RC48 Induces Senescence in HER2-Expressing Colon Cancer Cells by Activating the CDKN1A-RB-E2F1 Pathway

INTRODUCTION: RC48 is an anti-HER2 antibody‒drug conjugate that demonstrates antitumor activity in colon cancer with both high and low HER2 expression. Our study aimed to investigate the underlying mechanisms of RC48 on colon cancer cells with varying levels of HER2 expression. METHODS: The effects of RC48 on HER2 high-expressing HT29 colon cancer cells and low-expressing SW480 cells were assessed using transwell assays and flow cytometry. Proteomic analysis was carried out to explore the mechanisms by which RC48 inhibits the growth of HT29 and SW480 cells. β-Galactosidase staining and ELISAs were performed to validate the promotion of senescence in colon cancer cells by RC48. Bioinformatics analysis revealed the correlation between differentially expressed proteins and HER2 expression. RESULTS: RC48 inhibited the growth of HT29 and SW480 cells. Proteomic analysis revealed that RC48 enhanced CDKN1A expression in HT29 and SW480 cells. KEGG analysis indicated that CDKN1A expression was associated with the cellular senescence signaling pathway. RC48 upregulated CDKN1A expression in HT29 and SW480 cells, whereas the expression of other proteins related to the cellular senescence pathway was reduced. CDKN1A gene overexpression suppressed HT29 and SW480 cells. β-Gal staining demonstrated that RC48 promoted senescence in colon cancer cells, and RC48 enhanced senescence-associated secretory phenotype factor secretion in ELISAs. The close correlation between CDKN1A and HER2 expression was derived from the database. DISCUSSION: This study provides important insights into the antitumor mechanism of RC48, suggesting for the first time that CDKN1A-mediated cellular senescence may be a key mechanism underlying its therapeutic effect in HER2-high and low-expressing colon cancer, thereby offering a new theoretical basis for optimizing its clinical application. However, in vivo and clinical validation are lacking, and the functional role of CDKN1A, as well as its regulatory relationship with HER2, requires further investigation. CONCLUSION: RC48 exerts antitumor effects on both HER2-high- and HER2-low-expressing colon cancer cells. RC48 may induce senescence in both high- and low-HER2 expressing co-lon cancer cells by increasing CDKN1A protein expression. Moreover, there is a close correlation between CDKN1A and HER2 expression.