Synthesis and biological evaluation of xanthine derivatives with phenacyl group as tryptophan hydroxylase 1 (TPH1) inhibitors for obesity and fatty liver disease.

Tryptophan hydroxylase 1 (TPH1) has emerged as a target for the treatment of metabolic diseases including obesity and fatty liver disease. A series of xanthine derivatives were synthesized and evaluated for their TPH1 inhibition. Among the synthesized compounds, compound 40 showed good in vitro activity and liver microsomal stability. Docking studies revealed that compound 40 showed better binding to TPH1 via key intermolecular interactions involving the xanthine scaffold, imidazo-thiazolyl ring, and hydroxyl-containing phenacyl moiety. In addition, compound 40 effectively suppressed the adipocyte differentiation of 3 T3-L1 cells.

Discovery of Novel Coumarin Derivatives as Potential Dual Inhibitors against α-Glucosidase and α-Amylase for the Management of Post-Prandial Hyperglycemia via Molecular Modelling Approaches.

Coumarin derivatives are proven for their therapeutic uses in several human diseases and disorders such as inflammation, neurodegenerative disorders, cancer, fertility, and microbial infections. Coumarin derivatives and coumarin-based scaffolds gained renewed attention for treating diabetes mellitus. The current decade witnessed the inhibiting potential of coumarin derivatives and coumarin-based scaffolds against α-glucosidase and α-amylase for the management of postprandial hyperglycemia. Hyperglycemia is a condition where an excessive amount of glucose circulates in the bloodstream. It occurs when the body lacks enough insulin or is unable to correctly utilize it. With open-source and free in silico tools, we have investigated novel 80 coumarin derivatives for their inhibitory potential against α-glucosidase and α-amylase and identified a coumarin derivative, CD-59, as a potential dual inhibitor. The ligand-based 3D pharmacophore detection and search is utilized to discover diverse coumarin-like compounds and new chemical scaffolds for the dual inhibition of α-glucosidase and α-amylase. In this regard, four novel coumarin-like compounds from the ZINC database have been discovered as the potential dual inhibitors of α-glucosidase and α-amylase (ZINC02789441 and ZINC40949448 with scaffold thiophenyl chromene carboxamide, ZINC13496808 with triazino indol thio phenylacetamide, and ZINC09781623 with chromenyl thiazole). To summarize, we propose that a coumarin derivative, CD-59, and ZINC02789441 from the ZINC database will serve as potential lead molecules with dual inhibition activity against α-glucosidase and α-amylase, thereby discovering new drugs for the effective management of postprandial hyperglycemia. From the reported scaffold, the synthesis of several novel compounds can also be performed, which can be used for drug discovery.

Identification of New Non-BBB Permeable Tryptophan Hydroxylase Inhibitors for Treating Obesity and Fatty Liver Disease.

Serotonin (5-hydroxytryptophan) is a hormone that regulates emotions in the central nervous system. However, serotonin in the peripheral system is associated with obesity and fatty liver disease. Because serotonin cannot cross the blood-brain barrier (BBB), we focused on identifying new tryptophan hydroxylase type I (TPH1) inhibitors that act only in peripheral tissues for treating obesity and fatty liver disease without affecting the central nervous system. Structural optimization inspired by para-chlorophenylalanine (pCPA) resulted in the identification of a series of oxyphenylalanine and heterocyclic phenylalanine derivatives as TPH1 inhibitors. Among these compounds, compound 18i with an IC50 value of 37 nM was the most active in vitro. Additionally, compound 18i showed good liver microsomal stability and did not significantly inhibit CYP and Herg. Furthermore, this TPH1 inhibitor was able to actively interact with the peripheral system without penetrating the BBB. Compound 18i and its prodrug reduced body weight gain in mammals and decreased in vivo fat accumulation.

Effects of molecular shape on alcohol aggregation and water hydrogen bond network behavior in butanol isomer solutions.

