Structure-based design, synthesis, and evaluation of peptide-mimetic SARS 3CL protease inhibitors

The design and evaluation of low molecular weight peptide-based severe acute respiratory syndrome (SARS) chymotrypsin-like protease (3CL) protease inhibitors are described. A substrate-based peptide aldehyde was selected as a starting compound, and optimum side-chain structures were determined, based on a comparison of inhibitory activities with Michael type inhibitors. For the efficient screening of peptide aldehydes containing a specific C-terminal residue, a new approach employing thioacetal to aldehyde conversion mediated by N-bromosuccinimide was devised. Structural optimization was carried out based on X-ray crystallographic analyses of the R188I SARS 3CL protease in a complex with each inhibitor to provide a tetrapeptide aldehyde with an IC(50) value of 98 nM. The resulting compound carried no substrate sequence, except for a P(3) site directed toward the outside of the protease. X-ray crystallography provided insights into the protein-ligand interactions.     

Synthesis, biological activity, and structure-activity relationships for potent cytotoxic rhodium(III) polypyridyl complexes

The complexes mer-[RhCl 3(DMSO-kappa S)(pp)] 1a- 5a may be prepared by reaction of mer,cis-[RhCl 3(DMSO-kappa S) 2(DMSO-kappa O)] with the appropriate polypyridyl ligand (pp = bpy, phen, dpq, dppz, dppn) in CH 3OH/H 2O solution at 75 degrees C. The mer isomers of 1a- 5a are stable in chloroform solution but those of 1a and 2a isomerize rapidly to a mixture of fac and mer isomers in DMSO. The complexes are potent in vitro cytotoxic agents and exhibit IC 50 values that are strongly dependent on the size of the polypyridyl ligand. IC 50 values of, respectively, 4.0 (0.5) and 1.9 (0.5), 0.40 (0.06) and 0.19 (0.05), and 0.079 (0.012) and 0.069 (0.021) microM are observed for 1a- 3a against the human cell lines MCF-7 (breast cancer) and HT-29 (colon cancer). Cellular uptake studies showed a rapid and high accumulation of the polypyridyl compounds. Treatment of HT-29 and MCF-7 cells with 3a leads to significant decreases in cellular oxygen consumption and the rate of extracellular acidification.     

Enzymatic activity characterization of SARS coronavirus 3C-like protease by fluorescence resonance energy transfer technique

AIM: To characterize enzymatic activity of severe acute respiratory syndrome (SARS) coronavirus (CoV) 3C-like protease (3CL(pro)) and its four site-directed mutants. METHODS: Based on the fluorescence resonance energy transfer (FRET) principle using 5-[(2'-aminoethyl)-amino] naphthelenesulfonic acid (EDANS) and 4-[[4-(dimethylamino) phenyl] azo] benzoic acid (Dabcyl) as the energy transfer pair, one fluorogenic substrate was designed for the evaluation of SARS-CoV 3CL(pro) proteolytic activity. RESULTS: The kinetic parameters of the fluorogenic substrate have been determined as Km=404 micromol.L(-1), kcat=1.08 min(-1), and kcat/Km=2.7 mmol(-1).L.min(-1). SARS-CoV 3CL(pro) showed substantial pH and temperature-triggered activity switches, and site-directed mutagenesis analysis of SARS-CoV 3CL(pro) revealed that substitutions of His41, Cys145, and His163 resulted in complete loss of enzymatic activity, while replacement of Met162 with Ala caused strongly increased activity. CONCLUSION: This present work has provided valuable information for understanding the catalytic mechanism of SARS-CoV 3CL(pro). This FRET-based assay might supply an ideal approach for the exploration SARS-CoV 3CL(pro) putative inhibitors.     

