Studies On BRD4 And Aurora Kinase Small Molecule Inhibitors

BRD4 plays an important role in the regulation of the cell cycle of normal mammalian cells,affecting the processes of cell proliferation,cycle,apoptosis and transcription.In single and combined use,BRD4 inhibitors present good therapeutic effects and application prospects.The Aurora kinase of the serine family can regulate the process of cell mitosis,and it has a good application prospect in combination therapy.In this thesis,the BRD4 and Aurora kinase are the targets.The design,synthesis and biological activity of indole-2-one BRD4 inhibitors and novel2,4-disubstituted pyrimidine Aurora kinase inhibitors were studied,respectively.In the first part,a series of indole-2-one compounds were designed through the drug design strategy of scaffold hoping.Specifically,5-nitroindole-2-one was used as a raw material,and 28 benzenesulfonylindole-2-one-5-amines were synthesized through addition elimination,reduction,and acylation reactions.The kinase inhibitory activity of the compounds on BRD4（D1）was measured by time-resolved fluorescence analysis（TR-FRET）,and most of the compounds displayed good enzyme activities(IC₅₀<100 nM).In the cytotoxic activity experiments,compound37j was found to exhibit good antiproliferative activities against HL-60(IC₅₀=1.35±0.33μM)and HT-29(IC₅₀=4.75±0.63μM)cells,and moderate activity in WI-38(IC₅₀=44.07±1.89μM)cells,indicating that compound 37j may have better anti-tumor cells proliferation activity and lower cytotoxicity.However,compound 37j was similar to most BET inhibitors and had no selective inhibition of BRD2 and BRD4 protein subtypes.In the study of protein interaction mode,it was found that compound 37j could occupy the KAc recognition region and WPF shelf well.In cell cycle experiments,compound 37j was found to block HT-29 cell cycle in G₁ phase.In the study of c-Myc protein expression and drug-like properties,it was found that compound 37j could reduce the expression of c-Myc protein in HT-29 cells,and it complied with the five principles of drug-like properties and showed good drug-like properties.In this study,benzenesulfonylindole-2-one-5-amines were found to be a class of effective BRD4 inhibitors,which provided theoretical basis and a summary of experience for the study of this class of antitumor drugs.In the second part,a series of 2,4-disubstituted pyrimidine compounds were designed by using hybrid drug design strategies using Revesine and SNS-314 as two Aurora kinase inhibitors as templates.Using 2,4-dichloropyrimidine as a raw material,18 2,4-disubstituted pyrimidine compounds were synthesized through substitution,reduction and acylation reactions.In the study of anti-tumor cell proliferation activities,most compounds were found to have strong antiproliferative activities in A549 and HCT-116(IC₅₀<10μM)cells.Among them,compounds 10a and 12a exhibited good activities in HCT-116 cells,with IC₅₀ values of 1.09±0.12μM and1.31±0.41μM,respectively.In the enzyme protein activity experiments,compound12a was found to have good inhibitory activity on Aurora A and B.Subsequent pharmacological experiments also showed that in HCT-116 cells,compound 12a could inhibit the phosphorylation level of Aurora kinase,and induce apoptosis by regulating the expression levels of apoptotic proteins Bcl-xl and Bax.In addition,in the study of the interaction pattern with proteins and drug-like properties,it was found that compound 12a could bind to the hinge region well,and conformed to the five principles of drug-like,and had good drug-like properties.The findings of this study have accumulated experience for the study of 2,4-disubstituted pyrimidine Aurora kinase inhibitors.