Dihydropyrimidine Dehydrogenase Deficiency Research Articles

Patients homozygous for DPYD c.1129-5923C>G/haplotype B3 have partial DPD deficiency and require a dose reduction when treated with fluoropyrimidines.

Dihydropyrimidine dehydrogenase (DPD) is a critical determinant of 5-fluorouracil pharmacology, and reduced activity of DPD as a result of deleterious polymorphisms in the gene encoding DPD (DPYD) can result in severe treatment-related toxicity. Dosing recommendations to individualize treatment have been provided for three DPYD variants (DPYD*2A, c.2846A>T, and c.1679T>G). A fourth variant, c.1129-5923C>G/HapB3, has been shown to increase the risk of fluoropyrimidine-associated toxicity, but little is known about the functional effects of this variant. By performing a large retrospective screen for DPYD variants, we identified three patients who were homozygous for c.1129-5923C>G/HapB3. We describe their clinical course of treatment and analyzed DPD activity and DPYD gene expression, to provide insight into the phenotypic effects of c.1129-5923C>G/HapB3. DPD activity could be measured in two patients and was 4.1 and 5.4nmol/mg/h (DPD activity 41 and 55% compared to controls, respectively). The fluoropyrimidine dose had to be reduced during treatment in both patients. In line with partial DPD deficiency in both patients, sequence analysis of DPD cDNA demonstrated a normal-sized (wild type) cDNA fragment of 486 bp, as well as a larger-sized (mutant) 530-bp fragment containing an aberrant 44-bp insertion in intron 10. Patient three tolerated treatment well, but DPD activity measurement was not possible as the patient had deceased at the time of performing the study. The presented functional and clinical data indicate that the c.1129-5923C>G variant is both functionally and clinically relevant, and support an upfront dose reduction of the fluoropyrimidine starting dose in patients carrying c.1129-5923C>G homozygously.

Highlight on DPYD gene polymorphisms and treatment by capecitabine

Background: Sequencing of DPYD exome was conducted in a prospective cohort of advanced breast cancer patients receiving capecitabine.Methods: A total of 243 patients were analyzed. Digestive, neurologic and hematotoxicity over cycles 1–2 showed 10.3% G3 and 2.1% G4, including one toxic death. DPYD exome, flanking intronic regions (20 bp), 3’UTR and part of 5’UTR (500 bp) were sequenced on MiSeq Illumina (Integragen, 97% coverage, HWE checked).Results: In total, 48 SNPs were identified: three in 3’UTR, 19 in coding regions (four synonymous including E412E; 15 missenses including D949V, V732I, R592W, I560S, I543V, S534N, S492L, M406I, D342G, M166V, T65M, C29R), 19 in flanking intronic regions (including *2A) and seven in 5’UTR. In total, 11 SNPs have not been previously described, including three missense variations each heterozygous in three separate patients: R696H, F100L and A26T. The patient with a toxic death carried one D949V allele. The three consensual variants *2A, D949V and I560S were carried by seven patients (heterozygous). Analysis of consensual variants showed that they were associated with G3–4 toxicity (OR = 21.0, sensitivity 16.7%) but not with G4 toxicity. Adding the variants previously associated with DPD deficiency in vitro, i.e. R592W, S492L and D342N/G, increased sensitivity on G3–4 (23.3%, OR = 21.1) and was predictive of G4 toxicity (sensitivity 40%, OR = 19.0). Of note, adding the new F100L variant further improved predictivity of genotyping on G4 toxicity (sensitivity 60%, OR = 42.8).Conclusions: Present data establish the impact of consensual variants on capecitabine toxicity and reveal the existence of a novel DPYD variant, F100L, associated with G4 toxicity.

