靶向FLT3的抗肿瘤小分子抑制剂的虚拟筛选

摘要 目的：本研究旨在利用虚拟筛选技术,从ChEMBL数据库中发现新型FLT3抑制剂,为靶向FLT3的小分子抑制剂的开发提供理论基础。方法：选取ChEMBL数据库中约240万个小分子作为数据集,以FLT3蛋白为靶标,通过分子对接进行虚拟筛选,对筛选得到的目标化合物进行200 ns的分子动力学模拟,研究其与FLT3蛋白之间的结合能力和稳定性。结果：通过虚筛选成功发现了5个未见文献报道的新型潜在FLT3抑制剂（ChEMBL ID: 5186572、4845881、2151842、3642822、3916042）,对接分数（-10.93 ~ -12.58）和结合自由能（-82.06 ~ -88.49 kcal/mol）均优于参照组Gilteritinib（-8.73和-65.38 kcal/mol）;分子动力学模拟结果显示,目标化合物与FLT3蛋白均具有较强的结合能力,且与FLT3蛋白形成的复合物具有良好的稳定性。结论：本研究利用虚拟筛选技术成功发现了5个未见文献报道的具有潜在抗肿瘤活性的FLT3抑制剂,为新一代FLT3抑制剂的研发提供了重要的理论基础。     

Identification of a novel selective small-molecule inhibitor of protein arginine methyltransferase 5 (PRMT5) by virtual screening,resynthesis and biological evaluations

As one of the most promising anticancer target in protein arginine methyltransferase (PRMT) family, PRMT5 has been drawing more and more attentions, and many efforts have been devoted to develop its inhibitors. In this study, three PRMT5 inhibitors (9, 16, and 23) with novel scaffolds were identified by performing pharmacophore- and docking-based virtual screening combined with in vitro radiometric-based scintillation proximity assay (SPA). Substructure search based on the scaffold of the most active 9 afforded 26 additional analogues, and SPA results indicated that two analogues (9–1 and 9–2) showed increased PRMT5 inhibitory activity compared with the parental compound. Resynthesis of 9, 9–1, and 9–2 confirmed their PRMT5 enzymatic inhibition activity. In addition, compound 9–1 displayed selectivity against PRMT5 over other key homological members (PRMT1 and CARM1 (PRMT4)). While the structure–activity relationship (SAR) of this series of compounds was discussed to provide clues for further structure optimization, the probable binding modes of active compounds were also probed by molecular docking and molecular dynamics simulations. Finally, the antiproliferative effect of 9–1 on MV4-11 leukemia cell line was confirmed and its impact on regulating the target gene of PRMT5 was also validated. The hit compounds identified in this work have provided more novel scaffolds for future hit-to-lead optimization of small-molecule PRMT5 inhibitors.     

设计与筛选靶向PI3Kα的去氢骆驼蓬碱衍生物及其分子动力学验证

以PI3Kα作为靶蛋白,采用计算机辅助设计的方法优化去氢骆驼蓬碱结构,获得与靶蛋白结合力较高的化合物。在此基础上,利用分子对接,三维定量构效关系(3D-QSAR)和分子动力学技术验证了修饰化合物与靶蛋白的结合能力和稳定性。最后筛选出与靶标具有更好结合力的潜在去氢骆驼蓬碱结构衍生物。使用ACD/Percepta软件对去氢骆驼蓬碱的母核进行结构修饰,获得136 745个化合物的化合物库。通过Schrdinger软件从化合物库中依据MM/GBSA(分子力学广义玻恩表面积)筛选出结合能排在前三的化合物并使用3D-QSAR进行验证,最后通过分子动力学方法验证了三种结构衍生物,并预测了ADMET性质。通过软件设计、虚拟筛选、3D-QSAR模型构建和分子动力学模拟等一系列计算机辅助设计方法,获得了136 745种新的去氢骆驼蓬碱衍生物,评价了蛋白质结构复合物的稳定性,最终获得了3种结合能较高的化合物。其中,发现编号为37971的衍生物具有更好的3D-QSAR模型活性预测值与TPSA(拓扑极性表面积)值(87.84)和生物利用度(33.21%)。本研究结果为去氢骆驼蓬碱的结构修饰提供了设计思路,获得理论上与PI3Kα具有较好的靶向结合能力的化合物,通过分子对接和分子动力学验证,所获得的分子有望成为潜在的候选化合物,为进一步的实验验证奠定基础。     

