Identification of a novel mutation in the dihydropyrimidine dehydrogenase gene in a patient with a lethal outcome following 5-fluorouracil administration and the determination of its frequency in a population of 500 patients with colorectal carcinoma.

OBJECTIVES: Life-threatening toxic side-effects following 5-FU exposure have been related to deficiency of dihydropyrimidine dehydrogenase (DPD), the rate-limiting enzyme in its catabolism. We presently report a new DPYD gene SNP in a Spanish woman who died from multivisceral 5-FU-induced toxicity. DESIGN AND METHODS: We looked for 22 known SNPs by Pyrosequecing. Then, we sequenced the whole 23 exons of DPYD, along with adjacent intronic sequences. PCR was carried out to determine whether or not exons were deleted in the DPYD. To determine whether the predicted stop codon indeed resulted in a truncated protein, a bacterial expression vector was employed to generate the predicted protein. 500 patients were genotyped to determine allele frequency. RESULTS: A novel mutation 464 T>A was identified in DPYD gene exon 5, resulting in the replacement of leucine 155 by a stop codon in the protein. We confirmed this mutation by Pyrosequencing and its involvement by a protein truncation test. We genotyped the patient's family and the allele frequency was 0.2%. CONCLUSION: The involvement of this variant in 5-FU life-threatening toxicity supports its inclusion in pretherapeutic genetic screening.

Correlation between uracil and dihydrouracil plasma ratio, fluorouracil (5-FU) pharmacokinetic parameters, and tolerance in patients with advanced colorectal cancer: A potential interest for predicting 5-FU toxicity and determining optimal 5-FU dosage.

PURPOSE: Patients with genetic fluorouracil (5-FU) catabolic deficiencies are at high risk for severe toxicity. To predict 5-FU catabolic deficiencies and toxic side effects, we conducted a prospective study of patients treated for advanced colorectal cancer by high-dose 5-FU. PATIENTS AND METHODS: Eighty-one patients were treated with weekly infusions of 5-FU and folinic acid. The initial 5-FU dose of 1,300 mg/m(2) was individually adjusted according to a dose-adjustment chart. Plasma concentrations of uracil (U) and its dihydrogenated metabolite, dihydrouracil (UH(2)), were measured before treatment, and the ratio of UH(2) to U was calculated. Pharmacokinetic and pharmacodynamic studies were conducted to look for a relationship between the ratio of UH(2) to U and 5-FU metabolic outcome and tolerance. RESULTS: The UH(2)-U ratios were normally distributed (mean value, 2.82; range, 0.35 to 7.13) and were highly correlated to (1) 5-FU plasma levels after the first course of treatment (r =.58), (2) 5-FU plasma clearance (r =.639), and (3) individual optimal therapeutic 5-FU dose (r =.65). Toxic side effects were observed only in patients with initial UH(2)-U ratios of less than 1.8. No adverse effects were noted in patients with UH(2)-U ratios of greater than 2.25. CONCLUSION: The UH(2)-U ratio, easily determined before treatment, could help to identify patients with metabolic deficiency and, therefore, a risk of toxicity.

Determination of unbound platinum after oxaliplatin administration: comparison of currently available methods and influence of various parameters.

Variations in plasma protein binding may have profound effects on both disposition and activity of drugs, especially for those which are tightly bound to proteins, such as anticancer platinum derivatives. Methods of separation of the non-protein-bound fraction and some technical parameters may influence the results. We have compared ultrafiltration and ultracentrifugation, as well as the effect of potentially interfering factors, upon the determination of the plasma unbound platinum fraction after oxaliplatin administration to cancer patients. Ultrafiltration and ultracentrifugation provided very closely correlated results, so that either technique can be used. The ultrafiltration cut-off (3000-30,000 Da) devices, the type of tube for blood sampling and the type of anticoagulant (none, lithium heparinate or EDTA) did not influence the results markedly. In contrast, results were greatly influenced by freezing: erratic results were obtained on thawed plasmas when compared with those on fresh serum or plasma. Consequences may be important in usual practice, since many pharmacokinetic studies are carried out in multicentric trials with plasma processing centralized in one reference laboratory. The methods for the determination of protein-drug binding should be standardized and guidelines elaborated where optimal conditions for each type of binding assay are given.

Long-term weekly treatment of colorectal metastatic cancer with fluorouracil and leucovorin: results of a multicentric prospective trial of fluorouracil dosage optimization by pharmacokinetic monitoring in 152 patients.

PURPOSE: A relationship between fluorouracil (5-FU) dose and response has been previously shown in advanced colorectal cancer. In a previous study with 5-FU stepwise dose escalation in a weekly regimen, and pharmacokinetic monitoring, we defined a therapeutic range for 5-FU plasma levels: 2,000 to 3,000 microg/L (area under the concentration-time curve at 0 to 8 hours [AUC0-8], 16 to 24 mg x h/L). The current study investigated 5-FU therapeutic intensification with individual dose adjustment in a multicentric phase II prospective trial. PATIENTS AND METHODS: Weekly high-dose 5-FU was administered by 8-hour infusion with 400 mg/m2 leucovorin. The initial dose of 5-FU (1,300 mg/m2) was adapted weekly according to 5-FU plasma levels, to reach the therapeutic range previously determined. RESULTS: A total of 152 patients entered the study from December 1991 to December 1994: 117 patients with measurable metastatic disease and 35 with assessable disease. Toxicity was mainly diarrhea (39%, with 5% grade 3) and hand-foot syndrome (30%, with 2% grade 3). Among 117 patients with measurable disease, 18 had a complete response (CR), 48 a partial response (PR), 35 a minor response (MR) and stable disease (SD), and 16 progressive disease (PD). Median overall survival time was 19 months. The 5-FU therapeutic plasma range was rapidly reached with a variable 5-FU dose in the patient population: mean, 1,803 +/- 386 mg/m2/wk (range, 950 to 3,396). Thirteen patients were immediately in the toxic zone, whereas 51 required a > or = 50% dose increase. CONCLUSION: Individual 5-FU dose adjustment with pharmacokinetic monitoring provided a high survival rate and percentage of responses, with good tolerance.