Endogenous plasma and salivary uracil to dihydrouracil ratios and DPYD genotyping as predictors of severe fluoropyrimidine toxicity in patients with gastrointestinal malignancies

ObjectiveThe aim of this study was to evaluate the use of plasma and saliva uracil (U) to dihydrouracil (UH2) metabolic ratio and DPYD genotyping, as a means to identify patients with dihydropyrimidine dehydrogenase (DPD) deficiency and fluoropyrimidine toxicity. MethodsPaired plasma and saliva samples were obtained from 60 patients with gastrointestinal cancer, before fluoropyrimidine treatment. U and UH2 concentrations were measured by LC–MS/MS. DPYD was genotyped for alleles *7, *2A, *13 and Y186C. Data on toxicity included grade 1 to 4 neutropenia, mucositis, diarrhea, nausea/vomiting and cutaneous rash. Results35% of the patients had severe toxicity. There was no variant allele carrier for DPYD. The [UH2]/[U] metabolic ratios were 0.09–26.73 in plasma and 0.08–24.0 in saliva, with higher correlation with toxicity grade in saliva compared to plasma (rs=−0.515 vs rs=−0.282). Median metabolic ratios were lower in patients with severe toxicity as compared to those with absence of toxicity (0.59 vs 2.83 saliva; 1.62 vs 6.75 plasma, P<0.01). A cut-off of 1.16 for salivary ratio was set (AUC 0.842), with 86% sensitivity and 77% specificity for the identification of patients with severe toxicity. Similarly, a plasma cut-off of 4.0 (AUC 0.746), revealed a 71% sensitivity and 76% specificity. ConclusionsDPYD genotyping for alleles 7, *2A, *13 and Y186C was not helpful in the identification of patients with severe DPD deficiency in this series of patients. The [UH2]/[U] metabolic ratios, however, proved to be a promising functional test to identify the majority of cases of severe DPD activity, with saliva performing better than plasma.

Dihydro pyrimidine dehydrogenase deficiency in patients treated with capecitabine based regimens: a tertiary care centre experience.

Dihydropyrimidine dehydrogenase (DPD) enzyme is the rate limiting step in the metabolism of capecitabine, and its deficiency leads to severe toxicities and rarely, death. A total of 506 patients were treated in the GI Medical Oncology unit of our institution with capecitabine containing regimens with a dose range of 1,250 to 2,000 mg/m(2)/day during the period from June 2013 to June 2015. Patients with grade (Gr) 3/4 toxicities requiring in-patient care (life threatening complications) were planned for DPD activity testing by peripheral blood PCR sequencing. Thirty-one patients developed Gr 3/4 toxicities during cycle 1 of capecitabine. This included mucositis in 24 (77%), diarrhea in 29 (94%), hand-foot syndrome (HFS) in 13 (42%) and myelosuppression in 5 (16%) patients. Twenty-two (81.4%) were found to be DPD deficient with 6 patients negative for DPD mutation. Three patients did not undergo the DPD analysis as advised. More than one mutation was seen in 11 patients. The relative frequencies of the mutations were IVS14+1G→A in 39%, with 13% having homozygosity, 85 T→C in 36%, 1627 A→G in 32%, 496 A→G in 18% and 2194 G→A in 18%, respectively. After dose reduction in cycle 2 in 17 patients of the DPD mutation positive cohort, statistically significant reduction in the toxicities was seen. Dose reduction in DPD deficient patients, reduces risk of life threatening complications significantly but not completely. Upfront screening for DPD deficiency in Indian patients should be evaluated further in view of potentially high homozygous DPD mutation prevalence.

Development and validation of a rapid and sensitive UPLC–MS/MS method for determination of uracil and dihydrouracil in human plasma

Quantification of the endogenous dihydropyrimidine dehydrogenase (DPD) substrate uracil (U) and the reaction product dihydrouracil (UH2) in plasma might be suitable for identification of patients at risk of fluoropyrimidine-induced toxicity as a result of DPD deficiency. In this paper, we describe the development and validation of a rapid and sensitive ultra-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) assay for quantification of U and UH2 in human plasma.Analytes were extracted by protein precipitation, chromatographically separated on an Acquity UPLC® HSS T3 column with gradient elution and analyzed with a tandem mass spectrometer equipped with an electrospray ionization source. U was quantified in the negative ion mode and UH2 in the positive ion mode. Stable isotopes for U and UH2 were used as internal standards.Total chromatographic run time was 5min. Validated concentration ranges for U and UH2 were from 1 to 100ng/mL and 10 to 1000ng/mL, respectively. Inter-assay bias and inter-assay precision for U were within ±2.8% and ≤12.4%. For UH2, inter-assay bias and inter-assay precision were within ±2.9% and ≤7.2%. Adequate stability of U and UH2 in dry extract, final extract, stock solution and plasma was demonstrated. Stability of U and UH2 in whole blood was only satisfactory when stored up to 4hours at 2–8°C, but not at ambient temperatures.An accurate, precise and sensitive UPLC-MS/MS assay for quantification of U and UH2 in plasma was developed. This assay is now applied to support clinical studies with fluoropyrimidine drugs.