Despite butanol isomers such as n-butanol, sec-butanol, isobutanol and tert-butanol having the same chemical formula, their liquid-liquid phase diagrams are distinct. That is, tert-butanol is miscible in water at all concentrations, while the other three butanol isomers are partially miscible under ambient conditions. The molecular shape of tert-butanol is close to globular and differs from the other three butanol molecules with a relatively long carbon chain. By performing molecular dynamics simulations and graph theoretical analysis of the four water-butanol isomer mixtures at varying concentrations, we show how distinct butanol aggregates are formed which depend upon the molecular shape and affect the water H-bond network structure and phase diagram in the binary liquid. The three butanol isomers of n-butanol, sec-butanol and isobutanol at concentrated solutions form chain-like alcohol aggregates, but tert-butanol forms small aggregates due to the distinct packing behavior caused by its globular molecular shape. By employing the graph theoretical analysis such as the degree distribution and the eigenvalue spectrum from the adjacency matrix in the graphical representation of the alcohol H-bond network, we show that the tert-butanol aggregates have a different morphological structure from that of the other three butanol isomers in aqueous solution. The graph theoretically distinct butanol aggregates are categorized into two groups, water-compatible and water-incompatible, depending upon the interaction between the alcohol and water molecules. Based upon our observations, we propose that the water-incompatible networks of n-butanol, sec-butanol and isobutanol aggregates do not change the water structure significantly, forming two separate liquid phases that are alcohol-rich and water-rich. However, the water-compatible network of tert-butanol aggregates has a considerable interaction with the water molecules and causes significant disruption of the water H-bond network, forming a homogeneous solution. Understanding the alcohol aggregation behavior and water structure in butanol-water mixtures provides a critical clue in appreciating fundamental issues such as miscibility and phase separation in aqueous solution systems.

Peripheral Selective Oxadiazolylphenyl Alanine Derivatives as Tryptophan Hydroxylase 1 Inhibitors for Obesity and Fatty Liver Disease.

Tryptophan hydroxylase 1 (TPH1) has been recently suggested as a promising therapeutic target for treating obesity and fatty liver disease. A new series of 1,2,4-oxadiazolylphenyl alanine derivatives were identified as TPH1 inhibitors. Among them, compound 23a was the most active in vitro, with an IC50 (half-maximal inhibitory concentration) value of 42 nM, showed good liver microsomal stability, and showed no significant inhibition of CYP and hERG. Compound 23a inhibited TPH1 in the peripheral tissue with limited BBB penetration. In high-fat diet-fed mice, 23a reduced body weight gain, body fat, and hepatic lipid accumulation. Also, 23a improved glucose intolerance and energy expenditure. Taken together, compound 23a shows promise as a therapeutic agent for the treatment of obesity and fatty liver diseases.

Understanding alcohol aggregates and the water hydrogen bond network towards miscibility in alcohol solutions: graph theoretical analysis.

Under ambient conditions, methanol and ethanol are miscible in water at all concentrations, while n-butanol is partially miscible. This is the first study to quantitatively examine the miscibility of butanol and compare with miscible alcohols by employing molecular dynamics simulations and graph theoretical analysis of three water-alcohol mixtures at various concentrations. We show how distinct alcohol aggregates are formed, thereby affecting the water structure, which established the relationship between the morphological structure of the aggregates and the miscibility of the alcohol in aqueous solution. The aggregates of methanol and ethanol in highly concentrated solutions form an extended H-bond network that intertwines well with the H-bond network of water. n-Butanol tends to self-associate and form large aggregates, while such aggregates are segregated from water. Graph theoretical analysis revealed that the alcohol aggregates of methanol and ethanol solutions have a morphological structure different from that of n-butanol, although there is no significant difference in morphology between the three pure alcohols. These two distinct alcohol aggregates are classified as water-compatible and water-incompatible depending upon their interaction with the water H-bond network, and their effect on the water structure was investigated. Our study reveals that the water-compatible network of alcohol aggregates in methanol and ethanol solutions disrupts the water H-bond networks, while the water-incompatible network of n-butanol aggregates does not considerably alter the water structure, which is consistent with the experimental results. Furthermore, we propose that miscible alcohols form water-compatible networks in binary aqueous systems while partially miscible alcohols form water-incompatible networks. The bifurcating hypothesis on the alcohol aggregation behavior in liquid water is of critical use to understand the fundamental issues such as solubility and phase separation in solution systems.

