错义突变型p53调控DNA复制与损伤反应，促进ALK融合肺腺癌对靶向治疗的耐药

ALK gene fusions drive oncogenesis in ~5% of lung adenocarcinomas (LUADs), and ALK tyrosine kinase inhibitors (TKIs) such as lorlatinib have improved outcomes. However, acquired resistance remains a challenge, with >30% of mechanisms unknown. To uncover novel drivers of resistance, we performed multi-omic profiling of clinical samples and PDXs. Genomic analysis of 83 ALK+ LUADs revealed frequent co-occurring alterations, including CDKN2A deletion (34%), TP53 mutation (32%), and MYC amplification (13%). Transcriptomic profiling highlighted enrichment of DNA replication and repair pathways, MYC targets, epithelial-to-mesenchymal transition, and TGF-beta signaling in resistant tumors. scRNAseq of matched sensitive and resistant PDXs revealed extensive heterogeneity and diverse resistance pathways, mirroring those in clinical samples. Notably, elevated DNA replication and repair activity strongly correlated with TP53 missense mutations in clinical samples. Analysis of MSK clinical cohort showed that 80% of TP53 mutations in ALK+ LUAD occur in the DNA-binding domain, predominantly missense variants with potential gain-of-function (GOF) properties. The presence of TP53 missense mutations predicted worse prognosis in ALK-fusion LUAD patients treated with TKIs.To assess the role of mutp53 in resistance, we overexpressed R175H and R273H in ALK+ H3122 and H2228 cells. Co-immunoprecipitation and mass spectrometry revealed that mutp53 associates with DNA repair, cell cycle, and chromatin remodeling proteins. Mutp53-expressing cells were refractory to lorlatinib compared to isogenic p53-deficient clones. Overexpression of mutp53 upregulated DNA replication proteins, including ORC and MCM family members, as well as DNA repair proteins such as BRCA1, ATM, CHEK1, and MSH6. Notably, mutp53-expressing cells maintained DNA replication and repair protein levels following lorlatinib-induced DNA damage, unlike p53-deficient counterparts. Immunofluorescence of the DNA damage marker pH2A demonstrated complete repair of lorlatinib-induced lesions in the presence of mutp53, revealing a novel GOF role for these mutants. Furthermore, mutp53 enhanced DNA synthesis by promoting origin firing, thereby facilitating cellular survival under targeted therapy. Similar effects were observed in EGFR-driven LUAD cells, indicating that mutp53 contributes to resistance across oncogene-driven LUADs.To target this axis, we tested the proteasome inhibitor carfilzomib which inhibited mutp53 proteins and promoted the degradation of DNA replication proteins. Carfilzomib synergized with lorlatinib in resistant ALK+ p53 mutant cells and induced robust tumor response in TP53 mutant ALK+ PDX models resistant to lorlatinib, demonstrating therapeutic potential. In summary, missense mutp53 promotes ALK TKI resistance by enhancing DNA replication and repair.