The discovery of novel and selective fatty acid binding protein 4 inhibitors by virtual screening and biological evaluation

Adipocyte fatty acid binding protein (AFABP, FABP4) has been proven to be a potential therapeutic target for diabetes, atherosclerosis and inflammation-related diseases. In this study, a series of new scaffolds of small molecule inhibitors of FABP4 were identified by virtual screening and were validated by a bioassay. Fifty selected compounds were tested, which led to the discovery of seven hits. Structural similarity-based searches were then performed based on the hits and led to the identification of one high affinity compound 33b (K_i = 0.29 ± 0.07 μM, ΔT_m = 8.5 °C). This compound’s effective blockade of inflammatory response was further validated by its ability to suppress pro-inflammatory cytokines induced by lipopolysaccharide (LPS) stimulation. Molecular dynamics simulation (MD) and mutagenesis studies validated key residues for its inhibitory potency and thus provide an important clue for the further development of drugs.     

Discovery of novel inhibitors of vascular endothelial growth factor-A–Neuropilin-1 interaction by structure-based virtual screening

Neuropilin-1 (NRP-1), one of the most important co-receptors of vascular endothelial growth factor-A (VEGF-A), increases its angiogenic action in several chronic diseases including cancer by increasing the activity of associated tyrosine kinase receptors, VEGFR1 and VEGFR2. Binding of VEGF-A to NRP-1 plays a critical role in pathological angiogenesis and tumor progression. Today, targeting this interaction is a validated approach to fight against angiogenesis-dependent diseases. Only anti-NRP-1 antibodies, peptide and peptidomimetic drug-candidates or hits have been developed thus far. In order to identify potent orally active small organic molecules various experimental and in silico approaches can be used. Here we report, novel promising small drug-like molecules disrupting the binding of VEGF-A₁₆₅ to NRP-1. We carried out structure-based virtual screening experiments using the ChemBridge compound collection on the VEGF-A₁₆₅ binding pocket of NRP-1. After docking and two rounds of similarity search computations, we identified 4 compounds that inhibit the biotinylated VEGF-A₁₆₅ binding to recombinant NRP-1 with K_i of about 10 μM. These compounds contain a common chlorobenzyloxy alkyloxy halogenobenzyl amine scaffold that can serve as a base for further development of new NRP-1 inhibitors.     

Identification of (4-(9H-fluoren-9-yl) piperazin-1-yl) methanone derivatives as falcipain 2 inhibitors active against Plasmodium falciparum cultures

Background

Falcipain 2 (FP-2) is the hemoglobin-degrading cysteine protease of Plasmodium falciparum most extensively targeted to develop novel antimalarials. However, no commercial antimalarial drugs based on FP-2 inhibition are available yet due to the low selectivity of most FP-2 inhibitors against the human cysteine proteases.

Quantification and cell-to-cell variation of vascular endothelial growth factor receptors

The vascular endothelial growth factor receptors (VEGFR) play a significant role in angiogenesis, the formation of new blood vessels from existing vasculature. Systems biology offers promising approaches to better understand angiogenesis by computational modeling the key molecular interactions in this process. Such modeling requires quantitative knowledge of cell surface density of pro-angiogenic receptors versus anti-angiogenic receptors, their regulation, and their cell-to-cell variability. Using quantitative fluorescence, we systematically characterized the endothelial surface density of VEGFRs and neuropilin-1 (NRP1). We also determined the role of VEGF in regulating the surface density of these receptors. Applying cell-by-cell analysis revealed heterogeneity in receptor surface density and VEGF tuning of this heterogeneity. Altogether, we determine inherent differences in the surface expression levels of these receptors and the role of VEGF in regulating the balance of anti-angiogenic or modulatory (VEGFR1) and pro-angiogenic (VEGFR2) receptors.     

Structural studies of human purine nucleoside phosphorylase: towards a new specific empirical scoring function

Human purine nucleoside phosphorylase (HsPNP) is a target for inhibitor development aiming at T-cell immune response modulation. In this work, we report the development of a new set of empirical scoring functions and its application to evaluate binding affinities and docking results. To test these new functions, we solved the structure of HsPNP and 2-mercapto-4(3H)-quinazolinone (HsPNP:MQU) binary complex at 2.7 Å resolution using synchrotron radiation, and used these functions to predict ligand position obtained in docking simulations. We also employed molecular dynamics simulations to analyze HsPNP in two conditions, as apoenzyme and in the binary complex form, in order to assess the structural features responsible for stability. Analysis of the structural differences between systems provides explanation for inhibitor binding. The use of these scoring functions to evaluate binding affinities and molecular docking results may be used to guide future efforts on virtual screening focused on HsPNP.     