Germline pharmacogenomics in patients with dihydropyrimidine dehydrogenase (DPD) deficiency.

521 Background: DPD deficiency is a pharmacogenetic syndrome associated with dose-limiting toxicity to fluoropyrimidines. Oncologists are expected to recognize and diagnose this syndrome, as toxicities could be fatal. Over 40 single nucleotide polymorphisms (SNPs) and deletions have been identified within the DPYD gene. IVS14+1G&gt;A (DPYD*2A) is the most common (40-50%) and best studied of these SNPs. Yet, it showed a median sensitivity of 30% and is absent in Japanese, Korean and African Americans. Overall, the data on DPYD testing is insufficient to provide enough guidance to diagnose DPD deficiency. Herein we describe our experience with germline pharmacogenomics in patients with DPD deficiency. Methods: Between 2011 and 2015, 35 patients with gastrointestinal malignancies were tested for DPYD mutations; 17 were tested after developing toxicities to treatment and 18 were tested prior to treatment. IVS14+1G&gt;A (DPYD*2A) was tested in all patients. DPYD c.85T&gt;C (DPYD*9A), DPYD c.1679T&gt;G (DPYD*13A), DPYD c.-1590T&gt;C, and DPYD c.2846A&gt;T were tested in 24 patients (69%) only. We explored the association between DPYD mutations and fluoropyrimidine-related toxicity using Fisher’s exact test. Results: Median age was 60 years, 43% were male, 80% were Caucasian and 20% were African American. Capecitabine-based regimens (71%) and 5-Fluorouracil-based regimens (29%). 14 out of 35 patients (40%) had DPYD mutations. Grade 3 toxicities were encountered in 64% of patients with DPYD mutation and 48% of patients with no DPYD mutation. In patients who received full dose fluoropyrimidines (57% of patients with DPYD mutation and 81% of patients with no DPYD mutation), DPYD mutations were associated with a significantly higher rate of grade 3 diarrhea (p=0.026). In patients with DPYD mutation, 2 (14%) had DPYD*A2 mutation and 12 (86%) had DPYD*9A mutation. Conclusions: In patients treated with fluoropyrimidines, the rate of grade 3 diarrhea was significantly higher in patients with mutated DPYD gene. Testing for DPYD*2A alone to diagnose DPD deficiency is suboptimal. Testing for other DPYD mutation variants including DPYD*9A provides a more comprehensive approach. These data should further be validated in prospective clinical trials.

Dihydro pyrimidine dehydrogenase deficiency in patients treated with capecitabine-based regimens: A retrospective tertiary care centre experience.

712 Background: Capecitabine is an integral part of treatment of gastrointestinal cancers. Dihydropyrimidine dehydrogenase (DPD) enzyme is rate limiting in the metabolism of capecitabine, deficiency of which leads to myelsuppression, mucositis, diarrhea, hand foot syndrome (HFS) and rarely, death. Data regarding the toxicity of capecitabine in patients with DPD deficiency in the Indian context is scarce. Methods: 506 patients were treated with capecitabine containing regimens with a dose range of 1250 mg/m2/day to 2000 mg/m2/day during the period from June 2013 to May 2015 in the Gastrointestinal Medical Oncology Unit of our institution. Patients with Grade 3/4 toxicities requiring in-patient care (life threatening complications) were planned for DPD activity testing by Peripheral Blood PCR sequencing. Results: 27 patients developed Gr 3/4 toxicities during cycle 1 of capecitabine. It included mucositis in 22 (81.5 %), diarrhea in 25 (92.6%), HFS in 10 (37%) and myelosuppression in 4 (14.8%). 19 were found to be DPD deficient with 5 patients negative for DPD mutation. 3 patients did not do the DPD analysis as advised. Homozygous mutations were seen in 9 (33.3%) and heterozygous mutations in 10 (27%) of patients. More than one mutation was seen in 10 patients (37%). The relative frequencies of mutation were Exon 14 (44.4%), Exon 2 (25.9%), Exon 13 (25.9%), Exon 6 (11.1%) and Exon 18 (7.4 %). Post Cycle 1 of capecitabine, the drug was stopped in 5 patients (18.5%), regimen changed in 2 (7.4%) and dose reduction by 50% of the drug was done in the remaining patients. Despite dose reduction and change in therapy during Cycle 2, patients still had Grade 3/4 toxicities including mucositis in 7(25.9%), diarrhoea in 10 (37%), HFS in 7 (25.9 %) and myelosuppression in 6 (22%) of patients. Conclusions: Capecitabine can also lead to severe toxicities in DPD-deficient patients. Dose reduction of capecitabine in DPD deficient patients may not completely ameliorate the future risk of life – threatening complications. Screening for DPD deficiency prior to administration of capecitabine in toxicity prone nutritionally deficient Indian patients should be further evaluated based on this data.