Development of Human Serum Albumin Selective Fluorescent Probe Using Thieno[3,2-b]pyridine-5(4H)-one Fluorophore Derivatives.

The level of human serum albumin (HSA) in biological fluids is a key health indicator and its quantitative determination has great clinical importance. In this study, we developed a selective and sensitive fluorescent HSA probe by fluorescence-based high-throughput screening of a set of fluorescent thieno[3,2-b]pyridine-5(4H)-one derivatives against major plasma proteins: HSA, bovine serum albumin (BSA), globulin, fibrinogen, and transferrin. The fluorophore chosen finally (4) showed noticeable fluorescence enhancement in the presence of HSA (160-fold increase), and it exhibited rapid response, high sensitivity (detection limit 8 nM), and the ability to clearly distinguish HSA from BSA in pH 9 buffer condition. Moreover, the probe could be applicable to detect trace amounts of HSA in an artificial urine sample; further, it might be applied to the determination of the HSA concentration in complex biological samples for pre-clinical diagnosis.

Natural compounds as potential Hsp90 inhibitors for breast cancer-Pharmacophore guided molecular modelling studies.

Breast cancer is one of the major impediments affecting women globally. The ATP-dependant heat shock protein 90 (Hsp90) forms the central component of molecular chaperone machinery that predominantly governs the folding of newly synthesized peptides and their conformational maturation. It regulates the stability and function of numerous client proteins that are frequently upregulated and/or mutated in cancer cells, therefore, making Hsp90 inhibition a promising therapeutic strategy for the development of new efficacious drugs to treat breast cancer. In the present in silico investigation, a structure-based pharmacophore model was generated with hydrogen bond donor, hydrogen bond acceptor and hydrophobic features complementary to crucial residues Ala55, Lys58, Asp93, Ile96, Met98 and Thr184 directed at inhibiting the ATP-binding activity of Hsp90. Subsequently, the phytochemical dataset of 3210 natural compounds was screened to retrieve the prospective inhibitors after rigorous validation of the model pharmacophore. The retrieved 135 phytocompounds were further filtered by drug-likeness parameters including Lipinski's rule of five and ADMET properties, then investigated via molecular docking-based scoring. Molecular interactions were assessed using Genetic Optimisation for Ligand Docking program for 95 drug-like natural compounds against Hsp90 along with two clinical drugs as reference compounds - Geldanamycin and Radicicol. Docking studies revealed three phytochemicals are better than the investigated clinical drugs. The reference and hit compounds with dock scores of 48.27 (Geldanamycin), 40.90 (Radicicol), 73.04 (Hit1), 72.92 (Hit2) and 68.12 (Hit3) were further validated for their binding stability through molecular dynamics simulations. We propose that the non-macrocyclic scaffolds of three identified phytochemicals might aid in the development of novel therapeutic candidates against Hsp90-driven cancers.

Computational Simulations Identify Pyrrolidine-2,3-Dione Derivatives as Novel Inhibitors of Cdk5/p25 Complex to Attenuate Alzheimer's Pathology.

: Mechanistically, neurotoxic insults provoke Ca2+-mediated calpain activation, which cleaves the cytoplasmic region of membrane-embedded p35 and produces its truncated form p25. Upon physical interaction, cyclin-dependent kinase 5 (Cdk5) and p25 forms hyperactivated Cdk5/p25 complex and causes severe neuropathological aberrations including hyperphosphorylated tau-mediated neurofibrillary tangles formation, Alzheimer's symptoms, and neuronal death. Therefore, the inhibition of Cdk5/p25 complex may relieve p-tau-mediated Alzheimer's pathology. Herein, computational simulations have identified pyrrolidine-2,3-dione derivatives as novel inhibitors of Cdk5/p25 complex. A ligand-based pharmacophore was designed and employed as 3D query to retrieve drug-like molecules from chemical databases. By molecular docking, drug-like molecules obtaining dock score > 67.67 (Goldcore of the reference compound) were identified. Molecular dynamics simulation and binding free energy calculation retrieved four pyrrolidine-2,3-dione derivatives as novel candidate inhibitors of Cdk5/p25. The root means square deviation of Cdk5/p25 in complex with candidate inhibitors obtained an average value of ~2.15 Å during the 30 ns simulation period. Molecular interactions analysis suggested that each inhibitor occupied the ATP-binding site of Cdk5/p25 and formed stable interactions. Finally, the binding free energy estimation suggested that each inhibitor had lowest binding energy than the reference compound (-113.10 kJ/mol) to recapitulate their strong binding with Cdk5/p25. Overall, these inhibitors could mitigate tau-mediated Alzheimer's phenotype.