Discovery of novel dengue virus entry inhibitors via a structure-based approach

Dengue is a mosquito-borne virus that has become a major public health concern worldwide in recent years. However, the current treatment for dengue disease is only supportive therapy, and no specific antivirals are available to control the infections. Therefore, the need for safe and effective antiviral drugs against this virus is of utmost importance. Entry of the dengue virus (DENV) into a host cell is mediated by its major envelope protein, E. The crystal structure of the E protein reveals a hydrophobic pocket occupied by the detergent n-octyl-β-d-glucoside (β-OG) lying at a hinge region between domains I and II, which is important for the low-pH-triggered conformational rearrangement required for fusion. Thus, the E protein is an attractive target for the development of antiviral agents. In this work, we performed prospective docking-based virtual screening to identify small molecules that likely bind to the β-OG binding site. Twenty-three structurally different compounds were identified and two of them had an EC₅₀ value in the low micromolar range. In particular, compound 2 (EC₅₀ = 3.1 μM) showed marked antiviral activity with a good therapeutic index. Molecular dynamics simulations were used in an attempt to characterize the interaction of 2 with protein E, thus paving the way for future ligand optimization endeavors. These studies highlight the possibility of using a new class of DENV inhibitors against dengue.     

Search for Novel Remedies to Augment Radiation Resistance of Inhabitants of Fukushima and Chernobyl Disasters: Identifying DNA Repair Protein XRCC4 Inhibitors

Abstract

Two nuclear plant disasters occurring within a span of 25 years threaten health and genome integrity both in Fukushima and Chernobyl. Search for remedies capable of enhancing DNA repair efficiency and radiation resistance in humans appears to be a urgent problem for now. XRCC4 is an important enhancer in promoting repair pathway triggered by DNA double-strand break (DSB). In the context of radiation therapy, active XRCC4 could reduce DSB-mediated apoptotic effect on cancer cells. Hence, developing XRCC4 inhibitors could possibly enhance radiotherapy outcomes. In this study, we screened traditional Chinese medicine (TCM) database, TCM Database@Taiwan, and have identified three potent inhibitor agents against XRCC4. Through molecular dynamics simulation, we have determined that the protein-ligand interactions were focused at Lys188 on chain A and Lys187 on chain B. Intriguingly, the hydrogen bonds for all three ligands fluctuated frequently but were held at close approximation. The pi-cation interactions and ionic interactions mediated by o-hydroxyphenyl and carboxyl functional groups respectively have been demonstrated to play critical roles in stabilizing binding conformations. Based on these results, we reported the identification of potential radiotherapy enhancers from TCM. We further characterized the key binding elements for inhibiting the XRCC4 activities.     

Structure–function analysis of VEGF receptor activation and the role of coreceptors in angiogenic signaling

Vascular endothelial growth factors (VEGFs) constitute a family of six polypeptides, VEGF-A, -B, -C, -D, -E and PlGF, that regulate blood and lymphatic vessel development. VEGFs specifically bind to three type V receptor tyrosine kinases (RTKs), VEGFR-1, -2 and -3, and to coreceptors such as neuropilins and heparan sulfate proteoglycans (HSPG). VEGFRs are activated upon ligand-induced dimerization mediated by the extracellular domain (ECD). A study using receptor constructs carrying artificial dimerization-promoting transmembrane domains (TMDs) showed that receptor dimerization is necessary, but not sufficient, for receptor activation and demonstrates that distinct orientation of receptor monomers is required to instigate transmembrane signaling. Angiogenic signaling by VEGF receptors also depends on cooperation with specific coreceptors such as neuropilins and HSPG. A number of VEGF isoforms differ in binding to coreceptors, and ligand-specific signal output is apparently the result of the specific coreceptor complex assembled by a particular VEGF isoform. Here we discuss the structural features of VEGF family ligands and their receptors in relation to their distinct signal output and angiogenic potential.