Clinical trial experience with uridine triacetate for 5-fluorouracil toxicity.

655 Background: Life-threatening or lethal 5-FU toxicity occurs due to infusion pump/reservoir errors and 5-FU dosage miscalculations. Additionally, DPD-deficient patients often experience serious or lethal toxicity. Regardless of DPD status, some patients demonstrate severe, early-onset toxicities (as early as during 5-FU infusion) such as encephalopathy or cardiotoxicity. Uridine triacetate is an orally bioavailable prodrug of uridine, a direct antagonist of 5-FU, that competitively inhibits 5-FU incorporation in normal tissues and can counter 5-FU toxicity, reducing morbidity and mortality due to toxicity. Methods: 135 patients at excess risk of 5-FU toxicity due to overdose or accidental capecitabine ingestion (n = 111); or DPD deficiency and/or who showed rapid onset of severe toxicities (n = 24) have been treated with uridine triacetate under emergency IND or expanded access protocols. Patients were to receive uridine triacetate granules (10g q6h for 20 doses) up to 96h after the termination of 5-FU therapy. Clinical endpoints included survival compared with historical controls, time to resumption of chemotherapy, and safety. Results: A total of 130/135 (96%) patients treated with uridine triacetate recovered fully (within 30 days after receiving uridine triacetate), including rapid reversal of cardiotoxicity (EKG abnormalities, chest pain, etc) and neurotoxicity (altered mental status, ataxia, etc). Of the 111 overdose patients, 24 had a pump malfunction; 49 dose error; 5 had accidental capecitabine ingestion (2 adults/3 peds); 33 unknown. No patient who received uridine triacetate within 96 h after stopping 5-FU died. Comparatively, 38 of the 47 historical controls (81%) with 5-FU overexposure died. Of the 106 overdose patients with a diagnosis of cancer, 40 (37.7%) resumed chemotherapy within 30 days (median 20.0 days post-5-FU), highly indicative of rapid recovery from toxicity. Overall compliance was 92.5% (mean 18.3/20 doses). Mild, infrequent adverse events (diarrhea/nausea/vomiting) were attributed to uridine triacetate. Conclusions: In this study, uridine triacetate was a safe and effective life-saving antidote for 5-FU overexposure, and facilitated rapid resumption of chemotherapy. Clinical trial information: NCT01432301.

Phenotypic and clinical implications of variants in the dihydropyrimidine dehydrogenase gene

Dihydropyrimidine dehydrogenase (DPD) is the initial and rate-limiting enzyme in the catabolism of the pyrimidine bases uracil, thymine and the antineoplastic agent 5-fluorouracil. Genetic variations in the gene encoding DPD (DPYD) have emerged as predictive risk alleles for 5FU-associated toxicity. Here we report an in-depth analysis of genetic variants in DPYD and their consequences for DPD activity and pyrimidine metabolites in 100 Dutch healthy volunteers. 34 SNPs were detected in DPYD and 15 SNPs were associated with altered plasma concentrations of pyrimidine metabolites. DPD activity was significantly associated with the plasma concentrations of uracil, the presence of a specific DPYD mutation (c.1905+1G>A) and the combined presence of three risk variants in DPYD (c.1905+1G>A, c.1129-5923C>G, c.2846A>T), but not with an altered uracil/dihydrouracil (U/UH2) ratio. Various haplotypes were associated with different DPD activities (haplotype D3, a decreased DPD activity; haplotype F2, an increased DPD activity). Functional analysis of eight recombinant mutant DPD enzymes showed a reduced DPD activity, ranging from 35% to 84% of the wild-type enzyme. Analysis of a DPD homology model indicated that the structural effect of the novel p.G401R mutation is most likely minor. The clinical relevance of the p.D949V mutation was demonstrated in a cancer patient heterozygous for the c.2846A>T mutation and a novel nonsense mutation c.1681C>T (p.R561X), experiencing severe grade IV toxicity. Our studies showed that the endogenous levels of uracil and the U/UH2 ratio are poor predictors of an impaired DPD activity. Loading studies with uracil to identify patients with a DPD deficiency warrants further investigation.

