Dihydropyrimidine Dehydrogenase Phenotyping Using Pretreatment Uracil: A Note of Caution Based on a Large Prospective Clinical Study

Abstract

In clinical practice, 25-30% of the patients treated with fluoropyrimidines experience severe fluoropyrimidine-related toxicity. Extensively clinically validated DPYD genotyping tests are available to identify patients at risk of severe toxicity due to decreased activity of dihydropyrimidine dehydrogenase (DPD), the rate limiting enzyme in fluoropyrimidine metabolism. In April 2020, the European Medicines Agency recommended that, as an alternative for DPYD genotype-based testing for DPD deficiency, also phenotype testing based on pretreatment plasma uracil levels is a suitable method to identify patients with DPD deficiency. Although the evidence for genotype-directed dosing of fluoropyrimidines is substantial, the level of evidence supporting plasma uracil levels to predict DPD activity in clinical practice is limited. Notwithstanding this, uracil-based phenotyping is now used in clinical practice in various countries in Europe. We aimed to determine the value of pretreatment uracil levels in predicting DPD deficiency and severe treatment-related toxicity. To this end, we determined pretreatment uracil levels in 955 patients with cancer, and assessed the correlation with DPD activity in peripheral blood mononuclear cells (PBMCs) and fluoropyrimidine-related severe toxicity. We identified substantial issues concerning the use of pretreatment uracil in clinical practice, including large between-center study differences in measured pretreatment uracil levels, most likely as a result of pre-analytical factors. Importantly, we were not able to correlate pretreatment uracil levels with DPD activity nor were uracil levels predictive of severe treatment-related toxicity. We urge that robust clinical validation should first be performed before pretreatment plasma uracil levels are used in clinical practice as part of a dosing strategy for fluoropyrimidines.

Figure 1

Differences in measured pretreatment uracil levels between hospitals. Differences in uracil concentrations (ng/mL) between the participating hospitals in an explorative substudy of a prospective multicenter study in 955 patients (clinicaltrials.gov identifier NCT02324452). All the samples were measured centrally therefore, the central hospital was chosen to be the reference hospital (indicated in red). Differences between medians were determined using one‐way analysis of variance (Kruskal–Wallis). *P ≤ 0.05; ***P ≤ 0.001; ****P ≤ 0.0001. [Colour figure can be viewed at wileyonlinelibrary.com]

Figure 2

Correlations of endogenous uracil levels, DPD enzyme activity in PBMCs, toxicity, and DPYD genotype. Dots represent individual results. Black lines represent the median of the data. (a) Shows the correlation between endogenous uracil levels and DPD activity. (b) Shows the endogenous uracil concentration in patients with and without severe toxicity. DPYD variants were excluded from the analysis as they received initial dose reductions based on their genotype results. (c) Shows the endogenous uracil levels in patients by DPYD‐genotype. (d) Shows the DPD enzyme activity measured in PBMCs of 138 patients (both DPYD variant carriers and wild type patients). DPD, dihydropyrimidine dehydrogenase; DPYD, gene encoding dihydropyrimidine dehydrogenase; NS, not significant; PBMCs, peripheral blood mononuclear cells; P value; vs, versus. [Colour figure can be viewed at wileyonlinelibrary.com]