Abstract Introduction: Vitamin D and folate status are critical for cell signaling and functioning and low levels have been associated with an increased risk for colorectal cancer. Their role in colorectal cancer prognosis is an active area of research: higher vitamin D status is thought to improve outcomes, yet higher folate may increase risk of recurrence. As part of an international pilot study we evaluated differences and predictors of vitamin D and one-carbon biomarkers (folate/B12). Methods: Stage I-IV colorectal cancer cases (ages 18-90) and matched non-cancer controls had been recruited between 2006 and 2010 in Seattle, WA (n=98 cases, 33 controls); Heidelberg, Germany (40/40); and Tampa, FL (18/18). Plasma vitamin D status was measured by LC/MSD. Serum folate and B12 were determined by radioassay methods. To account for skewed distributions, Wilcoxon or Kruskal-Wallis tests were used to test for differences in biomarker concentrations by case status, tumor stage, study site, age, and season. Linear regression analysis with log-transformed outcome parameters was used for multivariate analyses. Results: As expected, folate levels overall were, on average, lower in Germany than in Tampa or Seattle (13ng/ml (range: 1-52) vs. 22 (4-70) and 23 (7-69) ng/ml), plausibly reflecting an absence of folic-acid fortification and lower supplement use in the German population. In Seattle, colorectal cancer patients who used folic-acid containing supplements had on average 1.5-fold higher concentrations of folate than non-users (30ng/ml vs.20ng/ml, p<0.01). For vitamin D, we observed generally consistent levels with wide interindividual differences across all three sites (Seattle: 24.1ng/ml (4.0-80) Moffitt: 20.8 (4.6-51.6) Heidelberg: 29.6 (11.0-83.0)). In Heidelberg we observed substantial seasonal variation (p=0.002), perhaps due to greater outdoor physical activity in the summer months and lower use of sun screen. In multivariate modeling, predictors for biomarkers included season (25(OH)D), study site (25(OH)D; folate, B12); and tumor stage (B12). Conclusion: Although the sample size in this pilot was small, these findings illustrate the value of an international cohort in studying colorectal cancer prognosis in the context of a broad range of exposures to discern different aspects of risk prediction. For folate, in addition to international differences, our pilot data show that the intra-individual differences at each site are large and that supplement use is a critical determinant of folate levels in colorectal cancer patients. For vitamin D in relation to outcomes, important determinants and confounders may include seasonal variation and sun exposure (e.g., oudoor physical activity and sun screen use), which need to be well measured and accounted for in prognostic studies. Citation Information: Cancer Prev Res 2010;3(12 Suppl):A82.