Beating the odds: efficacy and toxicity of dihydropyrimidine dehydrogenase-driven adaptive dosing of 5-FU in patients with digestive cancer.

5-FU is the backbone of most regimens in digestive oncology. Administration of standard 5-FU leads to 15-30% of severe side effects, and lethal toxicities are regularly reported with fluoropyrimidine drugs. Dihydropyrimidine dehydrogenase (DPD) deficiency is a pharmacogenetic syndrome responsible for most cases of life-threatening toxicities upon 5-FU intake, and pre-treatment checking for DPD status should help to reduce both incidence and severity of side effects through adaptive dosing strategies. We have used a simple method for rapidly establishing the DPD phenotype of patients with cancer and used it prospectively in 59 routine patients treated with 5-FU-based therapy for digestive cancers. No patient with total DPD deficiency was found but 23% of patients exhibited poor metabolizer phenotype, and one patient was phenotyped as profoundly deficient. Consequently, 5-FU doses in poor metabolizer patients were cut by an average 35% as compared with non deficient patients (2390 ± 1225 mg vs. 3653 ± 1371 mg, P < 0.003, t-test). Despite this marked reduction in 5-FU dosing, similar efficacy was achieved in the two subsets (clinical benefit: 40 vs. 43%, stable disease: 40 vs. 37%, progressive disease: 20% in both subsets, P = 0.893, Pearson's chi-square). No difference in toxicities was observed (P = 0.104, Fisher's exact test). Overall, only 3% of early severe toxicities were recorded, a value markedly lower than the 15-30% ones usually reported with 5-FU. This feasibility study shows how simplified DPD-based adaptive dosing of 5-FU can reduce sharply the incidence of treatment-related severe toxicities while maintaining efficacy as part of routine clinical practice in digestive oncology.

DPD deficiency in patients treated with fluorouracil

In The Lancet Oncology, Didier Meulendijks and colleagues report the findings from their meta-analysis of eight studies of DPYD genetic variants as predictors of fluoropyrimidine-induced severe toxicity. 1 Meulendjiks D Henricks LM Sonke GS et al. Clinical relevance of DPYD variants c.1679T>G, c.1236G>A/HapB3, and c.1601G>A as predictors of severe fluoropyrimidine-associated toxicity: a systematic review and meta-analysis of individual patient data. Lancet Oncol. 2015; (published online Oct 23.)http://dx.doi.org/10.1016/S1470-2045(15)00286-7 Google Scholar In Meulendijks and colleagues' meta-analysis of 7365 patients, DPYD c.1679T>G and c.1236G>A/HapB3 were significantly associated with fluoropyrimidine-associated toxicity (adjusted relative risk 4·40, 95% CI 2·08–9·30, p<0·0001; and 1·59, 1·29–1·97, p<0·0001, respectively). The investigators noted consistent associations of c.1679T>G and c.1236G>A/HapB3 with gastrointestinal and haematological toxicities, but not with hand-foot syndrome. Dihydropyrimidine dehydrogenase (DPD) deficiency is a pharmacogenetic syndrome resulting in an inability or impaired ability in some patients to detoxify fluorouracil in the liver. Consequently, DPD-deficient patients will have life-threatening toxicities with standard doses of fluorouracil. Clinical relevance of DPYD variants c.1679T>G, c.1236G>A/HapB3, and c.1601G>A as predictors of severe fluoropyrimidine-associated toxicity: a systematic review and meta-analysis of individual patient dataDPYD variants c.1679T>G and c.1236G>A/HapB3 are clinically relevant predictors of fluoropyrimidine-associated toxicity. Upfront screening for these variants, in addition to the established variants DPYD*2A and c.2846A>T, is recommended to improve the safety of patients with cancer treated with fluoropyrimidines. Full-Text PDF

