In silico investigations on the binding efficacy and allosteric mechanism of six different natural product compounds towards PTP1B inhibition through docking and molecular dynamics simulations.

Protein tyrosine phosphatase 1B (PTP1B) is a major negative regulator of both the insulin and leptin receptor phosphorylation which impacts insulin sensitivity and hence is a major therapeutic target for the treatment of type 2 diabetes and obesity. Identification of PTP1B active site inhibitors has proven to be difficult with none of them clearing the phase II clinical trials. Since the conventional methods of targeting the active site of PTP1B have failed to bring out effective PTP1B inhibitors as potential drugs, recent studies are focussing on identification of potential allosteric inhibitors of PTP1B with better specificity and activity. A complete understanding of the molecular features dynamically involved for allosteric site inhibition is still uncertain, and hence, this study is aimed at evaluating the allosteric effectiveness of six natural compounds isolated from medicinal plants which showed in vitro antidiabetic activity along with PTP1B inhibition. The allosteric binding and inhibition of these compounds are studied using computational methods such as molecular docking, homology modelling and molecular dynamics simulations for a timescale of 100 ns. The molecular dynamics simulations of native PTP1B, along with the modelled allosteric α-7 helix, for a timescale of 100 ns, revealed the spontaneous transition of the native PTP1B from open WPD loop (active) to closed WPD loop (inactive) conformations during the simulations. Similar dynamics was observed in the presence of the active site substrate pTyr (phosphotyrosine), whereas this transition was inhibited in the presence of the compounds at the allosteric site. Results of molecular dynamics simulations and principal component analysis reveal that the hindrance to WPD loop was mediated through structural interactions between the allosteric α-helical triad with Loop11 and WPD loop. The MM-PBSA (Molecular Mechanics - Poisson Boltzmann with Surface Area solvation) binding energy results along with H-bonding analysis show the possible allosteric inhibition of Aloe emodin glycoside (AEG), 3ß-taraxerol (3BT), chlorogenic acid (CGA) and cichoric acid (CHA) to be higher in comparison with (3ß)-stigmast-5-en-3-ol (SGS) and methyl lignocerate (MLG). The interaction analysis was further validated by scoring the allosteric complexes before and after MD simulations using Glide. These findings on spontaneous PTP1B fluctuations and the allosteric interactions provide a better insight into the role of PTP1B fluctuations in impacting the binding energy of allosteric inhibitors towards optimal drug designing for PTP1B. Graphical abstract.

Current trends in small molecule discovery targeting key cellular signaling events towards the combined management of diabetes and obesity.

Non-insulin dependent diabetes mellitus, also known as Type 2 diabetes is a polygenic disorder leading to abnormalities in the carbohydrate and lipid metabolism. The major contributors in the pathophysiology of type 2 diabetes (T2D) include resistance to insulin action, ß cell dysfunction, an abnormality in glucose metabolism and storage, visceral obesity and to some extent inflammation and oxidative stress. Insulin resistance, along with a defect in insulin secretion by the pancreatic ß cells is instrumental towards progression to hyperglycemia. Increased incidence of obesity is also a major contributing factor in the escalating rates of type 2 diabetes. Drug discovery efforts are therefore crucially dependent on identifying individual molecular targets and validating their relevance to human disease. The current review discusses bioactive compounds from medicinal plants offering enhanced therapeutic potential for the combined patho-physiology of diabetes and obesity. We have demonstrated that 3ß-taraxerol a pentacyclic triterpenoid (14-taraxeren-3-ol) isolated from the ethyl acetate extract of Mangifera indica, chlorogenic acid isolated from the methanol extract of Cichorium intybus, methyl tetracosanoate from the methanol extract of Costus pictus and vitalboside A derived from methanolic extract of Syzygium cumini exhibited significant effects on insulin stimulated glucose uptake causing insulin sensitizing effects on 3T3L1 adipocytes (an in vitro model mimicking adipocytes). Whereas, (3ß)-stigmast-5-en-3-ol isolated from Adathoda vasica and Aloe emodin isolated from Cassia fistula showed significant insulin mimetic effects favoring glucose uptake in L6 myotubes (an in vitro model mimicking skeletal muscle cells). These extracts and molecules showed glucose uptake through activation of PI3K, an important insulin signaling intermediate. Interestingly, cinnamic acid isolated from the hydro-alcohol extract of Cinnamomum cassia was found to activate glucose transport in L6 myotubes through the involvement of GLUT4 via the PI3K-independent pathway. However, the activation of glucose storage was effective in the presence of 3ß-taraxerol and aloe emodin though inhibition of GSK3ß activity. Therefore, the mechanism of improvement of glucose and lipid metabolism exhibited by the small molecules isolated from our lab is discussed. However, Obesity is a major risk factor for type-2 diabetes leading to destruction of insulin receptors causing insulin resistance. Identification of compounds with dual activity (anti-diabetic and antiadipogenic activity) is of current interest. The protein tyrosine phosphatase 1B (PTP1B) is an important negative regulator of the insulin and leptin-signaling pathway is of significance in target definition and discovery.

EGFR inhibition by pentacyclic triterpenes exhibit cell cycle and growth arrest in breast cancer cells.

AIMS: Pentacyclic triterpenes are a group of molecules with promising anticancer potential, although their precise molecular target remains elusive. The current work aims to investigate the antiproliferative and associated mechanisms of triterpenes in breast cancer cells in vitro. MAIN METHODS: Effect of triterpenes on cell cycle distribution, ROS and key regulatory proteins were analyzed in three breast cancer cells in vitro. Growth inhibition, new DNA synthesis, colony formation assays and Western blot analysis were performed to assess the EGFR inhibitory effect of triterpenes. Molecular docking was performed to study the interaction between EGFR and triterpenes. KEY FINDINGS: We have demonstrated the ability of dimethyl melaleucate (DMM), a pentacyclic triterpene to exhibit cell cycle arrest at G0/G1 phase by down-regulation of cyclin D1 through PI3K/AKT inhibition. Further, to identify the upstream target of DMM, potential EGFR inhibitory activity of DMM and three structurally related pentacyclic triterpenes, ursolic acid, 18α-glycyrrhetinic acid and carbenoxolone was investigated. Interestingly, pentacyclic triterpenes limit EGF mediated breast cancer proliferation through sustained inhibition of EGFR and its downstream effectors STAT3 and cyclin D1 in breast cancer lines. We also show pentacyclic triterpenes to bind at the ATP binding pocket of tyrosine kinase domain of EGFR leading to the hypothesis that pentacyclic triterpenes could be a novel class of EGFR inhibitors. In conclusion, pentacyclic triterpenes inhibit EGFR activation through binding with tyrosine kinase domain thereby suppressing breast cancer proliferation. SIGNIFICANCE: Pentacyclic triterpenes may serve as a potential platform for development of novel drugs against breast cancer.