Unique Molecular Interaction with the Histone Deacetylase 6 Catalytic Tunnel: Crystallographic and Biological Characterization of a Model Chemotype

Histone deacetylase 6 (HDAC6) is involved in multiple regulatory processes, ranging from cellular stress to intracellular transport. Inhibition of aberrant HDAC6 activity in several cancers and neurological diseases has been shown to be efficacious in both preclinical and clinical studies. While selective HDAC6 targeting has been pursued as an alternative to pan-HDAC drugs, identifying truly selective molecular templates has not been trivial. Herein, we report a structure–activity relationship study yielding TO-317, which potently binds HDAC6 catalytic domain 2 (Ki = 0.7 nM) and inhibits the enzyme function (IC50 = 2 nM). TO-317 exhibits 158-fold selectivity for HDAC6 over other HDAC isozymes by binding the catalytic Zn2+ and, uniquely, making a never seen before direct hydrogen bond with the Zn2+ coordinating residue, His614. This novel structural motif targeting the second-sphere His614 interaction, observed in a 1.84 Å resolution crystal structure with drHDAC6 from zebrafish, can provide new pharmacophores for identifying enthalpically driven, high-affinity, HDAC6-selective inhibitors.

组蛋白去乙酰化酶6（Histone deacetylase 6，HDAC6）参与诸多调控过程，涵盖细胞应激、胞内运输等多种生物学环节。多项临床前与临床研究均证实，在多种癌症及神经系统疾病中，抑制异常激活的HDAC6活性可展现出治疗有效性。尽管靶向选择性HDAC6已被作为泛HDAC抑制剂的替代方案加以探索，但筛选真正具备高选择性的分子模板并非易事。本研究通过构效关系分析，获得化合物TO-317：其可强效结合HDAC6催化结构域2（抑制常数Ki=0.7 nM），并抑制该酶的催化活性（半抑制浓度IC50=2 nM）。TO-317对HDAC6的选择性是其他HDAC同工酶的158倍，其通过结合催化位点的Zn²+，且尤为独特的是，可与Zn²+的配位残基His614形成此前从未被报道的直接氢键相互作用。本研究在斑马鱼drHDAC6的1.84埃分辨率晶体结构中，观察到这种靶向第二配位球His614相互作用的新型结构基序，可为筛选焓驱动、高亲和力的HDAC6选择性抑制剂提供全新的药效团设计模板。