p53 and p21 Status Influences Cellular Response to Metformin in KRAS-Mutant HCT116 Colorectal Cancer Cells

Colorectal cancer (CRC) remains a leading cause of cancer morbidity worldwide, highlighting the need for improved therapies. Metformin, a widely used antihyperglycemic agent, has gained attention for its potential antitumor properties. In this study, we evaluated the effects of the tumor suppressor genes TP53 and CDKN1A on metformin responsiveness in a KRAS-mutant CRC in vitro model using HCT116 cells harboring a G13D mutation in KRAS. Using parental (p53^+/+, p21^+/+) and isogenic knockout cell lines, we assessed cell-cycle distribution and transcriptomic responses following metformin treatment. In parental wild-type cells, metformin exposure was associated with a dose- and time-dependent reduction in cell viability and an increased proportion of cells in the G0/G1 phase, with significance levels across treatment conditions ranging from p = 0.03 to p < 0.0001. Loss of p53 or p21 was associated with attenuated cellular responses to metformin, with p21-deficient cells responding primarily at higher doses and prolonged exposure (72 h). Transcriptomic profiling revealed extensive differential gene expression in parental cells (1399 DEGs), compared with more limited responses in p53^-/- (270 DEGs) and p21^-/- cells (32 DEGs). Differentially expressed genes associated with MAPK signaling (DUSP5) and inflammatory regulation (TNFAIP3) were observed across genotypes, whereas pathway enrichment of DNA replication and chromatin organization was specific to p53-deficient cells. These findings provide a transcriptomic and phenotypic characterization of genotype-dependent cellular responses to metformin and establish a basis for future mechanistic and functional validation studies.