Clinical relevance of DPYD variants c.1679T>G, c.1236G>A/HapB3, and c.1601G>A as predictors of severe fluoropyrimidine-associated toxicity: a systematic review and meta-analysis of individual patient data

The best-known cause of intolerance to fluoropyrimidines is dihydropyrimidine dehydrogenase (DPD) deficiency, which can result from deleterious polymorphisms in the gene encoding DPD (DPYD), including DPYD*2A and c.2846A>T. Three other variants-DPYD c.1679T>G, c.1236G>A/HapB3, and c.1601G>A-have been associated with DPD deficiency, but no definitive evidence for the clinical validity of these variants is available. The primary objective of this systematic review and meta-analysis was to assess the clinical validity of c.1679T>G, c.1236G>A/HapB3, and c.1601G>A as predictors of severe fluoropyrimidine-associated toxicity. We did a systematic review of the literature published before Dec 17, 2014, to identify cohort studies investigating associations between DPYD c.1679T>G, c.1236G>A/HapB3, and c.1601G>A and severe (grade ≥3) fluoropyrimidine-associated toxicity in patients treated with fluoropyrimidines (fluorouracil, capecitabine, or tegafur-uracil as single agents, in combination with other anticancer drugs, or with radiotherapy). Individual patient data were retrieved and analysed in a multivariable analysis to obtain an adjusted relative risk (RR). Effect estimates were pooled by use of a random-effects meta-analysis. The threshold for significance was set at a p value of less than 0·0167 (Bonferroni correction). 7365 patients from eight studies were included in the meta-analysis. DPYD c.1679T>G was significantly associated with fluoropyrimidine-associated toxicity (adjusted RR 4·40, 95% CI 2·08-9·30, p<0·0001), as was c.1236G>A/HapB3 (1·59, 1·29-1·97, p<0·0001). The association between c.1601G>A and fluoropyrimidine-associated toxicity was not significant (adjusted RR 1·52, 95% CI 0·86-2·70, p=0·15). Analysis of individual types of toxicity showed consistent associations of c.1679T>G and c.1236G>A/HapB3 with gastrointestinal toxicity (adjusted RR 5·72, 95% CI 1·40-23·33, p=0·015; and 2·04, 1·49-2·78, p<0·0001, respectively) and haematological toxicity (adjusted RR 9·76, 95% CI 3·03-31·48, p=0·00014; and 2·07, 1·17-3·68, p=0·013, respectively), but not with hand-foot syndrome. DPYD*2A and c.2846A>T were also significantly associated with severe fluoropyrimidine-associated toxicity (adjusted RR 2·85, 95% CI 1·75-4·62, p<0·0001; and 3·02, 2·22-4·10, p<0·0001, respectively). DPYD variants c.1679T>G and c.1236G>A/HapB3 are clinically relevant predictors of fluoropyrimidine-associated toxicity. Upfront screening for these variants, in addition to the established variants DPYD*2A and c.2846A>T, is recommended to improve the safety of patients with cancer treated with fluoropyrimidines. None.

Discovery of novel mutations in the dihydropyrimidine dehydrogenase gene associated with toxicity of fluoropyrimidines and viewpoint on preemptive pharmacogenetic screening in patients.

BackgroundDihydropyrimidine dehydrogenase (DPD) is the initial and rate-limiting enzyme of the metabolic pathway of 5-fluorouracil (5-FU) and other fluoropyrimidines to inactive compounds. For this reason, severe, life-threatening toxicities may occur in patients with deficient DPD activity when administered standard doses of 5-FU and its prodrugs.Materials and methodsWe selected three patients with colorectal adenocarcinoma who displayed unexpected severe adverse reactions after treatment with 5-FU and capecitabine. To investigate the possible involvement of deficient variants of the DPD gene (DPYD), a denaturing HPLC (dHPLC) approach followed by target exon sequencing of DPYD was performed on DNA extracted from peripheral blood.ResultsThree novel non-synonymous mutations of DPYD, c.2509-2510insC, c.1801G>C, and c.680G>A, were detected in these subjects. Due to the absence of other deficient variants of DPYD and the compatibility of adverse reactions with fluoropyrimidine treatment, the novel variants were associated with a poor-metabolizer phenotype.ConclusionsStratification of patients on the basis of their genotype may help prevent toxicity, and the large body of evidence about the pathogenesis of fluoropyrimidine-induced adverse reactions strongly encourages the adoption of best practice recommendations to appropriately address this important clinical issue. This approach is of utmost importance within a preventive, prognostic, and personalized approach to patient care in the oncology setting.

