Comparative analysis of phenotypic markers of DPD activity as biomarkers of 5-FU toxicity

Nearly 300,000 cancer patients are treated with fluoropyrimidine chemotherapy in the U.S. annually. Approximately one-third of patients experience severe and life-threatening toxicities, which can result in death. Deleterious genetic variants in DPYD , which encodes the rate-limiting enzyme of fluoropyrimidine catabolism (dihydropyrimidine dehydrogenase, DPD), have been correlated with severe toxicity in clinical studies. However, these variants explain only 10-30% of toxicity cases. Phenotypic measures of DPD function are attractive biomarkers of fluoropyrimidine toxicity risk with the potential to identify unknown causes of DPD deficiency. In this study, we compared two measures of DPD enzyme activity, blood plasma dihydrouracil to uracil ratio (UH 2 :U) and direct measurement of DPD activity in peripheral blood mononuclear cells (PBMCs) using 204 subjects with comprehensive DPYD genotype data. DPD PBMC measurements showed higher overall inter-individual variation compared to UH 2 :U ratios. Low correlation between UH 2 :U and PBMC measurements were noted for matched samples taken from the same subject. As expected, DPD metabolites (U and UH 2 ) correlated with UH 2 :U ratio; however, no correlation was noted between U or UH 2 levels and the PBMC DPD activity. Furthermore, when data were analyzed by genotype, no correlations were noted between UH 2 :U and PBMC DPD activity. We observed the expected decrease in UH 2 :U ratio for carriers of the four clinically tested DPYD variants and identified two novel variants associated with a significantly decreased UH 2 :U ratio. To further investigate the impact of rare genotypes and previously identified DPYD haplotypes on the UH 2 :U ratio, additional cohorts of 1994 and 500 subjects, enriched for specific genotypes, were utilized. Our results suggest that phenotyping may have limited validity as a single test when predicting DPD deficiency at an individual level and call into question the often-reported belief that PBMC DPD activity represents the "gold standard" for identifying DPD deficiency. Additional studies are needed to investigate correlations between phenotypic measures and 5-FU toxicity in patients, potentially as secondary biomarkers in conjunction with genetic tests.