Design,Synthesis And Anti-tumor Activity Evaluation Of Novel Histone Deacetylase Inhibitors

As an epigenetic regulation of gene expression,there are many ways to covalently modify histones,including acetylation,methylation,phosphorylation,ubiquitination,sumoylation and so on.Acetylation is a common post-translational modification of proteins.Histone acetylation can dissociate DNA from histone octamer,relax the structure of nucleosome,and various transcription factors and synergistic transcription factors can specifically bind to DNA binding sites,thus activating gene transcription.Studies have shown that the regulation of intracellular Histone acetylation level is mainly regulated through two enzymes,Histone acetyltransferases（HAT）and Histone deacetylases（HDAC）.HAT transfers acetyl-coenzyme A to the amino group of lysine（Lys）residue of histone,which relaxes the conformation of chromosome DNA and is recognized by corresponding recognition proteins,and further recruits transcription factors for gene expression.HDAC can remove acetyl groups from histone Lys residues,make chromatin structure compact,and thus inhibit the transcription and expression of tumor suppressive genes.Dysregulation of HDAC can cause a variety of diseases such as cancer,affecting gene transcription and normal cell behavior.A large number of studies have shown that HDAC is associated with tumor proliferation and apoptosis,angiogenesis,metastasis and drug resistance,and HDAC inhibitors have shown good therapeutic effects as anti-tumor drugs.Therefore,HDAC inhibitors can be used to treat patients with abnormal levels of histone acetyltransferase,which is of great research value.In our work,two series of compounds were designed and synthesized with HDAC as the target,and the structure with anti-tumor activity reported in the literature as the parent nucleus（Cap）.1.Design,synthesis and antitumor activity evaluation of novel HDAC inhibitors with4,5,6,7-tetrahydrobenzothiazole as the skeletonIn this work,a series of sixteen hydroxamic acid based HDAC inhibitors were designed and synthesized with 4,5,6,7-tetrahydrobenzothiazole as the structural core.In a patent,a series of benzothiazole and 4,5,6,7-tetrahydrobenzothiazole derivatives as anticancer agents that were able to obstruct the SUMOylation,with some candidates displaying excellent antitumor activities in vitro and vivo.Inspired by these,（S）-2,6-diamino-4,5,6,7-tetrahydrobenzothiazole was selected as the structural nucleus to design novel HDACis.Different substituents installed on the 6-amino group together with the 4,5,6,7-tetrahydrobenzothiazole ring acted as the cap and were predicted to well accommodate with the entrance of the HDACs active pocket due to their flexibility and lipophilicity of 4,5,6,7-tetrahydrobenzothiazole ring.The hydroxamic acid was selected as the ZBG in consideration of its high affinity with the Zn²⁺.While,the six carbon long linker was derived from SAHA.A total of 16compounds were synthesized.Majority of them exhibited potent inhibitory activities against HDACs and one leading compound 6h was dug out.6h was proven to be a pan-HDAC inhibitor and displayed high cytotoxicity against seven human cancer cell lines with IC₅₀values in low micromolar range.6h could arrest cell cycle in G2/M phase and induce apoptosis in A549 cells.Moreover,compound 6h exhibited remarkable anti-migration and anti-angiogenesis activities.At the same time,6h was able to elevate the expression of acetylatedα-tubulin and acetylated histone H3 in a dose-dependent manner.Docking simulation revealed that 6h fitted well into the active sites of HDAC2 and 6.Finally,compound 6h also exerted potent antitumor effects in an A549 zebrafish xenograft model.Our study demonstrated that compound6h was a promising candidate for further preclinical studies.2.Design,synthesis and antitumor activity evaluation of novel dual targeting inhibitor of Tubulin and HDACMicrotubule,a major component of the eukaryotic cytoskeleton,is a hollowtube constituted ofα-andβ-tubulins by polymerization.Microtubule inhibitors can target microtubule dynamics and interfere with a variety of cellular functions,including mitosis,cell signaling,intracellular transport and angiogenesis.There are three different small molecule binding sites for tubulins:the paclitaxel binding site,the vinblastine binding site,and the colchicine binding sites.Taxane and periwinkle alkaloids have been successfully used in the clinical treatment of cancer.Although colchicine is a potent compound,its clinical use has been limited by its toxicity to normal tissues at effective drug concentrations.However,theα,β-enone structure of colchicine as the main pharmacophore of colchicine binding site provides a general idea for the structural design of our compounds.In this study,chalcone withα,β-enone structure was used as CAP,and was connected with N-hydroxycinnamamide as the zinc ion binding group,so that the designed compound could simultaneously act on Tubulin and HDAC.In this project,one compound was designed and synthesized,and verified by a series of enzyme activity,cell activity and other cell function experiments,it was found to have excellent HDAC8 selective inhibitory activity,and the IC₅₀of U87 cell line values in single digit nanomolar range.