Sukesh Kalva

Discovery of potent inhibitor for matrix metalloproteinase-9 by pharmacophore based modeling and dynamics simulation studies

Matrix metalloproteinase-9 (MMP-9) is an attractive target for anticancer therapy. In the present study ligand based pharmacophore modeling was performed to elucidate the structural elements for a diverse class of MMP-9 inhibitors. The pharmacophore model was validated through Güner-Henry (GH) scoring method. The final pharmacophore model consisted of three hydrogen bond acceptors (HBA), and two ring aromatic regions (RA). This model was utilized to screen the natural compound database to seek novel compounds as MMP-9 inhibitors. The identified hits were validated using molecular docking and molecular dynamics simulation studies. Finally, one compound named Hinokiflavone from Juniperus communis had high binding free energy of -26.54kJ/mol compared with the known inhibitors of MMP-9. Cytotoxicity for hinokiflavone was evaluated by MTT assay. Inhibition of MMP-9 in the presence of hinokiflavone was detected by gelatin zymography and gelatinolytic inhibition assay. Results revealed that the natural compounds derived based on the developed pharmacophore model would be useful for further design and development of MMP-9 inhibitors.

Potent inhibitors precise to S1′ loop of MMP-13, a crucial target for osteoarthritis

Matrix metalloproteinase-13 (MMP-13) is the primary MMP involved in cartilage degradation through its particular ability to cleave type-II collagen. This protein is expressed by chondrocytes and synovial cells in human osteoarthritis and rheumatoid arthritis; hence, it is an attractive target for the treatment of arthritic diseases. Currently available inhibitors lack specificity for metalloproteinase because of a common Zn binding site in MMPs; thus, there is a need to identify selective MMP-13 inhibitors for osteoarthritis therapy. Because selectivity is the major concern, both ligand-based and protein-based pharmacophore methodologies were used to identity potent and selective MMP-13 inhibitors. Different hypotheses were validated, and the best hypothesis was used to screen Zinc (natural and chemical) databases to seek novel scaffolds as MMP-13 inhibitors. The identified hits were validated using different strategies, such as Glide Standard precision, extra precision, E-model energies and receiver operating curve (ROC). In addition, potent inhibitors were selected based on two criteria: a similar binding mode as that of the active site PB3 crystal ligand and crucial amino acid interactions that are catalytically important for the function of MMP-13. The candidate potent inhibitors ZINC 02535232, ZINC 08399795, ZINC 12419118 and ZINC 00624580 nearly reproduced the H-bond interactions formed in the crystal structure of 1XUC with reasonable RMSD values exhibiting a novel interaction pattern that was not previously observed in MMP-13 inhibitors. The identified potent hits with diverse chemical scaffolds may be useful in designing new MMP-13 inhibitors.

Synthesis, antimicrobial activity and molecular docking studies of pyrano[2,3-d]pyrimidine formimidate derivatives

Abstract A new series of ten poly-functionalized pyrano[2,3-d]pyrimidine formimidate derivatives were successfully synthesized. All the synthesized compounds were characterized by 1H NMR, 13C NMR, HRMS, and FT-IR spectral analysis. All the synthesized compounds were evaluated for their antimicrobial activity by using the well plate method and MIC by the broth micro dilution method against the strains of bacteria as well as fungus. Four compounds exhibited good to excellent antimicrobial activity. The theoretical binding mode of the target molecules was evaluated by docking studies, which revealed a new molecular scaffold for enhancing the antimicrobial activity of compounds.Graphical AbstractStudies on synthesis, characterization, antimicrobial activity, and molecular docking of novel pyrano[2,3-d]pyrimidine formimidate derivatives (10) are reported. Four of the compounds exhibited excellent activity.

Lead identification and optimization of novel collagenase inhibitors; pharmacophore and structure based studies.

In this study, chemical feature based pharmacophore models of MMP-1, MMP-8 and MMP-13 inhibitors have been developed with the aid of HypoGen module within Catalyst program package. In MMP-1 and MMP-13, all the compounds in the training set mapped HBA and RA, while in MMP-8, the training set mapped HBA and HY. These features revealed responsibility for the high molecular bioactivity, and this is further used as a three dimensional query to screen the knowledge based designed molecules. These pharmacophore models for collagenases picked up some potent and novel inhibitors. Subsequently, docking studies were performed for the potent molecules and novel hits were suggested for further studies based on the docking score and active site interactions in MMP-1, MMP-8 and MMP-13.

