With the rapid development of computational techniques and hardware, more rigorous and precise theoretical models have been used to predict the binding affinities of a large number of small molecules to biomolecules. By employing continuum solvation models, the MM/GBSA and MM/PBSA methodologies achieve a good balance between low computational cost and reasonable prediction accuracy. In this study, we have thoroughly investigated the effects of interior dielectric constant, molecular dynamics (MD) simulations, and the number of top-scored docking poses on the performance of the MM/GBSA and MM/PBSA rescoring of docking poses for three tyrosine kinases, including ABL, ALK, and BRAF. Overall, the MM/PBSA and MM/GBSA rescoring achieved comparative accuracies based on a relatively higher solute (or interior) dielectric constant (i.e. ε = 2, or 4), and could markedly improve the 'screening power' and 'ranking power' given by Autodock. Moreover, with a relatively higher solute dielectric constant, the MM/PBSA or MM/GBSA rescoring based on the best scored docking poses and the multiple top-scored docking poses gave similar predictions, implying that much computational cost can be saved by considering the best scored docking poses only. Besides, compared with the rescoring based on the minimized structures, the rescoring based on the MD simulations might not be completely necessary due to its negligible impact on the docking performance. Considering the much higher computational demand of MM/PBSA, MM/GBSA with a high solute dielectric constant (ε = 2 or 4) is recommended for the virtual screening of tyrosine kinases.

Hemorrhagic stroke is a common and severe neurological disorder and is associated with high rates of mortality and morbidity, especially for intracerebral hemorrhage (ICH). Increasing evidence demonstrates that oxidative stress responses participate in the pathophysiological processes of secondary brain injury (SBI) following ICH. The mechanisms involved in interoperable systems include endoplasmic reticulum (ER) stress, neuronal apoptosis and necrosis, inflammation, and autophagy. In this review, we summarized some promising advances in the field of oxidative stress and ICH, including contained animal and human investigations. We also discussed the role of oxidative stress, systemic oxidative stress responses, and some research of potential therapeutic options aimed at reducing oxidative stress to protect the neuronal function after ICH, focusing on the challenges of translation between preclinical and clinical studies, and potential post-ICH antioxidative therapeutic approaches.

SUMMARYAdult mammalian cardiomyocytes have little capacity to proliferate in response to injury, a deficiency that underlies the poor regenerative ability of human hearts after myocardial infarction. By contrast, zebrafish regenerate heart muscle after trauma by inducing proliferation of spared cardiomyocytes, providing a model for identifying manipulations that block or enhance these events. Although direct genetic or chemical screens of heart regeneration in adult zebrafish present several challenges, zebrafish embryos are ideal for high-throughput screening. Here, to visualize cardiomyocyte proliferation events in live zebrafish embryos, we generated transgenic zebrafish lines that employ fluorescent ubiquitylation-based cell cycle indicator (FUCCI) technology. We then performed a chemical screen and identified several small molecules that increase or reduce cardiomyocyte proliferation during heart development. These compounds act via Hedgehog, Insulin-like growth factor or Transforming growth factor  signaling pathways. Direct examination of heart regeneration after mechanical or genetic ablation injuries indicated that these pathways are activated in regenerating cardiomyocytes and that they can be pharmacologically manipulated to inhibit or enhance cardiomyocyte proliferation during adult heart regeneration. Our findings describe a new screening system that identifies molecules and pathways with the potential to modify heart regeneration. DEVELOPMENT Drug treatmentsSAG (EMD Chemicals) was either purchased in solution or dissolved in water to a 10 mM stock solution. NBI-31772 (Calbiochem) was dissolved in DMSO for a 30 mM stock solution. SU5402 (Santa Cruz Biotechnology), -secretase inhibitor (Calbiochem), SB-431542 (Selleckchem), RA (Sigma-Aldrich) and DEAB (Sigma-Aldrich) were dissolved in DMSO to a final concentration of 10 mM. CyA (Toronto Research Chemicals), NVP-AEW541 (Cayman Chemical), and GSK-3 Inhibitor IX (BIO, Calbiochem) were dissolved in ethanol to stock solutions of 10 mM. For embryo studies, cmlc2:FUCCI embryos were collected within a 30-minute window from male-female pairings. From 1 day post-fertilization (dpf) onwards, embryos were treated with n-phenylthiourea (PTU; SigmaAldrich) to block development of pigmentation. Drug treatment was performed at 3-4 dpf in 3 ml of buffered embryo water in 6-well plates. For adult studies, injured adult fish were treated in 20 ml of buffered fish water per animal in finger bowls. For embryos, all drugs were used at 5 M except for NBI-31772, RA and BIO, which were used at 2.5 M. In adults, SAG was used at 2.5 µM, NBI-31772 was used at 10 M, CyA was used at 10 M and NVP-AEW541 was used at 2 M. HistologyInitial screening for FUCCI signals was performed using a Leica M205 FA dissecting microscope. z-stacks were acquired from 50 M-thick sections of paraformaldehyde (PFA)-fixed embryos using a Zeiss LSM 700 confocal microscope. Three-dimensional reconstructions were made using Imaris Version 7.3 software. Quantification of mCherry + and/or Venus + n...