Discovery of Small Molecules that Target Vascular Endothelial Growth Factor Receptor-2 Signalling Pathway Employing Molecular Modelling Studies.

Angiogenesis is defined as the formation of new blood vessels and is a key phenomenon manifested in a host of cancers during which tyrosine kinases play a crucial role. Vascular endothelial growth factor receptor-2 (VEGFR-2) is pivotal in cancer angiogenesis, which warrants the urgency of discovering new anti-angiogenic inhibitors that target the signalling pathways. To obtain this objective, a structure-based pharmacophore model was built from the drug target VEGFR-2 (PDB code: 4AG8), complexed with axitinib and was subsequently validated and employed as a 3D query to retrieve the candidate compounds with the key inhibitory features. The model was escalated to molecular docking studies resulting in seven candidate compounds. The molecular docking studies revealed that the seven compounds displayed a higher dock score than the reference-cocrystallised compound. The GROningen MAchine for Chemical Simulations (GROMACS) package guided molecular dynamics (MD) results determined their binding mode and affirmed stable root mean square deviation. Furthermore, these compounds have preserved their key interactions with the residues Glu885, Glu917, Cys919 and Asp1046. The obtained findings deem that the seven compounds could act as novel anti-angiogenic inhibitors and may further assist as the prototype in designing and developing new inhibitors.

Investigation of novel chemical scaffolds targeting prolyl oligopeptidase for neurological therapeutics.

Prolyl oligopeptidase (POP) is a potential therapeutic target for treatment of several neurological disorders and α-synucleinopathies including Parkinson's disease. Most of the known POP inhibitors failed in the clinical trials due to poor pharmacokinetic properties and blood-brain impermeability. Therefore, a training set of 30 structurally diverse compounds with a wide range of inhibitory activity against POP was used to generate a quantitative pharmacophore model, Hypo 3, to identify potential POP inhibitors with desirable drug-like properties. Validations through test set, cost analysis, and Fisher's randomization methods proved that Hypo 3 accurately predicted the known inhibitors among inactive compounds. Hypo 3 was employed as 3D query for virtual screening on an in-house drug-like chemical database containing compounds with good brain permeability and ADMET parameters. Database screening with Hypo 3 resulted in 99 compounds that were narrowed down to 21 compounds through molecular docking. Among them, five compounds were identified in our earlier studies, while two compounds showed in vitro POP inhibition. The current study proposed new 16 virtually screened compounds as potential inhibitors against POP that possess Gold docking score in the range of 64.61-75.74 and Chemscore of -32.25 to -38.35. Furthermore, the top scoring four hit compounds were subjected to molecular dynamics simulations to reveal their appropriate binding modes and assessing binding free energies. The hit compounds interacted with POP effectively via hydrogen bonds with important active site residues along with hydrophobic interactions. Moreover, the hit compounds had key inter-molecular interactions and better binding free energies as compared to the reference inhibitor. A potential new hydrogen bond interaction was discovered between Hit 2 with the Arg252 residue of POP. To conclude, we propose four hit compounds with new structural scaffolds against POP for the lead development of POP-based therapeutics for neurological disorders.

Exploring the Therapeutic Ability of Fenugreek against Type 2 Diabetes and Breast Cancer Employing Molecular Docking and Molecular Dynamics Simulations.

