Pretreatment serum uracil concentration as a predictor of severe and fatal fluoropyrimidine-associated toxicity

Abstract

Background: We investigated the predictive value of dihydropyrimidine dehydrogenase (DPD) phenotype, measured as pretreatment serum uracil and dihydrouracil concentrations, for severe as well as fatal fluoropyrimidine-associated toxicity in 550 patients treated previously with fluoropyrimidines during a prospective multicenter study.

Methods: Pretreatment serum concentrations of uracil and dihydrouracil were measured using a validated LC-MS/MS method. The primary endpoint of this analysis was global (any) severe fluoropyrimidine-associated toxicity, that is, grade ⩾3 toxicity according to the NCI CTC-AE v3.0, occurring during the first cycle of treatment. The predictive value of uracil and the uracil/dihydrouracil ratio for early severe fluoropyrimidine-associated toxicity were compared. Pharmacogenetic variants in DPYD (c.2846A>T, c.1679T>G, c.1129-5923C>G, and c.1601G>A) and TYMS (TYMS 5'-UTR VNTR and TYMS 3'-UTR 6-bp ins/del) were measured and tested for associations with severe fluoropyrimidine-associated toxicity to compare predictive value with DPD phenotype. The Benjamini-Hochberg false discovery rate method was used to control for type I errors at level q<0.050 (corresponding to P<0.010).

Results: Uracil was superior to the dihydrouracil/uracil ratio as a predictor of severe toxicity. High pretreatment uracil concentrations (>16 ng ml^-1) were strongly associated with global severe toxicity (OR 5.3, P=0.009), severe gastrointestinal toxicity (OR 33.7, P<0.0001), toxicity-related hospitalisation (OR 16.9, P<0.0001), as well as fatal treatment-related toxicity (OR 44.8, P=0.001). None of the DPYD variants alone, or TYMS variants alone, were associated with severe toxicity.

Conclusions: High pretreatment uracil concentration was strongly predictive of severe, including fatal, fluoropyrimidine-associated toxicity, and is a highly promising phenotypic marker to identify patients at risk of severe fluoropyrimidine-associated toxicity.

Figure 1

Selection of study population for analysis. DPD=dihydropyrimidine dehydrogenase; DPYD=dihydropyrimidine dehydrogenase (gene); TYMS=thymidylate synthase (gene).

Figure 2

Correlation between pretreatment serum uracil concentrations and DPD activity in healthy volunteers and distribution of uracil concentrations in patients. Correlation between dihydropyrimidine dehydrogenase activity in peripheral blood mononuclear cells and uracil plasma levels (r=−0.51, P=0.023) in 20 healthy volunteers (A, Figure adapted from Jacobs et al (2016a) with permission). Distribution of pretreatment serum uracil concentrations in the entire cohort of 550 patients treated with fluoropyrimidine-based chemotherapy (B). U=uracil; DPD=dihydropyrimidine dehydrogenase.

Figure 3

Associations of pretreatment serum uracil concentrations with toxicity outcomes. Associations of pretreatment serum uracil concentrations with toxicity outcomes in the entire population of 550 patients. * 2/17 patients (12%) in the uracil >16 ng ml^−l group had fatal treatment-related toxicity, compared to 2/500 patients (0.4%) among patients with uracil concentrations <13 ng ml⁻¹. Fatal treatment-related toxicity did not occur among patients with pretreatment U concentrations of 13–13.8 or 13.9–16 ng ml⁻¹. Associations with fatal toxicity were determined with adjustment for age and gender but not treatment regimen (due to the low number of events). OR=odds ratio; CI=confidence interval.

Figure 4

Risk of global severe toxicity and severe gastrointestinal toxicity at varying cutoff levels for pretreatment serum uracil. Results from the analysis to estimate the risk of global severe (grade ⩾3) toxicity and severe gastrointestinal toxicity at varying cutoffs for pretreatment uracil concentration in the original dataset, adjusted for age, gender, and treatment regimen. The solid line depicts the estimated odds ratio for risk of severe toxicity for patients with pretreatment uracil concentrations above the cutoff, vs patients with uracil concentrations below the cutoff. The dashed lines represent 95% confidence intervals. Odds ratios and 95% confidence intervals are shown on a log scale.

Figure 5

Results of the pharmacogenetic analysis. Results of the pharmacogenetic analysis investigating associations between DPYD variants (A) and TYMS variants (B) in the primary cohort of 550 patients. The frequencies of early hand-foot syndrome and cardiological toxicity were too low in the population to investigate associations with these individual types of toxicity. The associations mentioned as ‘not estimable’ could not be estimated due to too few events of severe toxicity. For the TYMS variants, the results are shown for the log-additive pharmacogenetic model. The other models (dominant, recessive, or risk score) resulted in similar, non-significant, associations between TYMS genotypes and toxicity outcomes (details not shown). Results of the pharmacogenetic analysis in the larger cohort of 1613 patients are shown in C. OR=odds ratio; CI=confidence interval.

Figure 6

Relationships between DPYD variants and pretreatment uracil concentrations. The figure shows pretreatment serum uracil concentrations by DPYD variant. The horizontal lines represent median concentrations. Overall, DPYD variants were associated with an increase of pretreatment uracil concentrations of 12% (P=0.003). The c.2846A>T and c.1679T>G variants were associated with significantly higher uracil concentrations (+82%, P<0.001 and +41%, P=0.024, respectively). In contrast, c.1129-5923C>G and c.1601G>A were not significantly associated with pretreatment uracil concentrations (+12%, P=0.105 and −1%, P=0.431, respectively). * wild type for DPYD*2A, c.2846A>T, c.1679T>G, c.1129-5923C>G, and c.1601G>A. ** the cohort of 550 patients contained 2 patients with the c.1679T>G variant. In view of the low frequency of this variant, the third patient who carried c.1679T>G from the entire cohort of 1613 patients was phenotyped solely for the purpose of this analysis investigating the association between DPYD variants and pretreatment serum uracil concentration (this patient received chemoradiotherapy and was therefore excluded from the main analysis).