Synthesis, antifungal activity, and structure-activity relationships of coruscanone A analogues

Coruscanone A, a plant-derived cyclopentenedione derivative, showed potent in vitro antifungal activity against Candida albicans and Cryptococcus neoformans comparable to amphotericin B and fluconazole. A series of analogues have been synthesized by modification of the cyclopentenedione ring, the enolic methoxy functionality, and the side chain styryl moiety of this natural product lead. A structurally close 1,4-benzoquinone analogue was also prepared. All the compounds were examined for their in vitro activity against major opportunistic fungal pathogens including C. albicans, C. neoformans, and Aspergillus fumigatus and fluconazole-resistant C. albicans strains, with several analogues demonstrating potent antifungal activity. Structure-activity relationship studies indicate that the 2-methoxymethylenecyclopent-4-ene-1,3-dione structural moiety is the pharmacophore responsible for the antifungal activity of this class of compounds while the side chain styryl-like moiety plays an important complementary role, presumably contributing to target binding.     

Synthesis, antibacterial activity and structure-activity relationships of N-substituted 3-methyl-4-diazo-5-pyrazolecarboxamides

A number of N-substituted 3-methyl-4-diazo-5-pyrazolecarboxamides (IVa-p) have been synthesized and tested for their in vitro antibacterial activity. All the compounds have been assayed against several representative Gram-negative and Gram-positive bacteria, as well as against some intestinal bacterial species. Some of the 4-diazopyrazoles IVa-c have shown a quite interesting broad-spectrum activity, while they are ineffective against the "protective" intestinal flora. The structure-activity relationships of the series have been studied quantitatively, via both univariate and multivariate methods: the results are consistent and permit some rationalization of the behaviour of the compounds investigated.     

Synthesis and structure-activity relationships of potent antitumor active quinoline and naphthyridine derivatives

The disease of cancer has been ranked second after cardiovascular diseases and plant-derived molecules have played an important role for the treatment of cancer. Nine cytotoxic plant-derived molecules such as vinblastine, vincristine, navelbine, etoposide, teniposide, taxol, taxotere, topotecan and irinotecan have been approved as anticancer drugs. Recently, epothilones are being emerging as future potential anti-tumor agents. However, targeted cancer therapy has now been rapidly expanding and small organic molecules are being exploited for this purpose. Amongst target specific small organic molecules, quinazoline was found as one of the most successful chemical class in cancer chemotherapy as three drugs namely Gefitinib, Erlotinib and Canertinib belong to this series. Now, quinazoline related chemical classes such as quinolines and naphthyridines are being exploited in cancer chemotherapy and a number of molecules such as compounds EKB-569 (52), HKI-272 (78) and SNS-595 (127a) are in different phases of clinical trials. This review presents the synthesis of quinolines and naphthyridines derivatives, screened for anticancer activity since year 2000. The synthesis of most potent derivatives in each prototype has been delineated. A brief structure activity relationship for each prototype has also been discussed. It has been observed that aniline group at C-4, aminoacrylamide substituents at C-6, cyano group at C-3 and alkoxy groups at C-7 in the quinoline ring play an important role for optimal activity. While aminopyrrolidine functionality at C-7, 2'-thiazolyl at N-1 and carboxy group at C-3 in 1,8-naphthyridine ring are essential for eliciting the cytotoxicity. This review would help the medicinal chemist to design and synthesize molecules for targeted cancer chemotherapy.     

感冒229E病毒3CL蛋白酶抑制剂金丝桃苷衍生物的合成及其构效关系研究

目的寻找作为感冒229E抗原型冠状病毒3CL蛋白酶抑制剂的新化学结构。方法运用分子对接方法在ACD化合物库中发现天然产物金丝桃苷是潜在的新型抑制剂,借助分子模拟的方法进行结构改造,设计并合成了5个金丝桃苷衍生物,采用表面等离子共振(SPR)法测试这些化合物与该蛋白酶的结合能力。结果与结论衍生物Ⅱ、Ⅲ、Ⅳ与蛋白酶的结合能力比原天然产物Ⅰ提高了3倍以上,它们的结合构象也明显不同于Ⅰ与SARS病毒3CL蛋白酶的结合构象。这些结合模式的差异为设计选择性更好的感冒病毒3CL蛋白酶抑制剂提供了有益的参考信息。将计算机辅助药物分子设计、有机合成和生物活性测试有机地结合起来,是发现和设计感冒229E型病毒3CL蛋白酶新型选择性抑制剂的有效途径。     

