PO.ET03.04 · 实验与分子治疗
VLX1570抑制蛋白酶体相关DUB以靶向UCHL5/USP14并增强顺铂在神经母细胞瘤中的疗效
Proteasome-associated DUB inhibition by VLX1570 targets UCHL5/USP14 and enhances cisplatin efficacy in neuroblastoma
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摘要 Abstract
中文摘要
神经母细胞瘤是儿童中最常见的颅外实体瘤，尽管采用强化的多模式治疗，高危疾病仍预后不良。与19S蛋白酶体相关的去泛素化酶（DUB），如泛素C端水解酶L5（UCHL5）和泛素特异性蛋白酶14（USP14），是蛋白质稳态的关键调控因子，并作为潜在治疗靶点引起关注。在本研究中，我们通过免疫组织化学检查了临床神经母细胞瘤标本中UCHL5和USP14的表达（与正常外周神经组织相比），并使用小分子抑制剂VLX1570在一组人神经母细胞瘤细胞系中研究了药理学抑制蛋白酶体相关DUB活性的效应，通过评估细胞活力、细胞周期分布、凋亡、克隆形成生长以及蛋白酶体应激、内质网应激和自噬的标志物，同时在异种移植小鼠模型中进行研究。顺铂仍是高危神经母细胞瘤诱导和巩固化疗的基石，而对基于顺铂方案的耐药或不耐受是治愈的主要障碍；因此，我们特别评估了VLX1570在体外和体内是否能增强顺铂诱导的细胞毒性。相对于正常外周神经，UCHL5和USP14在神经母细胞瘤组织中显著上调，VLX1570处理显著降低了神经母细胞瘤细胞生长，诱导G2/M期阻滞和caspase依赖性凋亡，促进多聚泛素化蛋白积累，激活未折叠蛋白反应信号，并调节自噬相关蛋白。值得注意的是，在体外，VLX1570与顺铂联合比任一单独治疗更强地抑制细胞活力和克隆存活，并增强了凋亡信号。一致地，VLX1570在异种移植模型中显著抑制肿瘤生长，而与顺铂共处理进一步延缓了肿瘤进展并降低了增殖指数，且无明显的全身毒性。综上所述，这些数据表明UCHL5和USP14在神经母细胞瘤中过表达，且用VLX1570药理学抑制蛋白酶体相关DUB活性发挥抗肿瘤效应并增强顺铂疗效，支持在神经母细胞瘤中进一步评估DUB抑制剂-顺铂联合策略。
查看英文原文 English abstract
Neuroblastoma is the most common extracranial solid tumor in children, and high-risk disease still carries an unfavorable prognosis despite intensive multimodality therapy. Deubiquitinating enzymes (DUBs) associated with the 19S proteasome, such as ubiquitin C-terminal hydrolase L5 (UCHL5) and ubiquitin-specific protease 14 (USP14), are key regulators of protein homeostasis and have attracted interest as potential therapeutic targets. In this study, we examined UCHL5 and USP14 expression in clinical neuroblastoma specimens compared with normal peripheral nerve tissue by immunohistochemistry, and investigated the effects of pharmacologic inhibition of proteasome-associated DUB activity using the small-molecule inhibitor VLX1570 in a panel of human neuroblastoma cell lines by assessing cell viability, cell-cycle distribution, apoptosis, clonogenic growth, and markers of proteasome stress, endoplasmic reticulum stress, and autophagy, as well as in a xenograft mouse model. Cisplatin remains a cornerstone of induction and consolidation chemotherapy for high-risk neuroblastoma, and resistance or intolerance to cisplatin-based regimens represents a major barrier to cure; therefore, we specifically evaluated whether VLX1570 could potentiate cisplatin-induced cytotoxicity in vitro and in vivo. UCHL5 and USP14 were markedly upregulated in neuroblastoma tissues relative to normal peripheral nerve, and VLX1570 treatment significantly reduced neuroblastoma cell growth, induced G2/M arrest and caspase-dependent apoptosis, promoted accumulation of polyubiquitinated proteins, activated unfolded protein response signaling, and modulated autophagy-related proteins. Notably, combining VLX1570 with cisplatin produced greater suppression of cell viability and clonogenic survival, and enhanced apoptotic signaling, than either treatment alone in vitro. Consistently, VLX1570 significantly suppressed tumor growth in xenograft models, while co-treatment with cisplatin further delayed tumor progression and reduced proliferative indices without overt systemic toxicity. Taken together, these data demonstrate that UCHL5 and USP14 are overexpressed in neuroblastoma and that pharmacologic inhibition of proteasome-associated DUB activity with VLX1570 exerts antitumor effects and enhances the efficacy of cisplatin, supporting further evaluation of DUB inhibitor-cisplatin combination strategies in neuroblastoma.
利益披露 Disclosure
Y. Kao, None..
C. Hsu, None..
S. Liao, None..
P. Chow, None..
K. Huang, None.