Discovery Of Small Molecule Inhibitors Targeting Tumor-associated Proteins BRD4 And RhoA And Investigation Of Mechanisms

In recent years, with the rapid development of the biomedical field, targeting specific proteins to screen their small molecule inhibitors has become the focus of this field. In this paper, we selected the epigenetics-related protein and cytoskeletal-related protein to screen for small molecules.As an important post-translation modification in epigenetics, histone acetylation not only plays an important role in regulating chromatin structure; gene transcription and many other fields, but also influences the occurrences and developments of cancers. Thus, histone acetylation has become a hot research at home and abroad. As a histone acetylation recognition factor, BRD4 is a member of the bromodomain and extra-terminal (BET) family, which can recognize and bind to the acetylation of lysine residues, recruits the transcription factors and regulates downstream oncogenes such as BCL2, CDK6, c-Myc and so on. Current studies has shown that BRD4 is abnormal expression in acute myelocytic leukemia, mixed-lineage leukemia, testis midline carcinoma and many other diseases, and BRD4 has become a definite drug target. High throughput screening method for BRD4 has developed based on Alphascreen technology and a series of novel small molecule inhibitors with good activities was obtained with preliminary screening, the lead compound can inhibit BRD4 protein to recognize acetylated histones significantly in vitro at 0.81±0.03μM.And the lead compound can inhibit the proliferation of the leukemia cells.As a number of GTPase, RhoA importantly mediate cytoskeleton , and is a member of the small G protein family, which can interact with the three upstream proteins and regulated by them, and thus converting between RhoA-GTP active state and RhoA-GDP inactive state. Futher research shows high expression of RhoA is closely related to the development of many kinds of tumors, such as breast cancer;prostate cancer; testicular cancer; lung cancer and so on. RhoA is a target protein contacting with different tumor types, and it is valuable and significant for us to find small molecules inhibitors in the treatment of cancer. High-throughput screening method was developed based protein thermal shift assay and the hit compound with inhibitory was screened, which can inhibit the enzyme activity of RhoA as well as the binding of RhoA and GEF proteins. Futhermore, the lead compound is a chemical probe and can reveal the variational mechanisms of RhoA.