Acyclic purine phosphonate analogues as antiviral agents. Synthesis and structure-activity relationships

A series of 9-(phosphonoalkyl)purines, which are analogues of 9-[2-(phosphonomethoxy)ethyl]purines (guanine, PMEG, 1; adenine, PMEA, 2), were synthesized. The analogues were tested for activity against herpes simplex virus types 1 and 2 (HSV-1 and HSV-2), human cytomegalovirus (HCMV), Rauscher murine leukemia virus (R-MuLV), and human immunodeficiency virus type 1 (HIV-1). With variations in the length of the alkyl chain, the optimal activity was achieved with two carbons between the purine base and the phosphonomethoxy functionality. Despite the structural similarity and the close pKa2 value of 8 to that of PMEG, no phosphorylation of 8 was observed by the bovine brain guanylate kinase. Since all isosteric analogues of PMEG (7-9) were not inhibitory against HSV-1 and HSV-2, the presence of the 3'-oxygen atom in the PME purines proved critical for anti-HSV activity. Introduction of the 1'-methyl group on the PMEG side chain significantly reduced its anti-HSV activity. Analogue 11, which is a mimic of the phosphate by incorporation of the alpha,alpha-difluoro carbon, was ineffective against HSV-1 and HSV-2. These results suggest that the structural requirements of PME purines for anti-HSV activity appear to be very strict.     

Synthesis and antiviral activity of 5-substituted cytidine analogues: identification of a potent inhibitor of viral RNA-dependent RNA polymerases

As part of our studies of lethal viral mutagens, a series of 5-substituted cytidine analogues were synthesized and evaluated for antiviral activity. Among the compounds examined, 5-nitrocytidine was effective against poliovirus (PV) and coxsackievirus B3 (CVB3) and exhibited greater activity than the clinically employed drug ribavirin. Instead of promoting viral mutagenesis, 5-nitrocytidine triphosphate inhibited PV RNA-dependent RNA polymerase (K(d) = 1.1 +/- 0.1 microM), and this inhibition is sufficient to explain the observed antiviral activity.     

Chemistry and structure activity relationships of bafilomycin A1, a potent and selective inhibitor of the vacuolar H+-ATPase.

Bafilomycin A1, a macrolide antibiotic isolated from the fermentation of Streptomyces spp., is a potent and selective inhibitor of vacuolar-type proton translocating ATP-ases (V-ATPases) and was used to study the physiological role of this class of enzymes. An extensive chemical effort on the unusual structure of this macrolide led to the synthesis of significantly different bafilomycin derivatives. None of the new analogues was more potent than the parent compound but provided a significant amount of information about the structural requirements for the inhibitory activity of bafilomycin A1 in particular on chicken osteoclast (cOc) ATPase. The vinylic methoxy group adjacent to a carbonyl function, the dienic system and the hydroxy group at position 7 were recognized to be essential features for bafilomycin V-ATPase-inhibitory activity. This information was utilized to design simplified novel derivatives as inhibitors of bone resorption.     

Zatosetron, a potent, selective, and long-acting 5HT3 receptor antagonist: synthesis and structure-activity relationships.

