Exploring the effect of E76K mutation on SHP2 cause gain-of-function activity by a molecular dynamics study.

Juvenile myelomonocytic leukemia (JMML), an invasive myeloproliferative neoplasm, is a childhood disease with very high clinical lethality. Somatic mutation E76K in SHP2 is the most commonly identified mutation found in up to 35% of patients with JMML. To investigate the effect of gain-of-function mutation-E76K on SHP2 activity, molecular dynamic simulations on the wild-type SHP2 (SHP2-WT) system and the mutated E76K (SHP2-E76K) system were performed. The evaluation of stability of these two systems indicated that the simulated trajectories were stable after simulation for 3 nanoseconds. The root mean square fluctuation and the per-residue root mean square deviation illustrated that there were two regions (residues Tyr 81-Glu 83 and Glu 258-Leu 261) in the wild-type system and the mutated system, which had large differences. The principal component analysis, dynamic cross correlation maps analysis, as well as secondary structure analysis suggested that the mutated E76K impacted the movement of these two regions in SHP2 protein. Furthermore, residue interaction network analysis, hydrogen bond occupancy, and binding free energies analysis were used to explain how the two regions were specifically affected by the mutant. The results indicated that the primary variances between SHP2-WT and SHP2-E76K were the different interactions between Glu/Lys 76 and Arg 265, Tyr 80 and Leu 77, Leu 77 and Tyr 81, Thr 73 and Glu 258, Ala 75 and Cys 259, Phe 71 and Tyr 81, Ala 75 and Glu 258, and Tyr 73 and Glu/Lys 76. Consequently, these findings here might provide insights into the increased activity in SHP2-E76K.

Puerarin regulates neovascular glaucoma through pigment epithelium­derived growth factor­induced NF­κB signaling pathway.

Neovascular glaucoma is an ophthalmic disease and a potentially blinding secondary glaucoma caused by the formation of abnormal new blood vessels on the iris, which can prevent the normal drainage of water from the anterior segment of the eye. Evidence from China has suggested that puerarin benefits many diseases including myocardial infarction, stable angina, cerebral ischemia and glaucoma in a clinical setting. In the present study, the aim was to investigate the efficacies of puerarin on neovascular glaucoma in a mouse model. The molecular mechanism of puerarin­mediated treatment for neovascular glaucoma was also investigated both in vitro and in vivo. Inflammatory responses in mice with neovascular glaucoma were analyzed by western blotting. Oxidative stress levels were investigated following treatment with puerarin in a mouse model of neovascular glaucoma. The results indicated that puerarin markedly improved growth of vascular endothelial cells. The present study reported that puerarin treatment markedly decreased interleukin (IL)­1ß, IL­17A and tumor necrosis factor­α expression levels in mice with neovascular glaucoma. It was found that puerarin significantly decreased oxidative stress levels by reducing reactive oxygen species, superoxide dismutase and malondialdehyde levels, as well as neuronal nitric oxide synthase (NOS) and inducible NOS expression levels. Results indicated that expression levels of pigment epithelium­derived growth factor were significantly inhibited following treatment with puerarin. Mechanism analysis demonstrated that treatment with puerarin effectively inhibited nuclear factor (NF)­κB activity and its target protein levels p65, inhibitor of NF­κB kinase subunit ß and inhibitor of NF­κB kinase subunit α in vascular endothelial cells. Increasing endothelial­derived growth factor (EDGF) expression levels could stimulate NF­κB activity and abolish the inhibitory effects of puerarin. An animal study reported that puerarin treatment presented therapeutic effects for mice with neovascular glaucoma. Numbers of new vessels in iris were recovered to normal following puerarin treatment. In conclusion, these results indicated that puerarin treatment can inhibit inflammatory responses and oxidative stress, platelet­derived growth factor (PDGF) expression and NF­κB activity, suggesting puerarin may be a potential agent for the treatment of neovascular glaucoma through PDGF­induced NF­κB signaling pathway.

Synthesis, bioactivity, 3D-QSAR studies of novel dibenzofuran derivatives as PTP-MEG2 inhibitors.

PTP-MEG2 plays a critical role in the diverse cell signalling processes, so targeting PTP-MEG2 is a promising strategy for various human diseases treatments. In this study, a series of novel dibenzofuran derivatives was synthesized and assayed for their PTP-MEG2 inhibitory activities. 10a with highest inhibitory activity (320 nM) exhibited significant selectivity for PTP-MEG2 over its close homolog SHP2, CDC25 (IC50 > 50 µM). By means of the powerful ''HipHop'' technique, a 3D-QSAR study was carried out to explore structure activity relationship of these molecules. The generated pharmacophore model revealed that the one RA, three Hyd, and two HBA features play an important role in binding to the active site of the target protein-PTP-MEG2. Docking simulation study indicated that 10a achieved its potency and specificity for PTP-MEG2 by targeting unique nearby peripheral binding pockets and the active site. The absorption, distribution, metabolism and excretion (ADME) predictions showed that the 11 compounds hold high potential to be novel lead compounds for targeting PTP-MEG2. Our findings here can provide a new strategy or useful insights for designing the effective PTP-MEG2 inhibitors.

Synthesis, biological evaluation and 3D-QSAR studies of imidazolidine-2,4-dione derivatives as novel protein tyrosine phosphatase 1B inhibitors.

Protein tyrosine phosphatase 1B (PTP1B) plays a vital role in the regulation of insulin sensitivity and dephosphorylation of the insulin receptor, so PTP1B inhibitors may be potential agents to treat type 2 diabetes. In this work, a series of novel imidazolidine-2,4-dione derivatives were designed, synthesized and assayed for their PTP1B inhibitory activities. These compounds exhibited potent activities with IC50 values at 0.57-172 µM. A 3D-QSAR study using CoMFA and CoMSIA techniques was carried out to explore structure activity relationship of these molecules. The CoMSIA model was more predictive with q(2) = 0.777, r(2) = 0.999, SEE = 0.013 and r(2)pred = 0.836, while the CoMFA model gave q(2) = 0.543, r(2) = 0.998, SEE = 0.029 and r(2)pred = 0.754. The contour maps derived from the best CoMFA and CoMSIA models combined with docking analysis provided good insights into the structural features relevant to the bioactivity, and could be used in the molecular design of novel imidazolidine-2,4-dione derivatives.

Yang-Dan-Tang, Identified from 15 Chinese Herbal Formulae, Inhibits Human Lung Cancer Cell Proliferation via Cell Cycle Arrest.

Lung cancer has long been one of the most deadly forms of cancer. The majority of lung cancers are of the non-small-cell lung cancer (NSCLC) type. Here we used the non-small-cell lung carcinoma cell line A549 to screen 15 different traditional Chinese herbal medicine (CHM) formulae to explore the possible mechanisms of alternative medicine in lung cancer therapy. We identified three formulae (Formulae 3, 5, and 14) that substantially decreased the survival of A549 cells but did not affect MRC5 normal lung tissue cells. Formula 14, Yang-Dan-Tang, a modified decoction of Ramulus Cinnamomi Cassiae, was chosen for further characterization. Flow cytometry analysis showed that treatment of Formula 14 induced cell cycle arrest in G1 and G2 phase without causing significant cell death. These results were also confirmed by Western blot analysis, with decreased expression of G1/S and G2/M promoting cell cycle machinery including cyclin D3, cyclin B1, CDK4, and CDK6. This study provides further insight into the possible working mechanism of Yang-Dan-Tang in patients.