Fenugreek (Trigonella foenum-graecum) is used as a spice throughout the world. It is known for its medicinal properties such as antidiabetic, anticarcinogenic, and immunological activities. The present study shows the properties and the nutritional quality of fenugreek seed extract and focuses on screening of active compounds in drug designing for type 2 diabetes and breast cancer. Quantitative analysis was used to calculate the percentages of protein, carbohydrates moisture, fatty acid, galactomannan, oil, and amino acid. Phytochemical analysis revealed the presence of flavonoids, terpenoids, phenols, proteins, saponins, and tannins in fenugreek seed extracts. Molecular docking and molecular dynamics simulation-based computational drug discovery methods were employed to address the role of fenugreek seed constituents against type 2 diabetes and breast cancer. The computational results reveal that the compound galactomannan can be ascribed as potential drug candidate against breast cancer and type 2 diabetes rendered by higher molecular dock scores, stable molecular dynamics (MD) simulations results, and lower binding energy calculations.

Targeting natural compounds against HER2 kinase domain as potential anticancer drugs applying pharmacophore based molecular modelling approaches.

Human epidermal growth factor receptors are implicated in several types of cancers characterized by aberrant signal transduction. This family comprises of EGFR (ErbB1), HER2 (ErbB2, HER2/neu), HER3 (ErbB3), and HER4 (ErbB4). Amongst them, HER2 is associated with breast cancer and is one of the most valuable targets in addressing the breast cancer incidences. For the current investigation, we have performed 3D-QSAR based pharmacophore search for the identification of potential inhibitors against the kinase domain of HER2 protein. Correspondingly, a pharmacophore model, Hypo1, with four features was generated and was validated employing Fischer's randomization, test set method and the decoy test method. The validated pharmacophore was allowed to screen the colossal natural compounds database (UNPD). Subsequently, the identified 33 compounds were docked into the proteins active site along with the reference after subjecting them to ADMET and Lipinski's Rule of Five (RoF) employing the CDOCKER implemented on the Discovery Studio. The compounds that have displayed higher dock scores than the reference compound were scrutinized for interactions with the key residues and were escalated to MD simulations. Additionally, molecular dynamics simulations performed by GROMACS have rendered stable root mean square deviation values, radius of gyration and potential energy values. Eventually, based upon the molecular dock score, interactions between the ligands and the active site residues and the stable MD results, the number of Hits was culled to two identifying Hit1 and Hit2 has potential leads against HER2 breast cancers.

Discovery of novel anti-leishmanial agents targeting LdLip3 lipase.

Leishmaniasis is a neglected tropical disease, caused by several species of Leishmania. Being an opportunistic lipid-scavenging pathogen, Leishmania relies extensively on lipid metabolism especially for host-pathogen interaction, utilizing host lipids for energy and virulence. The rational approach is to target lipid metabolism of the pathogen focusing lipid-catabolizing lipases. The LdLip3 lipase is considered as drug target as it is constitutively expressed in both promastigote and amastigote forms. Since the LdLip3 structure is not known, we modeled its three-dimensional structure to implement structure-based drug discovery approach. Similarity-based virtual screening was carried out to identify potential inhibitors utilizing NCI diversity set on ZINC database including natural products. Implementing computational and experimental approaches, four anti-leishmanial agents were discovered. The screened molecules ZINC01821375, ZINC04008765, ZINC06117316 and ZINC12653571 had anti-leishmanial activity with IC50 (% viable promastigotes vs. concentration) of 5.2±1.8µM, 13.1±2.6µM, 9.4±2.6µM and 17.3±3.1µM, respectively. The molecules showed negligible toxicity toward mouse macrophages. Based on the contact footprinting analysis, new molecules were designed with better predicted free energy of binding than discovered anti-leishmanial agents. Further validation for the therapeutic utility of discovered molecules can be carried out by the research community to combat leishmaniasis.

Unraveling the rationale behind organic solvent stability of lipases.

Organic solvent-stable lipases have pronounced impact on industrial economy as they are involved in synthesis by esterification, interesterification, and transesterification. However, very few of such natural lipases have been isolated till date. A study of the recent past provided few pillars to rely on for this work. The three-dimensional structure, inclusive of the surface and active site, of 29 organic solvent-stable lipases was analyzed by subfamily classification and protein solvent molecular docking based on fast Fourier transform correlation approach. The observations revealed that organic solvent stability of lipases is their intrinsic property and unique with respect to each lipase. In this paper, factors like surface distribution of charged, hydrophobic, and neutral residues, interaction of solvents with catalytically immutable residues, and residues interacting with essential water molecules required for lipase activity, synergistically and by mutualism contribute to render a stable lipase organic solvent. The propensity of surface charge in relation to stability in organic solvents by establishing repulsive forces to exclude solvent molecules from interacting with the surface and prohibiting the same from gaining entry to the protein core, thus stabilizing the active conformation, is a new finding. It was also interesting to note that lipases having equivalent surface-exposed positive and negative residues were stable in a wide range of organic solvents, irrespective of their LogP values.