Antagonists of 5HT3 receptors are clinically effective in treating nausea and emesis associated with certain oncolytic drugs, including cisplatin. Moreover, these agents may be useful in pharmacological management of several central nervous system disorders, including anxiety, schizophrenia, dementia, and substance abuse. Our studies on aroyltropanamides led to the discovery that dihydrobenzofuranyl esters and amides are potent 5HT3 receptor antagonists. Simple benzoyl derivatives of tropine and 3 alpha-aminotropane possessed weak 5HT3 receptor antagonist activity, as judged by blockade of bradycardia produced by iv injection of serotonin (5HT) to anesthetized rats. Within this series, use of benzofuran-7-carboxamide as the aroyl moiety led to a substantial increase of 5HT3 receptor affinity. The optimal 5HT3 receptor antagonist identified via extensive SAR studies was endo-5-chloro-2,3-dihydro-2,2-dimethyl-N-(8-methyl-8-azabicyclo[3.2.1]oc t- 3-yl)-7-benzofurancarboxamide (Z)-2-butenedioate (zatosetron maleate). The 7-carbamyl regiochemistry, dimethyl substitution, chloro substituent, and endo stereochemistry were all crucial elements of the SAR. Zatosetron maleate was a potent antagonist of 5HT-induced bradycardia in rats (ED50 = 0.86 micrograms/kg i.v.). Low oral doses of zatosetron (30 micrograms/kg) produced long-lasting antagonism of 5HT3 receptors, as evidenced by blockade of 5HT-induced bradycardia for longer than 6 h in rats. Moreover, this compound did not produce hemodynamic effects after i.v. administration to rats, nor did it block carbamylcholine-induced bradycardia in doses that markedly blocked 5HT3 receptors. Thus, zatosetron is a potent, selective, orally effective 5HT3 receptor antagonist with a long duration of action in rats.     

Antitumor agents. 2. Synthesis,structure-activity relationships,and biological evaluation of substituted 5H-pyridophenoxazin-5-ones with potent antiproliferative activity

New antiproliferative compounds, 5H-pyrido[3,2-a]phenoxazin-5-ones (1-10), 5H-benzophenoxazin-5-one (11), 5H-pyrido[2,3-a]phenoxazin-5-one (12), 5H-pyrido[3,4-a]phenoxazin-5-one (13), and 5H-pyrido[4,3-a]phenoxazin-5-one (14), were synthesized and evaluated against representative human neoplastic cell lines. The excellent cytotoxic activity of these polycyclic phenoxazinones, structurally related to the actinomycin chromophore, is discussed in terms of structural changes made to rings A and D (Chart 1). Electron-withdrawing or electron-donating substituents were introduced at different positions of ring A to probe the electronic and positional effects of the substitution. A nitro group in R(2) or in R(1) increases the cytotoxic activity, whereas electron-donating methyl groups in any position lead to 10- to 100-fold decreasing of the activity. The low antiproliferative activity of benzophenoxazinone 11 and pyridophenoxazinones 13 and 14 confirms the crucial role of pyridine nitrogen in the W position of ring D in DNA binding. The unexpected high activity exhibited by 12, which has the nitrogen in the X position, could be ascribed to a different mechanism of action, which needs further investigation.     

Synthesis and structure-activity relationships of 3-aminobenzophenones as antimitotic agents

A new series of 3-aminobenzophenone compounds as potent inhibitors of tubulin polymerization was discovered based on the mimic of the aminocombretastatin molecular skeleton. Lead compounds 5 and 11, with alkoxy groups at the C-4 position of B-ring, were potent cytotoxic agents and inhibitors of tubulin polymerization through the binding to the colchicine-binding site of tubulin. The corresponding antitubulin activities of 5 and 11 were similar to or greater than combretastatin A-4 and AVE-8063. Replacement of the methoxy group with a chloro group in the B ring of aminobenzopheneones (3, 8, and 9) caused drastic decrease in cytotoxic and antitubulin activity except in compounds 4 and 10, which could result from a unique alignment between chloro and amino groups located at the para position to each other. SAR information revealed that introduction of an amino group at the C-3 position in B ring of benzophenones, in addition to a methoxy group at the C-4 position, plays an important role for maximal cytotoxicity.     

