Metabolic profiling of a novel antithrombotic compound, S002-333 and enantiomers: metabolic stability, species comparison and in vitro-in vivo extrapolation.

OBJECTIVE: The aim of this research work was to characterize the metabolism of S002-333, (2-(4'-methoxy-benzenesulfonyl)-2,3,4,9-tetrahydro-1H-pyrido (3,4-b) indole-3-carboxylic acid amide) and its enantiomers, S004-1032 (R-form) and S007-1558 (S-form) in pooled human liver microsomes (PHLM) and pooled liver microsomes (LM) of rat (RLM), rabbit (RABLM), dog (DLM) and monkey (MLM). Another objective of this study was to identify suitable surrogate species to humans for further development of lead candidates. METHOD: In vitro metabolic stability and metabolite identification of S002-333 and enantiomers were carried out in PHLM and LM of various species. The prediction of surrogate species and in vitro in vivo extrapolation were performed based upon the calculated in vitro intrinsic clearance (CLint ). RESULTS/CONCLUSION: The in vitro CLint values for S002-333, S004-1032 and S007-1558 were 0.027 ± 0.005, 0.025 ± 0.004 and 0.036 ± 0.005 ml/min/mg, respectively, in PHLM, indicating that S007-1558 was the most metabolically unstable of the three. The LM of other species showed similar results. A common surrogate species to humans for S002-333 and enantiomers was predicted as rabbit where the extrapolated hepatic clearance (CLH ) did not show a significant difference to the in vivo CLH values. However, none of the species closely mimic humans with respect to the proportion of major metabolites (M-1-M-4) formed in vitro. Likewise, the CLH values were also predicted in humans for S002-333 and enantiomers using various mathematical models. During analysis, there was no chiral inversion evident among the individual isomers throughout in vitro and in vivo experiments. In conclusion, the in vitro results indicate a prominent role of phase I metabolism in the degradation of S002-333 and enantiomers and predict rabbit as an alternative species to conduct further safety and efficacy studies. Copyright © 2016 John Wiley & Sons, Ltd.

Pharmacokinetics, dose proportionality and permeability of S002-333 and its enantiomers, a potent antithrombotic agent, in rabbits.

1. S002-333 [(2-(4'-methoxy-benzenesulfonyl)-2,3,4,9-tetrahydro-1H-pyrido (3,4-b) indole-3-carboxylic acid amide)] is a novel and potent antithrombotic active agent. The present work investigates the pharmacokinetics, bioavailability, dose proportionality and permeability of the racemate, S002-333 in male New Zealand White (NZW) rabbits. 2. Rabbits were administered single intravenous (i.v.) (2 mg/kg) and three oral doses of 10, 20 and 40 mg/kg of S002-333, respectively, at different occasions to evaluate dose proportionality. Serial blood samples were collected and analyzed by a liquid chromatography tandem mass spectrometry (LC-MS/MS) method. Since S002-333 is a racemate consisting of S004-1032 (R) and S007-1558 (S), same samples were analyzed using a chiralcel column so as to evaluate the respective enantiomers. 3. The peak plasma concentration, after oral administration, occurred at ∼10 h post-dose. The clearance (CL) and volume of distribution (Vd) after i.v. dose were found to be 3.05 ± 0.09 l/h/kg and 6.73 ± 1.16 l/kg, respectively. The absolute oral bioavailability of S002-333 was 16.32%, whereas it was 6.62 and 5.90% for R- and S-enantiomers, respectively. The absolute bioavailability of 10, 20 and 40 mg/kg doses were found to be 27.91, 14.39 and 16.91%, respectively. The PAMPA (parallel artificial membrane permeability assay) assay shows that S002-333 has a low-passive permeability at gastric and intestinal environment. 4. In conclusion, S002-333 has low-passive permeability, low CL and large Vd. The R-enantiomer has a "slightly" greater bioavailability than the S-enantiomer.

Trichoderma species mediated differential tolerance against biotic stress of phytopathogens in Cicer arietinum L.

Trichoderma spp. have been reported to aid in imparting biotic as well as abiotic tolerance to plants. However, there are only few reports unfolding the differential ability of separate species of Trichoderma genera generally exploited for their biocontrol potential in this framework. A study was undertaken to evaluate the biocontrol potential of different Trichoderma species namely T. harzianum, T. asperellum, T. koningiopsis, T. longibrachiatum, and T. aureoviride as identified in the group of indigenous isolates from the agricultural soils of Eastern Uttar Pradesh, India. Their biocontrol potential against three major soilborne phytopathogens, i.e., Sclerotium rolfsii, Sclerotinia sclerotiorum, and Colletotrichum capsici was confirmed by dual culture plate technique. Efficient mycoparasitic ability was further assessed in all the isolates in relation to chitinase, ß-1,3 glucanase, pectinase, lipase, amylase, and cellulase production while equally consistent results were obtained for their probable phosphate solubilization and indole acetic acid (IAA) production abilities. The selected isolates were further subjected to test their ability to promote plant growth, to reduce disease incidence and to tolerate biotic stress in terms of lignification pattern against S. rolfsii in chickpea plants. Among the identified Trichoderma species, excellent results were observed for T. harzianum and T. koningiopsis indicating better biocontrol potential of these species in the group and thus exhibiting perspective for their commercial exploitation.

