Structural Modification Of1,3,4-thiadiazole Based HDAC Inhibitors

Histone acetylation and deacetylation are highly studied epigenetic phenomena, which balance is regulated by histone acetyltransferase (HAT) and histone deacetylase (HDAC). HDAC catalyze the removal of acetyl moieties from N-acetyllysine residues of histone. Human HDAC are subdivided into two classes based their mechanisms of action:zinc dependent and NAD+dependent HDAC. The zinc dependent HDAC can be tethered closely to tumor onset and progression considering aberrant expression of them in broad spectrum neoplasms. Small molecule HDAC inhibitors can induce tumor cell apoptosis, differentiation, growth arrest and inhibit angiogenesis. To date vorinostat (suberoylanilide hydroxamic acid, SAHA) and romidepsin have been FDA approved for use against cutaneous T cell lymphoma (CTCL). Many other HDAC inhibitors have demonstrated potent antitumor effects in clinical trials for treatment of hematological malignancies. Therefore, HDAC inhibitors emerge as a promising therapy for cancer.According to the enzyme-inhibitor interaction in the crystal structure of HDAC2complexed with SAHA, the structural characteristics of HDAC inhibitors are composed of a zinc binding group (ZBG), a linker and a cap attached to the linker. Previously we reported the discovery of1,3,4-thiadiazole containing hydroxamates as submicromolar HDAC inhibitors, of which compound6m showed inhibitory potency against HDAC enzymes similar to SAHA.6m exhibited as great antiproliferative activity as SAHA against ten cancer cell lines.6m can induce cell cycle arrest and apoptosis and inhibit migration of cancer cells. In order to improve the antiproliferative activity, modifications of1,3,4-thiadiazole based HDAC inibitors were investigated as follows:Series A came from introduction of various substituents into the C-5-phenyl of1,3,4-thiadiazole; Series B came from replacement of the C-5-phenyl of1,3,4-thiadiazole with other aromatic rings, such as naphthyl, pyridyl, furyl, and thienyl; Series C came from replacement of hydroxamate with other ZBGs, such as2-aminophenylamide; Series D came from bioisosterism of1,3,4-thiadiazole with thiazole and1H-pyrazole.Synthesis of Series A and B:The5-substituted-1,3,4-thiadiazol-2-amines were prepared by a facile one-pot reaction of the corresponding carboxylic acids, N-aminothiourea, and phosphoryl chloride. Treatment of the monomethyl esters with refluxing thionyl chloride and then the5-substituted-1,3,4-thiadiazol-2-amines gave1,3,4-thiadiazol-containing esters, which were converted to hydroxamic acids; Synthesis of Series C:Saponification of1,3,4-thiadiazol- containing ester above yielded the corresponding carboxylic acid. This was followed by amide coupling and protective group removal to afford compoud Cl; hydroxamic acids C2-C4was generated from different1,3,4-thiadiazol-containing esters; Synthesis of Series D:Thiazole and1H-pyrazole were prepared according to reported procedures and converted to hydroxamic acids as same as Series A and B.143novel chemical entities were synthesizd including71target compounds and72key intermediates. Structures of target compounds were confirmed with1H NMR,13C NMR, or HRMS.The bioactivity of target compounds was evaluated in enzymatic and/or cellular assays. Most compounds in Series A and B possessed better or similar HDAC inhibitory activity compared to SAHA. Structure-activity relationship (SAR) can be observed:1) addition of a bulky group on the phenyl led to a significant loss in HDAC inhibitory activity;2) compounds with an electron-withdrawing group exhibited generally poorer HDAC inhibition than those with an electron-donating group;3) a tendency in enzymatic potency of para-substitution<meta-substitution<ortho-substitution was noted. Replacement of the phenyl with a naphthalenyl group resulted in a considerable loss in HDAC inhibition, while pyridyl, furyl, and thienyl analogues had better or similar activity compared to SAHA. As for antiproliferative activity, the most active compound A31in Series A exerted higher activity toward all the six cell lines than6m and SAHA. The most active compound B12in Series B was comparable to6m and SAHA. The6carbon unit linker afforded appreciably potent cell growth inhibition.All the compounds in Series C were less active antiproliferative agents than6m and SAHA. N-hydroxybenzamide and N-hydroxycinnamamide analogues had poor inhibition on tumor cell growth.2-Aminophenylamide derivatives showed antiproliferative activity in the same order of magnitude as6m and SAHA.Compounds D1, D2and D3in Series D from replacement of1,3,4-thiadiazole with thiazole and1H-pyrazole exhibited profound growth-inhibitory activity towards all the ten cancer cell lines, which were remarkably better than that of6m and SAHA. Compounds D1, D2and D3had not only better antiproliferative activity on K562chronic myelogenous leukaemia cell, but also2-6fold greater potency on solid tumor cells than6m and SAHA.In conclusion, modifications of1,3,4-thiadiazole based HDAC inibitors were systematically investigated and SAR indicated that:introduction of small substituents into the ortho-position of the C-5-phenyl of1,3,4-thiadiazole improved both enzymatic and antiproliferative activity; replacement of the C-5-phenyl of1,3,4-thiadiazole with other aromatic rings or replacement of hydroxamate with other ZBGs failed to generate better antiproliferative agents; Series D from replacement of1,3,4-thiadiazole with thiazole and1H-pyrazole surprisingly exhibited much greater growth-inhibitory activity than6m and SAHA, representing highly promising antitumor agents with the potential for the treatment of human cancers.