Development of broad-spectrum halomethyl ketone inhibitors against coronavirus main protease 3CL(pro)

Coronaviruses comprise a large group of RNA viruses with diverse host specificity. The emergence of highly pathogenic strains like the SARS coronavirus (SARS-CoV), and the discovery of two new coronaviruses, NL-63 and HKU1, corroborates the high rate of mutation and recombination that have enabled them to cross species barriers and infect novel hosts. For that reason, the development of broad-spectrum antivirals that are effective against several members of this family is highly desirable. This goal can be accomplished by designing inhibitors against a target, such as the main protease 3CL(pro) (M(pro)), which is highly conserved among all coronaviruses. Here 3CL(pro) derived from the SARS-CoV was used as the primary target to identify a new class of inhibitors containing a halomethyl ketone warhead. The compounds are highly potent against SARS 3CL(pro) with K(i)'s as low as 300 nM. The crystal structure of the complex of one of the compounds with 3CL(pro) indicates that this inhibitor forms a thioether linkage between the halomethyl carbon of the warhead and the catalytic Cys 145. Furthermore, Structure Activity Relationship (SAR) studies of these compounds have led to the identification of a pharmacophore that accurately defines the essential molecular features required for the high affinity.     

Thenoyltrifluoroacetone, a potent inhibitor of carboxylesterase activity

Thenoyltrifluoroacetone (TTFA), a conventional mitochondrial complex II inhibitor, was found to inhibit purified porcine liver carboxylesterase non-competitively with a K(i) of 0.61x10(-6)M and an IC(50) of 0.54x10(-6)M. Both rat plasma and liver mitochondrial esterases were inhibited in a concentration-dependent fashion. Results indicate that TTFA is a potent inhibitor of carboxylesterase activity, in addition to its ability to inhibit mitochondrial complex II activity. Therefore, caution is warranted in using TTFA as a mitochondrial complex inhibitor in combination with esterase substrates, such as fluorescence probes or vitamin E esters.     

Synthesis of 3-deazaneplanocin A, a powerful inhibitor of S-adenosylhomocysteine hydrolase with potent and selective in vitro and in vivo antiviral activities

The neplanocin A analogue 3-deazaneplanocin A (2b) has been synthesized. A direct SN2 displacement on the cyclopentenyl mesylate 3 by the sodium salt of 6-chloro-3-deazapurine afforded the desired regioisomer 4 as the major product. After deprotection, this material was converted to 3-deazaneplanocin A in two steps. X-ray crystallographic analysis confirmed the assigned structure. Consistent with its potent inhibition of S-adenosylhomocysteine hydrolase, 3-deazaneplanocin A displayed excellent antiviral activity in cell culture against vesicular stomatitis, parainfluenza type 3, yellow fever, and vaccinia viruses. Antiviral activity was also displayed in vivo against vaccinia virus by using a mouse tailpox assay. The significantly lower cytotoxicity of 3-deazaneplanocin A, relative to its parent compound neplanocin A, may be due to its lack of conversion to 5'-triphosphate and S-adenosylmethionine metabolites.     

Discovery of potent anilide inhibitors against the severe acute respiratory syndrome 3CL protease

A diversified library of peptide anilides was prepared, and their inhibition activities against the SARS-CoV 3CL protease were examined by a fluorogenic tetradecapeptide substrate. The most potent inhibitor is an anilide derived from 2-chloro-4-nitroaniline, l-phenylalanine and 4-(dimethylamino)benzoic acid. This anilide is a competitive inhibitor of the SARS-CoV 3CL protease with K(i) = 0.03 muM. The molecular docking experiment indicates that the P1 residue of this anilide inhibitor is distant from the nucleophilic SH of Cys145 in the active site.     

Benarthin: a new inhibitor of pyroglutamyl peptidase. III. Synthesis and structure-activity relationships.

Benarthin, a new inhibitor of pyroglutamyl peptidase (PG-peptidase), has been isolated from the culture filtrate of Streptomyces xanthophaeus MJ244-SF1. The structure of benarthin has been determined to be L-(2,3-dihydroxybenzoyl)arginyl-L-threonine. This structure was confirmed by the total synthesis of benarthin. Moreover, we synthesized benarthin derivatives to obtain information on the relationship between structure and inhibitory activity. The results indicated that the catechol group of benarthin is the essential moiety for the inhibition of PG-peptidase.