RB1-I680T変異は、E2F1転写の抑制解除を介して非小細胞肺癌における腫瘍増殖と化学療法の感受性を増強する

Retinoblastoma Transcriptional Corepressor 1 (RB1) is a critical tumor suppressor restricting the malignant progression of cancer cells. Emerging evidence indicates that RB1 mutations typically promote tumorigenesis through loss of its tumor-suppressive functions. Yet, the biological significance of mutated RB1, specifically certain rare variants, in non-small cell lung cancer (NSCLC) remains elusive. Here, we first reported a rare and previously uncharacterized missense mutation in RB1, the 680th residue isoleucine replaced by threonine (RB1-I680T), in NSCLC.

To investigate the functional and mechanistic consequences of the RB1-I680T mutation in NSCLC, we first used CRISPR-Cas9 to knock out endogenous RB1 in NSCLC cells. Then, we generated cell models harboring the RB1-I680T mutation by infecting these knockout cells with lentivirus carrying either wild-type RB1 (RB1-WT) or RB1-I680T expression constructs. The biological phenotypes mediated by RB1-I680T were investigated using in vitro and in vivo experiments. The exploration of the molecular mechanism was performed primarily through co-immunoprecipitation, immunofluorescence, dual-luciferase reporter assays, western blot analysis, and protein docking and dynamics simulation.

Our study demonstrated that the I680T mutation caused faster tumor growth and potentiated chemotherapy-induced tumor regression compared to RB1-WT control. Mechanistic studies illustrated that the I680T mutation in RB1 disrupted its inhibition of E2F1 transcriptional activity by weakening the physical interaction between RB1 and E2F1 in a manner dependent on conformational flexibility of RB1 pocket B domain, which is essential for sustaining the enhanced proliferation and chemosensitivity in NSCLC cells.

Our findings elucidate that the I680T mutation-induced loss-of-function of RB1 simultaneously confers invasive proliferation and chemotherapeutic vulnerability to tumor cells, suggesting that RB1-I680T could serve as a predictive biomarker for chemotherapy response in NSCLC. Stratifying patients based on the RB1-I680T mutation status may enable personalized therapeutic strategies, particularly for tumors with E2F1 dysregulation.