Field- and Gaussian-based 3D-QSAR studies on barbiturate analogs as MMP-9 inhibitors

Matrix metalloproteinase-9 (MMP-9) is one of the important enzymes belongs to gelatinase family and involved in multiple cellular processes including proliferation, angiogenesis, and metastasis. Various studies show that N-substituted homopiperazine barbiturates are promising and also selective for the MMP-9 inhibition. In this study, we selected and reported 49 barbiturate derivatives as inhibitors of MMP-9 and performed structure-based 3D-QSAR studies to elucidate the important structural features responsible for binding affinity. Crystal structure of MMP-9 complexed with barbiturate available in PDB was selected and performed quantum-polarized ligand docking for the reported inhibitors. Receptor-based alignment obtained from the docked poses were used for field- and Gaussian-based 3D-QSAR study. The results of both field and Gaussian models gave the good predictive correlation coefficient (test set) q2 of 0. 77 and 0.85, and the conventional correlation coefficient (training set) r2 of 0.845 and 0.928, respectively. The results of 3D-QSAR and docking studies validate each other and provided insight into the structural requirements for MMP-9 inhibition. The outcomes of the contour maps for both steric and Gaussian models results indicate that steric and hydrophobic interactions are major contributing factor for the activity and these findings will be useful to design the new inhibitors against MMP-9.

E-Pharmacophore and Molecular Dynamics Study of Flavonols and Dihydroflavonols as Inhibitors Against DiHydroOrotate DeHydrogenase

DiHydroOrotate DeHydrogenase [huDHODH] is a therapeutic target for Rheumatoid arthritis [RA]. Leflunomide [A771726] is a widely used synthetic inhibitor against huDHODH. We to find more efficient lead like compounds. A four featured E-Pharmacophore A1D4H6R7 was built based on the inhibitor A771726. This pharmacophore was validated by checking its ability to identify known highly active inhibitors of huDHODH and assigning higher fitness scores to them. A reverse validation was also performed where random 4 featured pharmacophores were built and its efficiency in identifying actives was compared with our E-Pharmacophore. Our Epharmacophore was very efficient, since it passed both validations by picking the known active molecules with high fitness scores. This validated E- pharmacophore was searched against the KEGG phytochemicals subset database. This search resulted in 18 molecules which were subjected to docking with huDHODH. The molecules with docking score greater than that of A771726 were selected. The docking results were further validated using MM/GBSA which gave similar ranking with high binding free energy values. The four molecules 6-Methoxytaxifolin, Rhamnetin, Rhamnazin and Pinoquercetin were taken for explicit 3ns simulation and it was observed that all four molecules had acceptable RMSD values and stable interactions. Thus our study, suggests four phytomolecules that might inhibit huDHODH more efficiently than A771726. Interestingly, some of the obtained hits have already been proven in vitro anti-inflammatory activity which confirms that, the developed E-pharmacophore can be used to identify novel small molecules against inflammatory target, huDHODH.

Discovery of Novel 5-lox Inhibitors: Pharmacophore Design, Homology and Docking Studies

Inhibition of leukotriene biosynthesis has been extensively studied as a potential for the development of novel therapies for inflammation, respiratory diseases and, in particular, for asthma. We have designed specific functional inhibitors against 5-Lipoxygenase (5-LOX) using several virtual screening techniques like Homology modeling, MD simulations, Docking and Pharmacophore studies. We have identified 6 analogues of novel 5-LOX inhibitors (IC50 < 19 nM) using pharmacophore models and docking studies and their drug like properties were evaluated. These 6 compounds can be taken for further study including synthesis. 5-LOX (5- Lipoxygenase) was found to be a cytosolic enzyme and contained a non-heme iron atom and calcium. This iron atom is critical to determine the catalytic potential of the enzyme, with oxidation to the ferric (Fe3+) form conferring activity (1). 5-LOX can be activated by an increase in intracellular Ca2+ concentration, diacylglycerols, phosphorylation by MAPKAP (mitogen-activated protein kinase-activated protein kinase) kinase-2 and ERK (Extracellular Receptor Kinase) (2). In the absence of calcium, 5-LOX has minimal catalytic activity. The addition of calcium stimulates both the oxygenation and dehydration reactions of 5-LOX. This enzyme is expressed mainly in leukocytes, in line with the function of leukotrienes as mediators of immune reactions (3). Granulocytes, monocytes/macrophages, mast cells, dendritic cells and B lymphocytes express 5-LOX, whereas platelets, endothelial cells, T cells and erythrocytes do not. 5-lipoxygenase (5- LOX) catalyzes the first two steps in the synthesis of all LTs (Leukotrienes), namely the oxidation of AA to 5- hydroperoxyeicosatetraenoic acid (5HPETE) followed by the dehydration of 5-HPETE to LTA (4). 5-Lipoxygenase has a central role in the biosynthesis of leukotrienes. This enzyme initializes the formation of proinflammatory leukotrienes from arachidonic acid (AA). The potent biological effects of Leukotrienes (LTs) includes leukocyte

Interactome based biomarker discovery for irritable bowel syndrome—A systems biology approach

