Department of Pathology, The Johns Hopkins University, Baltimore, Maryland, United States of America.
Department of Oncology, The Johns Hopkins University, Baltimore, Maryland, United States of America.
PLoS One. 2021 Sep 10;16(9):e0256937. doi: 10.1371/journal.pone.0256937. eCollection 2021.
Bortezomib and the other licensed 20S proteasome inhibitors show robust activity against liquid tumors like multiple myeloma, but have disappointed against solid tumors including ovarian cancer. Consequently, interest is mounting in alternative non-peptide based drugs targeting the proteasome's 19S regulatory particle subunit, including its ubiquitin receptor RPN13. RA183 and RA375 are more potent analogs of the prototypic inhibitor of RPN13 (iRPN13) called RA190, and they show promise for the treatment of ovarian cancer. Here we demonstrate that rendering these candidate RPN13 inhibitors chiral and asymmetric through the addition of a single methyl to the core piperidone moiety increases their potency against cancer cell lines, with the S-isomer being more active than the R-isomer. The enhanced cancer cell cytotoxicities of these compounds are associated with improved binding to RPN13 in cell lysates, ATP depletion by inhibition of glycolysis and mitochondrial electron chain transport, mitochondrial depolarization and perinuclear clustering, oxidative stress and glutathione depletion, and rapid accumulation of high molecular weight polyubiquitinated proteins with a consequent unresolved ubiquitin proteasome system (UPS) stress response. Cytotoxicity was associated with an early biomarker of apoptosis, increased surface annexin V binding. As for cisplatin, BRCA2 and ATM deficiency conferred increased sensitivity to these iRPN13s. Ubiquitination plays an important role in coordinating DNA damage repair and the iRPN13s may compromise this process by depletion of monomeric ubiquitin following its sequestration in high molecular weight polyubiquitinated protein aggregates. Indeed, a synergistic cytotoxic response was evident upon treatment of several ovarian cancer cell lines with either cisplatin or doxorubicin and our new candidate iRPN13s, suggesting that such a combination approach warrants further exploration for the treatment of ovarian cancer.
硼替佐米和其他已获许可的 20S 蛋白酶体抑制剂对多发性骨髓瘤等液体肿瘤具有强大的活性，但对包括卵巢癌在内的实体瘤作用不大。因此，人们对靶向蛋白酶体 19S 调节颗粒亚基的替代非肽类药物越来越感兴趣，包括其泛素受体 RPN13。RA183 和 RA375 是 RPN13 的原型抑制剂（iRPN13）RA190 的更有效类似物，它们有望用于治疗卵巢癌。在这里，我们证明通过在核心哌啶酮部分添加一个甲基，使这些候选 RPN13 抑制剂具有手性和不对称性，可以提高它们对癌细胞系的活性，其中 S-异构体比 R-异构体更有效。这些化合物增强癌细胞的细胞毒性与它们在细胞裂解物中与 RPN13 的结合改善、通过抑制糖酵解和线粒体电子链传递导致 ATP 耗竭、线粒体去极化和核周聚集、氧化应激和谷胱甘肽耗竭以及高分子量多聚泛素化蛋白的快速积累有关，从而导致未解决的泛素蛋白酶体系统 (UPS) 应激反应。细胞毒性与凋亡的早期生物标志物，即表面膜联蛋白 V 结合增加有关。与顺铂一样，BRCA2 和 ATM 缺陷赋予对这些 iRPN13 更高的敏感性。泛素化在协调 DNA 损伤修复中起着重要作用，而 iRPN13 可能通过将其隔离在高分子量多聚泛素化蛋白聚集体中耗尽单体泛素来破坏这一过程。事实上，在用顺铂或多柔比星和我们的新候选 iRPN13 治疗几种卵巢癌细胞系时，观察到协同的细胞毒性反应，这表明这种联合治疗方法值得进一步探索用于治疗卵巢癌。
