# Genetic Variants Associated with Fluoropyrimidine-Induced Toxicity in Real-World Patients After Pre-Emptive DPYD Pharmacogenetic Testing

## TL;DR

This study finds that some patients still experience severe side effects from fluoropyrimidine chemotherapy even after standard genetic testing, suggesting the need for more comprehensive genetic strategies.

## Contribution

The study identifies rare DPYD variants and the roles of ENOSF1 and TYMS in toxicity, expanding beyond standard pharmacogenetic testing.

## Key findings

- ENOSF1 rs2612091 C allele is linked to shorter time to dose reduction and fluoropyrimidine withdrawal.
- TYMS rs11280056 is associated with toxicity in patients treated with 5-fluorouracil.
- Rare DPYD variants, including a splice site and stop-gained mutation, were more common in patients with toxicity.

## Abstract

Background/Objectives: As a treatment, fluoropyrimidines are often associated with early moderate-to-severe toxicity. Although pre-emptive DPYD genotyping enables genotype-guided dosing, significant adverse events still occur in patients classified as DPYD wild-type (WT). The aim of this study was to identify DPYD variants and evaluate the contributions of TYMS, ENOSF1, CDH4, and CDA variants to fluoropyrimidine toxicity. Methods: A total of 256 European ancestry patients (aged ≥ 18 years, had completed ≥6 cycles of chemotherapy, and were WT for the DPYD variants routinely tested for) underwent genotyping for TYMS, ENOSF1, CDH4, and CDA variants, and full DPYD exon sequencing was performed in 56 of these patients. Toxicity was defined as fluoropyrimidine-related adverse events requiring a dose reduction. Multivariable models were adjusted for sex, fluoropyrimidine type, and the Charlson Comorbidity Index, and time-to-event was assessed using the Kaplan–Meier/Cox proportional hazards models. Results: A subgroup of 117 patients experienced toxicity requiring a dose reduction. The most frequent events were asthenia, gastrointestinal toxicity, hand–foot syndrome, and haematological toxicity. The ENOSF1 rs2612091 C allele was associated with fluoropyrimidine withdrawal and a shorter time to dose reduction. In the patients treated with 5-fluorouracil, TYMS rs11280056 was associated with toxicity. DPYD exon sequencing identified thirteen variants, nine of which were more prevalent in the toxicity group. These included a canonical splice site (c.150+1G>A) and a stop-gained (c.1863G>A) variant, which is predicted to result in loss of function. Conclusions: In real-world practice, despite undergoing standard DPYD genotyping, DPYD WT patients receiving a full dose of fluoropyrimidines develop clinically relevant toxicity. The presence of rare DPYD variants and associated genes (ENOSF1 and TYMS) suggests that broader, prospectively validated pharmacogenetic strategies may improve toxicity prevention.