BACKGROUND: Colorectal cancer (CRC) is a common cause of cancer-related mortality and is mainly influenced by metabolic dysfunction. The β-alanine metabolic pathway plays an important role in altering the aggressiveness and metabolic characteristics of the cancer cells. This study aimed to investigate the genetic role of the β-alanine metabolic pathway in CRC patient survival. METHODS: Using a Cox regression model, we assessed the impact of 27 single-nucleotide polymorphisms (SNPs) from 31 genes in the β-alanine metabolic pathway on overall survival (OS) and progression-free survival (PFS) in 287 patients with CRC. Additional methods, including differential expression analysis, expression quantitative trait loci analysis, dual-luciferase reporter assay, and cell phenotype assay, were used to evaluate the genetic function of candidate SNPs in tumor progression. RESULTS: We identified that SNP rs2811182 A > G allele located in DPYD was significantly associated with poorer prognosis of CRC, with hazard ratio (HR) of 0.63 for OS [95% confidence interval (CI) = 0.45-0.88, P = 7.12 × 10] and 0.68 for PFS (95% CI = 0.52-0.89, P = 5.01 × 10). Mechanistically, the G allele of rs2811182 increased DPYD transcriptional activity and expression by mediating the binding affinity of the transcription factor POU1F1. Importantly, the overexpression of DPYD reduced the malignant cell phenotypes of proliferation, migration, and invasion. CONCLUSIONS: This study indicates a pivotal genetic role for the β-alanine metabolic pathway, particularly rs2811182 in DPYD, in influencing CRC prognosis. These findings offer new perspectives for personalized treatment strategies and enhance our understanding of CRC pathogenesis.