CRISPRi screening identifies epigenetic vulnerabilities and an ARID1A-PRC2 synthetic-lethal axis sensitizing cutaneous T-cell lymphoma to combined JAK/STAT and EZH2 inhibition

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.