Poster 86 Rehabilitation Process of 5-Fluorouracil Induced Severe Encephalopathy in a Patient with Dihydropyrimidine Dehydrogenase Deficiency: A Case Report

Poster 86 Rehabilitation Process of 5-Fluorouracil Induced Severe Encephalopathy in a Patient with Dihydropyrimidine Dehydrogenase Deficiency: A Case Report

Impairment of energy metabolism in cardiomyocytes caused by 5-FU catabolites can be compensated by administration of amino acids.

Identification of patients with increased risk of 5-fluorouracil (5-FU)-related toxicity is an important challenge for cancer treatment. Research often focus on dihydropyrimidine dehydrogenase (DPYD) deficiency in this context. However, patients with normal DPYD activity may also develop life-threatening 5-FU adverse effects. DPYD initiates the catabolic route of 5-FU generating metabolites such as fluoroacetate (FAC). The catabolite FAC is known to inhibit the TCA cycle enzyme aconitase, which is supposed to impair mitochondrial energy metabolism. Therefore, we aim for a systems understanding of the association of 5-FU-related cardiac side effects with aconitase inhibition caused by FAC. Using a mitochondrial model of cardiomyocytes we found strong depletion of ATP production and citrate accumulation as main effects of aconitase inhibition. Shadow price analysis revealed that the uptakes of valine, arginine, proline and glutamate are most effective in compensating the impairment of energy metabolism. Our findings suggest that 5-FU catabolism contributes to the occurrence of cardiac adverse effects and are the basis for further biomarker identifications and development of side effect treatment.

Translating DPYD genotype into DPD phenotype: using the DPYD gene activity score.

The dihydropyrimidine dehydrogenase enzyme (DPD, encoded by the gene DPYD) plays a key role in the metabolism of fluoropyrimidines. DPD deficiency occurs in 4-5% of the population and is associated with severe fluoropyrimidine-related toxicity. Several SNPs in DPYD have been described that lead to absent or reduced enzyme activity, including DPYD*2A, DPYD*13, c.2846A>T and c.1236G>A/haplotype B3. Since these SNPs differ in their effect on DPD enzyme activity, a differentiated dose adaption is recommended. We propose the gene activity score for translating DPYD genotype into phenotype, accounting for differences in functionality of SNPs. This method can be used to standardize individualized fluoropyrimidine dose adjustments, resulting in optimal safety and effectiveness.

DPD and UGT1A1 deficiency in colorectal cancer patients receiving triplet chemotherapy with fluoropyrimidines, oxaliplatin and irinotecan.

Triplet chemotherapy with fluoropyrimidines, oxaliplatin and irinotecan is a standard therapy for metastatic colorectal cancer (CRC). Single nucleotide polymorphisms (SNPs) in DPYD and UGT1A1 influence fluoropyrimdines and irinotecan adverse events (AEs). Low frequency DPYD variants (c.1905 + 1G > A, c.1679 T > G, c.2846A > T) are validated but more frequent ones (c.496A > G, c.1129-5923C > G and c.1896 T > C) are not. rs895819 T > C polymorphism in hsa-mir-27a is associated with reduced DPD activity. In this study, we evaluated the clinical usefulness of a pharmacogenetic panel for patients receiving triplet combinations. Germline DNA was available from 64 CRC patients enrolled between 2008 and 2013 in two phase II trials of capecitabine, oxaliplatin and irinotecan plus bevacizumab or cetuximab. SNPs were determined by Real-Time PCR. We evaluated the functional variants in DPYD (rare: c.1905 + 1G > A, c.1679 T > G, c.2846A > T; most common: c.496A > G, c.1129-5923C > G, c.1896 T > C), hsa-mir-27a (rs895819) and UGT1A1 (*28) genes to assess their association with grade 3-4 AEs. None of the patients carried rare DPYD variants. We found DPYD c.496A > G, c.1129-5923C > G, c.1896 T > C in heterozygosity in 19%, 5% and 8%, respectively, homozygous rs895819 in hsa-mir-27a in 9% and homozygous UGT1A1*28 in 8%. Grade 3-4 AEs were observed in 36% patients and were associated with DPYD c.496A > G (odds ratio (OR) 4.93, 95% CI 1.29, 18.87; P = 0.021) and homozygous rs895819 in hsa-mir-27a (OR 11.11, 95% CI 1.21, 102.09; P = 0.020). Carriers of DPYD c.1896 T > C and homozygous UGT1A1*28 showed an OR of 8.42 (95% CI 0.88, 80.56; P = 0.052). Multivariate analysis confirmed an independent value for DPYD c.496A > G and c.1896 T > C. Concomitant assessment of DPYD variants and the UGT1A1*28 allele is a promising strategy needing further validation for dose personalization.

