Marcadores farmacogenéticos asociados a toxicidad severa a fluoropirimidinas. Utilidad y viabilidad en la clínica

Abstract

Introduction: Fluoropyrimidines (5-fluorouracil and capecitabine) are chemotherapy drugs with proven efficacy for the treatment of solid tumors, but with important toxicity associated. The search for biomarkers that could help identify those patients with a higher risk of adverse events is paramount to individualize treatment and diminish the impact these toxicities have on patients and on the Healthcare System. Several genetic polymorphisms, mainly in DPYD gen, have been associated to fluoropyrimidine adverse events, but in many cases evidence is still insufficient to incorporate them into clinical practice and/or the cost-effectiveness of their implementation has not been evaluated. This is why the objectives of this work were: to evaluate the cost-effectiveness of the implementation in the clinical practice of a DPYD genotyping test that includes three variants clearly associated with fluoropyrimidine toxicity, to identify polymorphisms associated with fluoropyrimidinebased chemotherapy in genes MTHFR, CDA, TYMS, ABCB1 and ENOSF1 and to identify new variants in DPYD associated with DPD deficit and early severe fluoropyrimidine toxicity. Material and methods: Observational ambispective multicenter study in patients with solid tumors treated with fluoropyrimidine-based chemotherapy (5-fluorouracil or capecitabina). The study was approved by the Ethics Committees of all participating hospitals and all patients signed and informed consent form. The genotyping techniques used included real-time PCR with TaqMan probes, SNapshot, PCR and fragment length analysis by electrophoresis and Sanger sequencing. DPD activity was measured indirectly by measurement of endogenous uracil and dihydrouracil. Adverse events were classified according to Common Terminology Criteria for Adverse Events (CTCAE). Results: Cost per patient of the DPYD genotyping test including 3 SNPS in the proposed conditions was 6.4€ and the estimated cost for the management of one severe neutropenia was 3,044€. This test would therefore be cost-effective if it managed to avoid at least 2.21 cases per 1,000 treated patients. In patients with colorectal cancer treated with capecitabina, ABCB1*1 haplotype was associated with greater risk of severe global toxicity, diarrhea and dose delays and reductions. CDA rs2072671 polymorphism was associated with greater risk of severe global toxicity, TYMS rs45445694 with hand-foot syndrome (HFS), dose delays and reductions and TYMS rs45445694 and ENOSF1 rs2612091 with moderate-severe HFS. A score based on the combination of CDA- ABCB1 risk polymorphisms was able to predict the risk of severe toxicity with a PPV of 54% and a sensitivity of 43%. DPYD exon sequencing in patients with early severe fluoropyrimidine toxicity revealed that 90% of them presented at least one coding variant or close to the exon in DPYD and 42% were categorized as possibly damaging by the algorithms used. This technique allowed us to discover variant c.2242+1G>T, that generates a non-functional protein and to increase available knowledge on poorly studied deleterious variants such as p.R696H and p.L775W. Measurement of uracil and dihydrouracil in plasma confirmed partial DPD activity deficit in heterozygous carriers of damaging variants c.2242+1G>T, c.1645G>C, c.257C>T and c.2324T>G. Conclusions: Genotyping of rs3918290, rs67376798 and rs55886062 in DPYD with TaqMan probes is cost-effective to avoid severe adverse reactions with fluoropyrimidine treatment. The addition of polymorphisms in ABCB1, CDA, ENOSF1 y TYMS to the screening could help predict severe adverse reactions to capecitabina and improve the test´s predictive capability, although more studies are needed to confirm its clinical utility. In patients highly suspicious of DPD activity deficit, the sequential approach with whole gene sequencing, variant analysis and measurement of protein activity proved to be useful to identify and characterize rare deleterious variants and helped establish a causal relationship with the observed toxicity.