Genotyping of a family with a novel deleterious DPYD mutation supports the pretherapeutic screening of DPD deficiency with dihydrouracil/uracil ratio.

Despite the growing evidence that dihydropyrimidine dehydrogenase deficiency (DPD, encoded by the DPYD gene) confers a higher risk of developing severe toxicity, most patients are not screened for DPD deficiency before fluoropyrimidine treatment. We report here the genetic and phenotypic analyses of DPD in a family related to a patient who died after a first cycle of 5-fluorouracil and in 15 additional retrospective patients having a partial DPD deficiency (as measured by plasma dihydrouracil/uracil ratio). The patient with lethal toxicity was found to be a compound heterozygote for two DPYD mutations: a novel 8-bp duplication (c.168_175dupGAATAATT, p.Phe59Ter) and c.1679T>G (Ile560Ser). The patient's dihydrouracil/uracil ratio indicates complete DPD deficiency. The novel mutation was found in two members of the patient's family. Deleterious DPYD mutations were identified in 9 out of the 15 patients. The relationship between genotype and dihydrouracil/uracil values in the 22 patients of the present study was significant (P = 0.01).

Pharmacokinetics of heated intraperitoneal oxaliplatin.

OBJECTIVE: To evaluate the pharmacokinetic inter-patient variability of 30-min hyperthermic intraperitoneal oxaliplatin chemotherapy. PATIENTS AND METHODS: Data were obtained from 24 patients who were treated according to two procedures of heated intra-operative intraperitoneal oxaliplatin. For the first procedure (12 patients), the solution instilled within the peritoneal cavity contained oxaliplatin, and a delay of 8-10 min was necessary to reach a temperature of 42-43 degrees C. For the second procedure (12 patients), the cavity was initially filled only with the dextrose solution, and oxaliplatin was added to the peritoneal instillate when temperature reached 42-43 degrees C. Plasma and peritoneal fluid oxaliplatin concentrations were analyzed according to a population pharmacokinetic approach using NONMEM. RESULTS: Peritoneal and total plasma data were simultaneously analyzed according to a three-compartment pharmacokinetic model. The peritoneal half-life ranged between 18 and 42 min. The mean peritoneal clearance was 5.47 L/h (+/-21%), and the mean plasma clearance was 3.71 L/h (+/-47%). The heated intra-operative procedure did not have any impact on oxaliplatin pharmacokinetics. CONCLUSION: The inter-individual variability was larger for plasma pharmacokinetic parameters than that for peritoneal parameters. However, the percentage of oxaliplatin dose absorbed during a 30-min hyperthermic intraperitoneal chemotherapy may vary from 40 to 68%. The present pharmacokinetic model will be useful to implement pharmacokinetic evaluation of further clinical trials of hyperthermic intraperitoneal chemotherapy based on platinum compounds' administration.

Dexamethasone as a probe for CYP3A4 metabolism: evidence of gender effect.

BACKGROUND: A study was conducted to evaluate prospectively the correlation between docetaxel clearance and pharmacokinetics of dexamethasone previously obtained in 21 patients. PATIENTS AND METHODS: Dexamethasone pharmacokinetics were performed in 17 patients 24 h before docetaxel treatment as monochemotherapy. Dexamethasone and docetaxel plasma concentrations were determined by HPLC methods. Determination of docetaxel unbound fraction in plasma was performed using microequilibrium dialysis. RESULTS: Significant correlation was observed between observed plasma docetaxel clearances (CL(docetaxel)) and values predicted from dexamethasone plasma clearance (CL(dexa)), unbound plasma docetaxel fraction estimated from serum alpha1-acid glycoprotein level (fu(alpha1-AAG)), and hepatic metastasis status. However, after splitting of the prospective data set according to gender, no correlation was observed for males (R(2) = 0.08, NS, n = 10), then strong correlation was observed for females (R(2) = 0.78, P < 0.01, n = 7). Multivariate analysis was performed from data obtained in the women included in the first study and those of this prospective study (n = 18). Docetaxel CL was significantly correlated with CL(dexa) (P = 0.001) and fu(alpha1-AAG) (P = 0.01) according to the relationship (with +/-95% confidence intervals): CL(docetaxel) (l/h) = 1.92 (+/-0.94) x CL(dexa) (l/h) + 2.68 (+/-1.95) x fu(alpha1-AAG) (%) (R(2) = 0.68). CONCLUSION: Dexamethasone may be used to predict docetaxel clearances in females, but not in males.

A phase I clinical and pharmacokinetic study of capecitabine (Xeloda) and irinotecan combination therapy (XELIRI) in patients with metastatic gastrointestinal tumours.

Capecitabine is a highly active oral fluoropyrimidine that is an attractive alternative to 5-fluorouracil in colorectal cancer treatment. The current study, undertaken in 27 patients with gastrointestinal tumours, aimed to assess the toxicity and potential for significant pharmacokinetic interactions of a combination regimen incorporating capecitabine with 3-weekly irinotecan (XELIRI). Irinotecan (200 and 250 mg m(-2)) was administered as a 90-min infusion on day 1 in combination with escalating capecitabine doses (700-1250 mg m(-2) twice daily) administered on days 2-15 of a 3-week treatment cycle. Pharmacokinetics were characterised on days 1 and 2 of the first two cycles. A total of 103 treatment cycles were administered. The principal dose-limiting toxicities were diarrhoea and neutropenia. Capecitabine 1150 mg m(-2) twice daily with irinotecan 250 mg m(-2) was identified as the maximum-tolerated dose and capecitabine 1000 mg m(-2) with irinotecan 250 mg m(-2) was identified as the recommended dose for further study. Analyses confirmed that there were no significant pharmacokinetic interactions between the two agents. The combination was clinically active, with complete and partial responses achieved in heavily pretreated patients. This study indicates that XELIRI is a potentially feasible and clinically active regimen in patients with advanced gastrointestinal cancer.