Irritable bowel syndrome (IBS) is a common functional gastrointestinal (GI) disorder around world with no standard therapy. Till date, the pathophysiology of IBS is not clearly understood due to complexity of the disease. Current study reveals the underlying mechanism of IBS using systems biological approach. The complexity of IBS was explained by constructing protein-protein interaction (PPI) network from the text mined genes/proteins. PPI network displayed 68822 interactions from 3595 proteins. IBS interactome was mapped with colon tissue interactome which resulted in a sub-network containing 153 genes. Further, MCL algorithm was applied to sub-network to identify six major clusters. These cluster genes are involved in several pathways such as MAPK, PI3K/Akt, and NF-kappa B. The obtained clustered genes were prioritized using differentially expressed transcriptome data of 45 IBS and 45 normal volunteers. Among the differentially expressed genes, FUS, UNC5CL and BCLAF1 were found in the clusters, suggesting that the identified clusters could play a potential role in the regulation of IBS. Further pathway analysis of cluster genes revealed their molecular association with IBS. Gene prioritization studies identified top 10 genes that can be used as candidate biomarkers for early diagnosis of IBS. Out of top ten genes, PRPF31 was expressed in all biofluids (serum, saliva and urine).

Whole genome sequencing of Oryza sativa L. cv. Seeragasamba identifies a new fragrance allele in rice

Fragrance of rice is an important trait that confers a large economic benefit to the farmers who cultivate aromatic rice varieties. Several aromatic rice varieties have limited geographic distribution, and are endowed with variety-specific unique fragrances. BADH2 was identified as a fragrance gene in 2005, and it is essential to identify the fragrance alleles from diverse geographical locations and genetic backgrounds. Seeragasamba is a short-grain aromatic rice variety of the indica type, which is cultivated in a limited area in India. Whole genome sequencing of this variety identified a new badh2 allele (badh2-p) with an 8 bp insertion in the promoter region of the BADH2 gene. When the whole genome sequences of 76 aromatic varieties in the 3000 rice genome project were analyzed, the badh2-p allele was present in 13 varieties (approximately 17%) of both indica and japonica types. In addition, the badh2-p allele was present in 17 varieties that already had the loss-of-function allele, badh2-E7. Taken together, the frequency of badh2-p allele (approximately 40%) was found to be greater than that of the badh2-E7 allele (approximately 34%) among the aromatic rice varieties. Therefore, it is suggested to include badh2-p as a predominant allele when screening for fragrance alleles in aromatic rice varieties.

Molecular Docking and Dynamics Simulation Study on the Influence of Zn 2+ on the Binding Modes of Aggrecanase with its Inhibitors

Zinc plays a vital role in structural organization, regulation of function and stabilization of the folded protein, which ultimately activates or inactivates the binding sites of the protein. Its transition makes a major change in the protein and its binding affinity. The ligand binding aggrecanases can be influenced by Zn2+ ions; therefore the study focuses on checking the binding mode in the presence and absence of zinc using Docking and Molecular dynamics simulation. The crystal structure with zinc was considered as wild type (ADAMTS-4-1Zn2+, ADAMTS-5-1Zn2+) and the crystal structure without zinc was considered as the mutant type (ADAMTS-4-0Zn2+, ADAMTS-5-0Zn2+). Mutations were made manually by deleting the zinc atom. ADAMTS-4-1Zn2+ had the best Glide score of -12.66 kcal·mol−1, whereas ADAMTS-4-0Zn2+ had -11.69 kcal·mol−1. ADAMTS-4-1Zn2+ had the best glide energy of -72.29 kcal·mol−1, whereas ADAMTS-4-0Zn2+ had-68.44 kcal·mol−1. ADAMTS-4-1Zn2+ had the best glide e-model of -116.34, whereas ADAMTS-4-0Zn2+ had -104.264. The RMSD value for ADAMTS-4-1Zn2+ and ADAMTS-4-0Zn2+ was 1.9. These results suggested that the absence of zinc decreases the binding affinity of ADAMTS-4 with its inhibitor. ADAMTS-5-1Zn2+ had the best Glide score of -8.32 kcal·mol−1, whereas ADAMTS-5-0Zn2+ had -6.62 kcal·mol−1. ADAMTS-5-1Zn2+ had the best glide energy of -70.28 kcal·mol−1, whereas ADAMTS-5-0Zn2+ had -66.02 kcal·mol−1. ADAMTS-5-1Zn2+ had the best glide e-model of-108.484, whereas ADAMTS-5-0Zn2+ had -93.81. The RMSD value for ADAMTS-5-1Zn2+ and ADAMTS-5-0Zn2+ was 0.48Å. These results confirmed that the absence of zinc decreased the binding affinity of ADAMTS-5 with its inhibitor whereas the presence extended the docking energy range and strengthened the binding affinity. Per-residue interaction study, MM-GBSA and Molecular Dynamics showed that all the four complexes underwent extensive structural changes whereas the complex with zinc was stable throughout the simulation period.