In silico characterization of thermostable lipases.

Thermostable lipases are of high priority for industrial applications as they are endowed with the capability of carrying out diversified reactions at elevated temperatures. Extremophiles are their potential source. Sequence and structure annotation of thermostable lipases can elucidate evolution of lipases from their mesophilic counterparts with enhanced thermostability hence better industrial potential. Sequence analysis highlighted the conserved residues in bacterial and fungal thermostable lipases. Higher frequency of AXXXA motif and poly Ala residues in lid domain of thermostable Bacillus lipases were distinguishing characteristics. Comparison of amino acid composition among thermostable and mesostable lipases brought into light the role of neutral, charged and aromatic amino acid residues in enhancement of thermostability. Structural annotation of thermostable lipases with that of mesostable lipases revealed some striking features which are increment of gamma turns in thermostable lipases; being first time reported in our paper, longer beta strands, lesser beta-branched residues in helices, increase in charged-neutral hydrogen bonding pair, hydrophobic-hydrophobic contact and differences in the N-cap and C-cap residues of the α helices. Conclusively, it can be stated that subtle changes in the arrangement of amino acid residues in the tertiary structure of lipases contributes to enhanced thermostability.

Deciphering role of amino acids for the stability of Staphylococcus aureus lipase (SAL3).

Lipase from Staphylococcus aureus (SAL3) has been gaining attention because of its potential application in detergent industries against its thermostability and pH stability. It has been also used in synthesis of starch oleate recently. Protein engineering of SAL3 will give insight in development of better lipase for industrial applications. The present work aims at bringing out the role of amino acid residues in the stability of SAL3 using prosa2003 (Protein Structure Analysis) program. 127 amino acid residues were identified as structurally/strategically important for stability of the protein among the modeled 386 residues and further categorized into four classes. In the first class, 47 positions are predicted at which substitution of any one of these 47 positions by any other amino acids will lead to destabilization of the protein. The second class comprises of 33 positions which can be mutated by one corresponding amino acid without affecting the stability of the protein. The third class comprises of 21 positions which can be mutated by two corresponding amino acids without affecting the stability of the protein. The fourth class comprises of 26 positions which can be substituted by three corresponding amino acids without affecting the stability of the protein. Further five residues Glu9, Glu136, Phe286, Lys305 and Leu358 are identified in which substitution by any amino acid does not affect protein stability. Hence mutation can be carried out at these positions in the direction of increasing protein stability. In conclusion, this study prefigures well as a prototype for mutation and protein engineering studies of SAL3.

In silico characterization of thermoactive, alkaline and detergent-stable lipase from a Staphylococcus aureus strain.

Bacterial true lipases having thermo and alkaline stability are highly attractive for their industrial production of pharmaceuticals, agrochemicals, cosmetics, and flavour. Staphylococcus aureus lipase (SAL3) remains active at temperatures 40-60°C, with an optimum temperature of 55°C and an optimum pH of 9.5 stable over a range of 5-12. Detailed understanding of the structure and insight into the activity of such lipase would aid in engineering lipases that would function in the desired extreme industrial environments. In the present study, we carried out in silico characterization and structural modeling of SAL3 which is thermoactive, alkaline and detergent-stable. Comparison of SAL3 with other staphylococcal lipases indicates that SAL3 is a true lipase having the catalytic triad (residues Ser119, Asp310 & His352) and the calcium binding site (residues Asp351, Asp354, Asp359, Asp362 and Gly286). Conservation in sequence implies that interfacial activation mechanism is possible in SAL3 with the lid formed by helix (residues 180-196) and loop (residues 197-206). Three dimensional (3D) structure model of SAL3 has been predicted for the first time and aims at understanding its function and biochemical characteristics of possessing relatively high thermal and pH stability.