNCATED AT 400 WORDS)     

Synthesis and biological evaluation of some novel triazol-3-ones as antimicrobial agents

A new series of triazol-3-one derivatives bearing 4-methyl-4H-thieno[3',2': 5,6]thiopyrano[4,3-d][1,3]thiazolyl or 4-(thiophene-3-yl) thiazolyl moiety at 4-position and alkyl substitution at 2-position are synthesized. All the synthesized compounds are characterized by elemental analysis, IR, (1)H NMR, (13)C NMR, and mass spectral data. The newly synthesized compounds are screened for antifungal and antibacterial activities.     

Synthesis of DNA oligodeoxynucleotides containing structurally defined N6-(2-hydroxy-3-buten-1-yl)-adenine adducts of 3,4-epoxy-1-butene

3,4-Epoxy-1-butene (EB) is generated by cytochrome P450-mediated epoxidation of 1,3-butadiene (BD), an important environmental and industrial chemical classified as a probable human carcinogen. The ability of EB to induce point mutations at GC and AT base pairs has been attributed to its reactions with DNA to form covalent nucleobase adducts. Guanine alkylation is preferred at the endocyclic N7 nitrogen, while adenine can be modified at the N1-, N3-, N7-, and the N6 positions. For each of these sites, a pair of regioisomeric 2-hydroxy-3-buten-1-yl and 1-hydroxy-3-buten-2-yl adducts is produced as a result of epoxide ring opening at the terminal C-4 or the internal C-3 carbon position of EB, respectively. The N6-EB-adenine adducts are of particular interest because of their stability in DNA, potentially leading to their accumulation in vivo. In the present work, synthetic DNA oligomers containing structurally defined N6-(2-hydroxy-3-buten-1-yl)-dA (N6-HB-dA) adducts were prepared for the first time by a postoligomerization approach that involved coupling 6-chloropurine-containing DNA with synthetic 1-amino-3-buten-2-ol. N6-HB-dA-containing DNA oligomers were isolated by reversed phase HPLC, and the presence of N6-HB-dA in their structure was confirmed by molecular weight determination from HPLC-ESI- -MS of the intact strands and by HPLC-ESI+-MS/MS and MS/MS/MS analyses of the enzymatic digests using synthetic N6-HB-dA as an authentic standard. N6-HB-dA-containing oligomers generated in this study will be used for structural and biological studies.