Multimodal interrogation establishes the AAA+ ATPase RUVBL1 as a prognostic driver and tractable dependency in malignant pleural mesothelioma

<title>Abstract</title> <p>Malignant pleural mesothelioma (MPM) is an inexorably lethal malignancy, yet its therapeutic landscape remains largely devoid of actionable drivers. Here, by leveraging a stringent prioritization framework on genome-scale CRISPR dependency maps integrated with orthogonal molecular interrogation, we identify the AAA+ ATPase RUVBL1, an obligate component of the RUVBL1/2 complex, as a non-redundant and pharmacologically tractable vulnerability in MPM. This liability reflects a network-level addiction to RUVBL1/2-containing multiprotein assemblies, operating as survival hubs where subunit integrity appears indispensable. Genetic depletion of RUVBL1 induces profound cytostasis and cytotoxicity, compromising tumor cell fitness and clonogenic expansion in adherent monolayers and multicellular spheroids. Mechanistically, RUVBL1 loss triggers canonical replication stress with destabilization of the PIKK family kinases ATM and ATR. This leads to γH2AX accumulation, induction of p21, and activation of apoptotic signaling, enforcing checkpoint engagement and subsequent cell death. In contrast, ectopic RUVBL1 expression amplifies proliferative kinetics and potentiates aggressive traits, including epithelial-to-mesenchymal transition (EMT), invasion, and anchorage-independent growth. Correlative transcriptomics consensually links RUVBL1 to core oncogenic programs governing DNA damage repair, cell fitness, and EMT plasticity. Remarkably, pharmacologic inhibition of the RUVBL1/2 complex phenocopies the hallmarks of genetic perturbation. Clinically, tissue microarray profiling demonstrates significantly elevated RUVBL1 in mesothelioma, while multi-cohort analyses position it as a disease-specific marker and negative prognostic indicator. Critically, patient-derived, treatment-naïve ex vivo MPM cultures retain tumor-intrinsic RUVBL1 expression and exhibit strong sensitivity to ATPase inhibition, providing preclinical validation in patient-proximal models. Together, our findings establish RUVBL1 as a bona fide driver of MPM pathobiology and nominate the RUVBL1/2 complex as a rational target for therapeutic exploitation.</p>