In vitro metabolism of a novel antithrombotic compound, S002-333, and its enantiomers: quantitative cytochrome P450 phenotyping, metabolic profiling and enzyme kinetic studies.

1. S002-333, (2-(4'-methoxy-benzenesulfonyl)-2,3,4,9-tetrahydro-1H-pyrido (3,4-b) indole-3-carboxylic acid amide) is a novel potent antithrombotic molecule currently under development phase. It is the racemic mixture of two enantiomers, namely S004-1032 (R-form) and S007-1558 (S-form). 2. The contribution of five major isoenzymes, namely CYP2B6, 2C9, 2C19, 2D6 and 3A4 was quantified using recombinant P450s in the phase-I metabolism through relative activity factor approach. CYP2C19 was found to be the major contributor for S002-333 and S007-1558, while CYP3A4 showed greater involvement in S004-1032 metabolism. Chemical inhibition and immunoinhibition studies reconfirmed the results in human liver microsomes (HLM). 3. Four major phase-I metabolites of S002-333; M-1 and M-3 (oxidative), M-2 (O-demethylated) and M-4 (dehydrogenated) were characterized in HLM. These metabolites constituted 11.2, 11.3 and 21.5% of the parent in comparison with the net phase-I metabolism of 29.9, 31.4 and 38.3% of S002-333, S004-1032 and S007-1558, respectively. 4. Among CYP2C9, 2C19 and 3A4, the relative contribution of CYP2C9 was found to be maximum during M-1 through M-4 formation. Enzyme kinetic analysis for detected metabolites indicated that M-1 to M-3 followed classical hyperbolic kinetics, whereas M-4 showed evidence of autoactivation. In conclusion, the results suggest prominent role of CYP2C9, 2C19 and 3A4 isoforms for enantioselective disposition of S002-333 in vitro.

Chilli Anthracnose: The Epidemiology and Management.

Indian cuisine is renowned and celebrated throughout the world for its spicy treat to the tongue. The flavor and aroma of the food generated due to the use of spices creates an indelible experience. Among the commonly utilized spices to stimulate the taste buds in Indian food, whole or powdered chilli constitutes an inevitable position. Besides being a vital ingredient of of Indian food, chilli occupy an important position as an economic commodity, a major share in Indian economy. Chilli also has uncountable benefits to human health. Fresh green chilli fruits contain more Vitamin C than found in citrus fruits, while red chilli fruits have more Vitamin A content than as found in carrots. The active component of the spice, Capsaicin possesses the antioxidant, anti-mutagenic, anti-carcinogenic and immunosuppressive activities having ability to inhibit bacterial growth and platelet aggregation. Though introduced by the Portuguese in the Seventeenth century, India has been one of the major producers and exporters of this crop. During 2010-2011, India was the leading exporter and producer of chilli in the world, but recently due to a decline in chilli production, it stands at third position in terms of its production. The decline in chilli production has been attributed to the diseases linked with crop like anthracnose or fruit rot causing the major share of crop loss. The disease causes severe damage to both mature fruits in the field as well as during their storage under favorable conditions, which amplifies the loss in yield and overall production of the crop. This review gives an account of the loss in production and yield procured in chili cultivation due to anthracnose disease in Indian sub-continent, with emphasis given to the sustainable management strategies against the conventionally recommended control for the disease. Also, the review highlights the various pathogenic species of Colletotrichum spp, the causal agent of the disease, associated with the host crop in the country. The information in the review will prove of immense importance for the groups targeting the problem, for giving a collective information on various aspects of the epidemiology and management of the disease.

2,4-Dimethoxyphenylsemicarbazones with anticonvulsant activity against three animal models of seizures: synthesis and pharmacological evaluation.

Various 2,4-dimethoxyphenylsemicarbazones were synthesized starting from 2,4-dimethoxyaniline via a phenylcarbamate intermediate. The structures were confirmed by spectral and elemental analyses. The anticonvulsant activity of the synthesized compounds was established after intraperitoneal administration in three seizure models in mice which include maximal electroshock seizure, subcutaneous pentylenetetrazole, and subcutaneous strychnine-induced seizure screens. Nine compounds exhibited protection in all the three seizure models, and N1-(2,4-dimethoxyphenyl)-N4-(propan-2-one)semicarbazone (17) emerged as the most active compound with no neurotoxicity. These compounds were found to elevate gamma-aminobutyric acid (GABA) levels in the midbrain and medulla oblongata regions equipotent to clobazam.