Background: Despite several plausible biological mechanisms linking proton pump inhibitors (PPIs) and H2 receptor antagonists (H2RAs) with colorectal tumorigenesis, their association with risk of colorectal cancer (CRC) has not been adequately assessed in prospective epidemiological studies. Methods: We evaluated the association of acid-suppressive medication use with CRC risk among 175,871 (PPI) and 208,831 (H2RA) participants from three large prospective cohort studies. Medication use was assessed at baseline and updated biennially. The association was evaluated using multivariate Cox proportional hazards regression models. Results: There was no significant association between baseline PPI use (hazard ratio (HR) = 0.89, 95% confidence interval (CI), 0.71–1.12) or PPI use after a lag of 8–10 years (HR = 1.12, 95% CI, 0.78–1.59) with CRC risk. We observed no significant association between H2RA use after a lag of 8–10 years and CRC risk (HR = 1.02, 95% CI, 0.81–1.28), while risk was lower for participants with baseline H2RA use (HR = 0.76, 95% CI, 0.60–0.95). Duration of PPI use or H2RA use was not associated with CRC risk (P-trend = 0.21 and 0.95, respectively). Conclusions: Among participants from three large prospective cohorts, use of PPI or H2RA was not associated with higher risk of colorectal cancer.