PO.ET09.02 · 实验与分子治疗
CRISPRi筛选鉴定表观遗传脆弱性和ARID1A-PRC2合成致死轴使皮肤T细胞淋巴瘤对JAK/STAT和EZH2联合抑制敏感
CRISPRi screening identifies epigenetic vulnerabilities and an ARID1A-PRC2 synthetic-lethal axis sensitizing cutaneous T-cell lymphoma to combined JAK/STAT and EZH2 inhibition
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
晚期皮肤T细胞淋巴瘤(CTCL)危及生命且治疗选择有限。异常的JAK/STAT激活是其决定性的分子特征,一项II期试验证明JAK1/2抑制剂ruxolitinib在T细胞淋巴瘤中有效。然而,反应有限且短暂,凸显了对基于机制的联合策略的需求。多梳抑制复合物2(PRC2)由其催化亚基EZH2驱动,调控H3K27me3介导的转录抑制,并在T细胞淋巴瘤中作为关键致癌驱动因子发挥作用。ARID1A是SWI/SNF的核心亚基,在生理上拮抗PRC2介导的染色质沉默。ARID1A的缺失或减少破坏了这种拮抗作用,增加了细胞对PRC2的依赖,并使细胞对EZH2抑制敏感。为鉴定ruxolitinib反应的遗传修饰因子,我们在暴露于JAK/STAT阻断的CTCL细胞系(HH和Hut78)中进行了全基因组CRISPR干扰筛选。Ruxolitinib处理富集了一个与PRC2功能直接相关的连贯表观遗传网络,提示表观遗传修饰因子可能与ruxolitinib协同作用。为评估临床相关性,我们分析了已发表的晚期CTCL皮肤活检(n = 70)和正常皮肤(n = 29)的转录组数据。EZH2表达在CTCL中显著升高(P = 0.0014),提示晚期疾病中PRC2依赖性增强。然后,我们使用药物浓度矩阵在原代CTCL PDX来源的肿瘤细胞中评估JAK和EZH2的联合抑制,产生15种配对组合。该联合在三个PDX样本中产生了强烈的、剂量依赖的生长抑制和凋亡诱导协同作用,Loewe协同评分分别为17.54、27.27和20.597。从机制上讲,CRISPRi分析揭示了在ruxolitinib选择压力下ARID1A sgRNA的耗竭(Hut78:−27.5%,P = 0.0071;HH:−14%,P = 0.069),表明JAK/STAT阻断增加了CTCL对EZH2生存的依赖。这种ARID1A-PRC2合成致死相互作用为观察到的药物协同作用提供了生物学基础。总之,这些发现表明JAK/STAT抑制将CTCL细胞驱动至PRC2依赖、EZH2高表达的表观遗传状态,产生了一个治疗上可行的脆弱性。这些数据为联合JAK和EZH2抑制提供了强有力的理论依据,正在进行的CTCL PDX体内研究将进一步明确该策略的转化潜力。
查看英文原文 English abstract
Advanced-stage cutaneous T-cell lymphoma (CTCL) is life-threatening and has limited treatment options. Aberrant JAK/STAT activation is a defining molecular feature, and a phase II trial demonstrated that ruxolitinib, a JAK1/2 inhibitor, is efficacious in T-cell lymphomas. However, responses are modest and short-lived, highlighting the need for mechanism-based combination strategies.Polycomb repressive complex 2 (PRC2), driven by its catalytic subunit EZH2, regulates H3K27me3-mediated transcriptional repression and functions as a key oncogenic driver in T-cell lymphomas. ARID1A, a core SWI/SNF subunit, physiologically counteracts PRC2-mediated chromatin silencing. Loss or reduction of ARID1A disrupts this antagonism, increasing cellular reliance on PRC2 and sensitizing cells to EZH2 inhibition.To identify genetic modifiers of ruxolitinib response, we performed a genome-wide CRISPR interference screen in CTCL cell lines (HH and Hut78) exposed to JAK/STAT blockade. Ruxolitinib treatment enriched a coherent epigenetic network directly linked to PRC2 function, suggesting that epigenetic modifiers may act synergistically with ruxolitinib.To assess clinical relevance, we analyzed published transcriptomic data from advanced CTCL skin biopsies (n = 70) and normal skin (n = 29). EZH2 expression was significantly elevated in CTCL (P = 0.0014), suggesting heightened PRC2 dependence in advanced-stage disease. We then evaluated the combined inhibition of JAK and EZH2 in primary CTCL PDX-derived tumor cells using a matrix of drug concentrations, yielding 15 paired combinations. This combination produced strong, dose-dependent synergy in growth inhibition and apoptosis induction across three PDX samples, with Loewe synergy scores of 17.54, 27.27, and 20.597, respectively.Mechanistically, CRISPRi profiling revealed depletion of ARID1A sgRNAs under ruxolitinib selection pressure (Hut78: −27.5%, P = 0.0071; HH: −14%, P = 0.069), indicating that JAK/STAT blockade increases CTCL dependence on EZH2 for survival. This ARID1A-PRC2 synthetic-lethal interaction provides a biological basis for the observed drug synergy.Together, these findings demonstrate that JAK/STAT inhibition drives CTCL cells into a PRC2-dependent, EZH2-high epigenetic state, creating a therapeutically actionable vulnerability. These data provide a strong rationale for combining JAK and EZH2 inhibition, and ongoing CTCL PDX in vivo studies will further define the translational potential of this strategy.
利益披露 Disclosure
Y. Liu, None..
L. Pincus, None..
Y. Chang, None.