Background The cisplatin-resistance is still a main course for chemotherapy failure of lung cancer patients. Cisplatin-resistant cancer cells own higher malignance and exhibited increased metastatic ability, but the mechanism is not clear. In this study, we investigated the effects of Ataxia Telangiectasia Mutated (ATM) on lung cancer metastasis. Materials and methods Cisplatin-resistant A549CisR and H157CisR cell line were generated by long-term treating parental A549 and H157 cells (A549P and H157P) with cisplatin. Cell growth, cell migration and cell invasion were determined. Gene expressions were determined by Western Blot and qPCR. Tumor metastasis was investigated using a xenograft mouse model. Results The IC50 of the cisplatin-resistant cells (A549CisR and H157CisR cells) to cisplatin was 6–8 higher than parental cells. The A549CisR and H157CisR cells expressed lower level of E-cadherin and higher levels of N-cadherin, Vimentin and Snail compared to the parental A549P and H157P cells, and exhibited stronger capabilities of metastatic potential compared to the parental cells. The ATM expression was upregulated in A549CisR and H157CisR cells and cisplatin treatment also upregulated expression of ATM in parental cells, The inhibition of ATM by using specific ATM inhibitor CP466722 or knock-down ATM by siRNA suppressed Epithelial-to-Mesenchymal transition (EMT) and metastatic potential of A549CisR and H157CisR cells. These data suggest that ATM mediates the cisplatin-resistance in lung cancer cells. Expressions of JAK 1,2, 、 STAT 3 、PD-L1 and ATM were increased in A549CisR and H157CisR cells and could by induced by cisplatin in parental lung cancer cells. Interestedly, ATM upregulated PD-L1 expression via JAK 1,2 /STAT 3 pathway and inhibition of ATM decreased JAK/STAT3 signaling and decreased PD-L1 expression. The treatment of PD-L1 neutralizing Ab reduced EMT and cell invasion. Inhibition of JAK 1,2 /STAT 3 signaling by specific inhibitors suppressed ATM-induced PD-L1 expression, EMT and cell invasion. Importantly, inhibition of ATM suppressed EMT and tumor metastasis in cisplatin-resistant lung cancer cells in an orthotopic xenograft mouse model. Conclusions Our results show that ATM regulates PD-L1 expression through activation of JAK/STAT3 signaling in cisplatin-resistant cells. Overexpression of ATM contributes to cisplatin-resistance in lung cancer cells. Inhibition of ATM reversed EMT and inhibited cell invasion and tumor metastasis. Thus, ATM may be a potential target for the treatment of cisplatin-resistant lung cancer. Electronic supplementary material The online version of this article (10.1186/s13046-019-1161-8) contains supplementary material, which is available to authorized users.

As model organism-based research shifts from forward to reverse genetics approaches, largely due to the ease of genome editing technology, a low frequency of abnormal phenotypes is being observed in lines with mutations predicted to lead to deleterious effects on the encoded protein. In zebrafish, this low frequency is in part explained by compensation by genes of redundant or similar function, often resulting from the additional round of teleost-specific whole genome duplication within vertebrates. Here we offer additional explanations for the low frequency of mutant phenotypes. We analyzed mRNA processing in seven zebrafish lines with mutations expected to disrupt gene function, generated by CRISPR/Cas9 or ENU mutagenesis methods. Five of the seven lines showed evidence of altered mRNA processing: one through a skipped exon that did not lead to a frame shift, one through nonsense-associated splicing that did not lead to a frame shift, and three through the use of cryptic splice sites. These results highlight the need for a methodical analysis of the mRNA produced in mutant lines before making conclusions or embarking on studies that assume loss of function as a result of a given genomic change. Furthermore, recognition of the types of adaptations that can occur may inform the strategies of mutant generation.