DPYD c.1905+1G&gt;A and c.2846A&gt;T and UGT1A1*28 allelic variants as predictors of toxicity: Pharmacogenetic translational analysis from the phase III TRIBE study in metastatic colorectal cancer.

3532 Background: Adverse drug reactions (ADRs) caused by fluoropyrimidines depend, at least in part, from DPD deficiency resulting from the loss-of-function mutations c.1905+1G > A and c.2846A > T....

Influence of metastatic disease on the usefulness of uracil pharmacokinetics as a screening tool for DPD activity in colorectal cancer patients.

Dihydropyrimidine dehydrogenase (DPD) deficiency can lead to severe toxicity in patients treated with a standard dose of a fluoropyrimidine such as 5-fluorouracil or capecitabine (CAP). Administration of oral uracil and subsequent measurement of uracil and dihydrouracil (DHU) plasma concentrations has been used to identify patients with DPD deficiency. Liver metastasis might influence systemic DPD activity. The aim of the study was to investigate the effect of metastatic disease on the pharmacokinetics of uracil and DHU after oral administration of uracil. 500mg/m(2) uracil was administered orally to 12 subjects with stages II-III colorectal cancer (CRC) who were treated in the adjuvant setting and to 12 subjects with stage IV metastasized CRC, all treated with CAP containing therapy. All subjects had a normal DPD activity defined as >6nmol/mg/h determined in peripheral blood mononuclear cells. The mean uracil clearance [CL 51.7 (SD 6.4) vs. 46.7 (SD 13.0) l/h], area under the curve [AUC0-220min 20.6 (SD 6.4) vs. 21.0 (SD 5.7) hmg/l], elimination half-life [t 1/2 21 (SD 7) vs. 21 (SD 8) min], maximum concentration time [T max 27 (SD 9) vs. 25 (SD 9) min], volume of distribution [V 26.58 (SD 10.11) vs. 21.10 (SD 8.48) l] and the elimination constant [k el 2.01 (SD 0.56) vs. 2.41 (SD 0.72) h(-1)] did not differ significantly (p>0.05) non-metastatic CRD versus metastatic CRC. Metastasis does not alter uracil pharmacokinetics andis similar in CRC patients with and without metastasis. Therefore, the uracil test dose could be used as a DPD phenotype test in both adjuvantly treated and metastatic CRC patients using similar cutoff criteria to identify patients with DPD deficiency.

Dihydropyrimidine Dehydrogenase 85T&gt;C Mutation Is Associated With Ocular Toxicity of 5-Fluorouracil

5-Fluorouracil (5-FU), the mainstay of solid tumor chemotherapy over the past 40 years, induces grade III-IV toxicities in up to 15% of patients with polymorphisms in the dihydropyrimidine dehydrogenase (DPYD), thymidylate synthase (TYMS), and methylenetetrahydrofolate reductase (MTHFR) genes. These toxicities include mucositis, neutropenia, nausea, diarrhea, myelosuppression, hand-foot syndrome, and rare ocular adverse effects. Here, we present the case of a female patient with rectal cancer who received 5-FU-based chemotherapy and developed grade III hand-foot syndrome and rare acute ocular adverse effects. Genetic analysis revealed that the patient had an 85T>C mutation in the DPYD gene resulting in a DPYD*9A allele. The clinical and molecular observations indicate that DPYD deficiency may be responsible for the severe ocular adverse effects observed in 5-FU-treated patients. Application of personalized therapy based on molecular testing should help clinicians provide the most effective chemotherapy agents and dose modifications for each patient, although further population-based pharmacogenetic trials for the 5-FU metabolism-related genes are necessary to minimize adverse effects and enhance clinical outcomes.