Ohoka Nobumichi, Ujikawa Osamu, Shimokawa Kenichiro, Sameshima Tomoya, Shibata Norihito, Hattori Takayuki, Nara Hiroshi, Cho Nobuo, Naito Mikihiko
Division of Molecular Target and Gene Therapy Products, National Institute of Health Sciences.
Pharmaceutical Research Division, Takeda Pharmaceutical Co., Ltd.
Chem Pharm Bull (Tokyo). 2019 Mar 1;67(3):203-209. doi: 10.1248/cpb.c18-00567. Epub 2018 Oct 26.
Targeted protein degradation by small molecules is an emerging modality with significant potential for drug discovery. We previously developed chimeric molecules, termed specific and non-genetic inhibitor of apoptosis protein (IAP)-dependent protein erasers (SNIPERs), which induce the ubiquitylation and proteasomal degradation of target proteins. This degradation is mediated by the IAPs; the target proteins include bromodomain-containing protein 4 (BRD4), an epigenetic regulator protein. The SNIPER that degrades this particular protein, SNIPER(BRD)-1, consists of an IAP antagonist LCL-161 derivative and a bromodomain and extra-terminal (BET) inhibitor, (+)-JQ-1. SNIPER(BRD)-1 also degrades a cellular inhibitor of apoptosis protein 1 (cIAP1) and an X-linked inhibitor of apoptosis protein (XIAP), the mechanisms of which are not well understood. Here, we show that the degradation of cIAP1 and XIAP by SNIPER(BRD)-1 is induced via different mechanisms. Using a chemical biology-based approach, we developed two inactive SNIPERs, SNIPER(BRD)-3 and SNIPER(BRD)-4, incapable of degrading BRD4. SNIPER(BRD)-3 contained an N-methylated LCL-161 derivative as the IAP ligand, which prevented it from binding IAPs, and resulted in the abrogated degradation of cIAP1, XIAP, and BRD4. SNIPER(BRD)-4, however, incorporated the enantiomer (-)-JQ-1 which was incapable of binding BRD4; this SNIPER degraded cIAP1 but lost the ability to degrade XIAP and BRD4. Furthermore, a mixture of the ligands, (+)-JQ-1 and LCL-161, induced the degradation of cIAP1, but not XIAP and BRD4. These results indicate that cIAP1 degradation is triggered by the binding of the IAP antagonist module to induce autoubiquitylation of cIAP1, whereas a ternary complex formation is required for the SNIPER-induced degradation of XIAP and BRD4.
小分子靶向蛋白质降解是一种新兴的药物研发模式，具有巨大的潜力。我们之前开发了嵌合分子，称为凋亡蛋白（IAP）依赖性蛋白清除剂（SNIPERs）的特异性非遗传抑制剂，它能诱导靶蛋白的泛素化和蛋白酶体降解。这种降解由IAP介导；靶蛋白包括含溴结构域蛋白4（BRD4），一种表观遗传调节蛋白。降解这种特定蛋白的SNIPER，即SNIPER(BRD)-1，由IAP拮抗剂LCL-161衍生物和含溴结构域及额外末端（BET）抑制剂(+)-JQ-1组成。SNIPER(BRD)-1还能降解细胞凋亡蛋白1（cIAP1）和X连锁凋亡抑制蛋白（XIAP），但其机制尚不清楚。在此，我们表明SNIPER(BRD)-1对cIAP1和XIAP的降解是通过不同机制诱导的。使用基于化学生物学的方法，我们开发了两种无活性的SNIPER，即SNIPER(BRD)-3和SNIPER(BRD)-4，它们无法降解BRD4。SNIPER(BRD)-3含有N-甲基化的LCL-161衍生物作为IAP配体，这阻止了它与IAP结合，导致cIAP1、XIAP和BRD4的降解被消除。然而，SNIPER(BRD)-4包含对映体(-)-JQ-1，它无法结合BRD4；这种SNIPER能降解cIAP1，但失去了降解XIAP和BRD4的能力。此外，配体(+)-JQ-1和LCL-161的混合物能诱导cIAP1的降解，但不能诱导XIAP和BRD4的降解。这些结果表明，cIAP1的降解是由IAP拮抗剂模块的结合触发，以诱导cIAP1的自泛素化，而SNIPER诱导的XIAP和BRD4降解需要形成三元复合物。