To investigate whether IL‐6 signaling affects the susceptibility of castration‐resistant prostate cancer (CRPC) cells to cytotoxic action of natural killer (NK) cells, CRPC cell lines (having different IL‐6 levels) were developed by lentiviral transduction. While observing no secreted IL‐6 level in parental C4‐2 and CWR22Rv1 cells, we found the IL‐6 expression/secretion in these cells was induced after the transduction process and the IL‐6 level difference in C4‐2siIL‐6/sc and CWR22siIL‐6/sc cell CRPC cell sets could be detected. We then found that IL‐6‐knockdown cells were more susceptible to NK cell cytotoxicity than control cells due to lowered programmed death receptor ligand 1 (PD‐L1) and increased NK group 2D (NKG2D) ligand levels. In animal studies, to concur with the in vitro results, we found that IL‐6‐expressing cell‐derived tumors were more resistant to NK cell action than the tumors of IL‐6‐knockdown cells. Further, we discovered that JAK‐Stat3 is the most critical IL‐6 downstream signaling that modulates PD‐L1/NKG2D ligand levels in CRPC cells. Furthermore, inhibition of the JAK or Stat3 signaling effectively increased the susceptibility of C4‐2sc and CWRsc cells to NK cell cytotoxicity. We observed the most effective cytotoxicity when the PD‐L1 Ab and JAK inhibitor (or Stat 3 inhibitor) were used together. These results suggest that the strategy of targeting IL‐6 signaling (or its downstream signaling) may enhance the NK cell‐mediated immune action to CRPC tumors, thus yielding clinical implications in developing future immunotherapeutics of exploiting this strategy to treat patients with CRPC.

How does a type II inhibitor bind to/unbind from a kinase target is still a confusing question because the small molecule occupies both the ATP pocket and the allosteric pocket of the kinase binding site. Here, by using enhanced sampling simulations (umbrella sampling, US) and two-end-state free energy calculations (MM/GSBA), we systemically studied the dissociation processes of two distinct small molecules escaping from the binding pocket of p38 MAP kinase through the allosteric channel and the ATP channel. The results show that the unbinding pathways along the allosteric channel have much lower PMF depths than those along the ATP channel, suggesting that the allosteric channel is more favorable for the dissociations of the two inhibitors and thereby supporting the general understanding that the largest channel of a target is usually the entry/exit pathway for the binding/dissociation of small molecules. Interestingly, the MM/GBSA approach yielded similar PMF profiles compared with those based on US, a much time consuming approach, indicating that for a general study, such as detecting the important transition state of a ligand binding/unbinding process, MM/GBSA may be a feasible choice.

BackgroundIn this work, we analyzed and compared the distribution profiles of a wide variety of molecular properties for three compound classes: drug-like compounds in MDL Drug Data Report (MDDR), non-drug-like compounds in Available Chemical Directory (ACD), and natural compounds in Traditional Chinese Medicine Compound Database (TCMCD).ResultsThe comparison of the property distributions suggests that, when all compounds in MDDR, ACD and TCMCD with molecular weight lower than 600 were used, MDDR and ACD are substantially different while TCMCD is much more similar to MDDR than ACD. However, when the three subsets of ACD, MDDR and TCMCD with similar molecular weight distributions were examined, the distribution profiles of the representative physicochemical properties for MDDR and ACD do not differ significantly anymore, suggesting that after the dependence of molecular weight is removed drug-like and non-drug-like molecules cannot be effectively distinguished by simple property-based filters; however, the distribution profiles of several physicochemical properties for TCMCD are obviously different from those for MDDR and ACD. Then, the performance of each molecular property on predicting drug-likeness was evaluated. No single molecular property shows good performance to discriminate between drug-like and non-drug-like molecules. Compared with the other descriptors, fractional negative accessible surface area (FASA-) performs the best. Finally, a PCA-based scheme was used to visually characterize the spatial distributions of the three classes of compounds with similar molecular weight distributions.ConclusionIf FASA- was used as a drug-likeness filter, more than 80% molecules in TCMCD were predicted to be drug-like. Moreover, the principal component plots show that natural compounds in TCMCD have different and even more diverse distributions than either drug-like compounds in MDDR or non-drug-like compounds in ACD.