Targeting a cryptic allosteric site of SIRT6 with small-molecule inhibitors that inhibit the migration of pancreatic cancer cells

Qiufen Zhang; Yingyi Chen; Duan Ni; Zhimin Huang; Jiacheng Wei; Li Feng; Jun-Cheng Su; Yingqing Wei; Shaobo Ning; Xiuyan Yang; Mingzhu Zhao; Yuran Qiu; Kun Song; Zhengtian Yu; Jianrong Xu; Xinyi Li; Houwen Lin; Shaoyong Lu; Jian Zhang

doi:10.1016/j.apsb.2021.06.015

Targeting a cryptic allosteric site of SIRT6 with small-molecule inhibitors that inhibit the migration of pancreatic cancer cells

Acta Pharm Sin B. 2022 Feb;12(2):876-889. doi: 10.1016/j.apsb.2021.06.015. Epub 2021 Jul 2.

Authors

Qiufen Zhang^{1

2}, Yingyi Chen¹, Duan Ni¹, Zhimin Huang¹, Jiacheng Wei¹, Li Feng², Jun-Cheng Su¹, Yingqing Wei¹, Shaobo Ning¹, Xiuyan Yang¹, Mingzhu Zhao¹, Yuran Qiu², Kun Song², Zhengtian Yu³, Jianrong Xu⁴, Xinyi Li², Houwen Lin¹, Shaoyong Lu¹, Jian Zhang^{1

2

5}

Affiliations

¹ State Key Laboratory of Oncogenes and Related Genes, Department of Pharmacy, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200127, China.
² Medicinal Chemistry and Bioinformatics Center, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China.
³ Nutshell Therapeutics, Shanghai 201203, China.
⁴ Academy of Integrative Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China.
⁵ School of Pharmaceutical Sciences, Zhengzhou University, Zhengzhou 450001, China.

Abstract

SIRT6 belongs to the conserved NAD⁺-dependent deacetylase superfamily and mediates multiple biological and pathological processes. Targeting SIRT6 by allosteric modulators represents a novel direction for therapeutics, which can overcome the selectivity problem caused by the structural similarity of orthosteric sites among deacetylases. Here, developing a reversed allosteric strategy AlloReverse, we identified a cryptic allosteric site, Pocket Z, which was only induced by the bi-directional allosteric signal triggered upon orthosteric binding of NAD⁺. Based on Pocket Z, we discovered an SIRT6 allosteric inhibitor named JYQ-42. JYQ-42 selectively targets SIRT6 among other histone deacetylases and effectively inhibits SIRT6 deacetylation, with an IC₅₀ of 2.33 μmol/L. JYQ-42 significantly suppresses SIRT6-mediated cancer cell migration and pro-inflammatory cytokine production. JYQ-42, to our knowledge, is the most potent and selective allosteric SIRT6 inhibitor. This study provides a novel strategy for allosteric drug design and will help in the challenging development of therapeutic agents that can selectively bind SIRT6.

Keywords: ADPr, ADP-ribose; Allosteric inhibitor; BSA, bull serum albumin; CCK-8, Cell Counting Kit-8; Cell migration; Cytokine production; DMSO, dimethyl sulfoxide; FBS, fetal bovine serum; FDL, Fluor de Lys; H3K18, histone 3 lysine 18; H3K56, histone 3 lysine 56; H3K9, histone 3 lysine 9; HDAC, histone deacetylase; HPLC, high-performance liquid chromatography; IC50, half-maximum inhibitory concentration; IPTG, isopropyl-β-d-thiogalactoside; MD, molecular dynamics; Molecular dynamics simulations; NAD+, nicotinamide adenine dinucleotide; NAM, nicotinamide; PBS, phosphate buffer saline; PMA, phorbol 12-myristate 13-acetate; PMSF, phenylmethanesulfonyl fluoride; Pancreatic cancer; RMSD, root-mean-square deviation; RT-qPCR, real-time quantitative PCR; Reversed allostery; SDS-PAGE, SDS-polyacrylamide gel electrophoresis; SIRT6; SIRT6, sirtuin 6.