Comprehensive profiling of ALK fusion lung adenocarcinomas identifies mutant p53 driven DNA replication and repair deregulation in targeted therapy resistance [ChIP-Seq]

Acquired resistance to ALK tyrosine kinase inhibitors (TKIs) remains a major barrier in ALK-fusion lung adenocarcinoma (LUAD), with up to 50% of resistant tumors lacking known resistance drivers. Here, we performed integrated genomic, transcriptomic, and single-cell analyses of clinical specimens and patient-derived xenografts (PDXs) to define previously uncharacterized mechanisms of resistance. Transcriptomic profiling revealed marked intratumoral heterogeneity, with multiple distinct and independent pathways contributing to ALK inhibitor resistance. While secondary ALK mutations were detected in ~30% of resistant tumors, approximately half harbored no known actionable alterations. However, TP53 mutations, particularly DNA-binding domain missense variants, were significantly enriched in these TKI-resistant tumors and were associated with activation of DNA replication and repair programs. We find that missense mutant p53 acquired gain-of-function activity by directly binding promoters of replication and repair genes and engaging the chromatin remodeler EP400, thereby enhancing DNA damage repair and promoting survival under ALK inhibition. Therapeutically, combining lorlatinib with the proteasome inhibitor carfilzomib selectively depleted mutant p53, suppressed replication programs, and overcame resistance in TP53-mutant ALK-fusion PDX models. Together, these findings identify mutant p53–driven replication and repair as a previously unrecognized common mechanism of ALK TKI resistance and nominate a rational combination strategy for targeting a substantial fraction of TKI-refractory ALK-fusion LUAD.