Molecular Simulations Based On Selective CDK9 Inhibitors

Cyclin dependent kinase(CDK)9 is a member of the CDKs family.Unlike many other CDKs,CDK9 is not directly involved in the regulation of cell cycle.Instead,it binds to the subunit cyclin T1 to form the positive transcription elongation factor b(P-TEFb),and then participate in the regulation of gene transcription by the phosphorylation of RNA polymerase II.Therefore,CDK9 is considered as a potential therapeutic target in oncology,virology,cardiology and inflammation.The identification of selective CDK9 inhibitors is of great significance not only for understanding its biological functions,but also for establishing safe and effective therapies for cancer treatment.However,since the CDKs are highly conserved in their ATP binding sites,the discovery of selective CDK inhibitors has always been a crucial task and a critical challenge for medicinal chemists.Herein,a variety of molecular simulation methods were applied to a range of selective CDK9 inhibitors reported previously.There are four chapters in this dissertation,which involves different aspects on molecular modeling of CDK9 inhibitors.In the first chapter,the research highlights on kinases and CDK9 were reviewed,and molecular simulation strategies previously applied to kinase inhibitors were also introduced.In particular,the applications of molecular simulation methods on selective kinase inhibitors were emphasized.In the second chapter,in silico model-based virtual screening was exploited to search for novel CDK9 inhibitors,especially selective inhibitors.Bayesian classification models were first established on the basis of known CDK9 inhibitors,and a complex-based pharmacophore was then constructed according to the crystal structures of CDK9-inhibitors.Both models were used for the preliminary screening,and then molecular docking was applied for further refining.Forty-six compounds were prioritized for biological evaluation,in which eight compounds showed percentage inhibition above 50%under the concentration of 100 ?mol/L or 10 ?mol/L.The most active compound C-33 showed IC50 values of 0.22 ?mol/L and 3.5 ?mol/L against CDK9 and CDK2,respectively.In the third chapter,a variety of molecular simulation methods,including quantitative structure-selectivity relationship(QSSR),protein-protein interaction and allosteric inhibition studies,were attempted to model selective CDK9 inhibitors.In the first section,2D-QSSR models with good predictive ability for external test set were established based on the reported selective data against CDK9/2.Analysis of the molecular descriptors sheds lights on the structural basis for selective inhibition and provides crucial clues for further molecular design and optimization of selective CDK9 inhibitors.In the second section,starting from a known class of small molecules that potentially blocks the protein-protein interaction between Cyclin T1 and Tat,molecular docking was used to determine the possible acting sites of these compounds on a newly resolved super elongation complex(SEC)crystal.Based on the interaction modes revealed by docking,a library was generated by fragment-replacement on the active compound C3 and screened to give potential selective CDK9 inhibitors through blocking PPI.In the third section,CDK9 DFG-out conformation model was established,which provides a structural basis for further screening and design of type 2 CDK9 inhibitors.The sequence alignment of amino acids and the comparation of three-dimensional structures based on the type 2 binding sites in CDKs were also carried out to elucidate the selective basis for type 2 CDK inhibitors.In the fourth chapter,molecular modeling was applied to indolin-2-ones previously prepared in our group to explain the loss of CDK9 inhibitory activity in this compound class.Flow cytometry and immunochemical assays were performed to explore the anti-tumor mechanism of these compounds.In summary,a variety of molecular simulation methods were applied to CDK9 inhibitors,which provides conducive clues for further discovery and optimization of selective CDK9 inhibitors.