Impact of the biochemical assay for serum creatinine measurement on the individual carboplatin dosing: a prospective study.

We previously developed a formula to estimate the individual carboplatin clearance (CL) based on serum creatinine (Scr) determined by an enzymatic assay using creatinine amidohydrolase. An analytical comparison had shown systematic differences between this method and the commonly used Jaffé method (with Jaffé Scr (in microM)=1.08 x enzymatic Scr+1.6, as regression equation). We performed a pharmacokinetic prospective clinical study using the Jaffé assay to evaluate the impact of the method used for Scr measurement on the prediction of the carboplatin CL. In forty patients, carboplatin dosing was performed according to the Chatelut formula where the serum creatinine level was corrected according to the above equation. The population pharmacokinetics of carboplatin were analysed using the NONMEM program to determine the individual carboplatin CL from a limited sampling strategy. Thanks to the correction of the Jaffé Scr, no significant difference was observed between the administered and the optimal dose. In contrast, if no correction of the Scr was done, the patients would have been significantly under-dosed. Moreover, a covariate analysis using NONMEM gave a very consistent result showing that Scr should be decreased by 11.6% when the Jaffé value is used within the Chatelut equation. This study confirmed that differences in the Scr assay has consequences with regard to carboplatin dosing. The correction we propose for Scr obtained by the Jaffé method may help to standardise clinical practice.

Dihydropyrimidine dehydrogenase activity in normal, inflammatory and tumour tissues of colon and liver in humans.

BACKGROUND/PURPOSE: Dihydropyridmidine dehydrogenase (DPD) is the initial and rate-limiting enzyme in the catabolism of 5-fluorouracil (5FU). Although this catabolism is likely to occur in the liver in humans, there may be a local inactivation in tumours, modifying the efficacy of 5FU. The aim of this study was to examine the DPD activity in normal, inflammatory and malignant tissues from both the colon and the liver to assess the modifications of DPD activity in the process of tumourigenesis. METHODS: DPD activity was evaluated in 107 patients, corresponding to 194 samples (70 colorectal tumour and normal colon, nine metastases secondary to a colon cancer, ten inflammatory colon, 20 samples of normal liver, seven from primary liver cancer, and eight from inflammatory liver). DPD activity was determined using an enzymatic reaction followed by analysis of 5FU and its catabolite dihydro-5FU by high-performance liquid chromatograph. Results were expressed as pmol of 5FU catabolized/min x mg protein. RESULTS: DPD was highly variable in tumour and normal tissues, both from colon and liver. In colon, the correlation between DPD activity in tumour and normal mucosa was weak, even if it was statistically significant due to the higher number of samples. In inflammatory colon tissue (ulcerative colitis or Crohn's disease), DPD activity was significantly higher than in normal tissue (P = 0.006). In liver metastases from colon cancer, DPD activity was not significantly different from that observed in primary colon tumour (P = 0.32). In liver, DPD activity was significantly lower in primary liver tumour than in uninvolved liver specimens (P = 0.001). In inflammatory liver tissue (hepatitis), DPD activity ranged between normal and tumour tissues, and did not differ significantly either from normal tissue or primary liver cancer. CONCLUSIONS: DPD activity was modified in colon and in liver during a pathological process and the dysregulation of DPD increased from a benign to a malignant tissue.

CPT-11 converting carboxylesterase and topoisomerase activities in tumour and normal colon and liver tissues.

CPT-11 is a prodrug activated by carboxylesterases to the active metabolite SN-38 which is a potent inhibitor of topoisomerase I. CPT-11 is of clinical interest in the treatment of colorectal cancer. We evaluated the activities of CPT-11 converting carboxylesterase (CPT-CE) and topoisomerase I (topo I) in 53 colorectal tumours, in eight liver metastases and in normal tissue adjacent to the tumours. Both CPT-CE and topo I activities were widely variable in the malignant and the normal tissue of patients with colorectal carcinomas. CPT-CE was only two to threefold lower in primary tumours compared to normal liver, suggesting that a local conversion to SN-38 might occur in tumour cells. CPT-CE was similar in liver and in normal colon tissues. Levels of topo I in tumour ranged from 580 to 84 900 U mg protein(-1) and was above 40 000 U mg protein(-1) in 11 of 53 patients. Similarly, a very high ratio (> 5) between tumour and normal tissues were observed in 12 of 53 patients. An inverse correlation was observed between the topo I activity and the clinical stage of disease. Clinical studies are in progress in our institution to explore a possible relationship between CPT-CE and topo I activities in tumour cells and the response to CPT-11-based chemotherapy in patients with colorectal cancer.

A limited-sampling strategy for estimation of etoposide pharmacokinetics in cancer patients.

Etoposide (VP16), a widely used anticancer drug, is a topoisomerase II inhibitor. A number of studies have highlighted a correlation between hematologic toxicity and pharmacokinetic or physiological parameters. Other studies have also suggested that the anti-tumor response could be related to the plasma etoposide concentration. Therefore, it would seem of interest to individualize VP16 dose regimens on the basis of pharmacokinetic parameters. The aim of this study was to develop and validate a limited-sampling strategy allowing VP16 pharmacokinetic evaluation with minimal disturbance to the patient. A total of 34 patients (54 kinetics) received VP16 at various dose regimens, with doses ranging between 30 and 200 mg and infusion times varying between 0.5 and 2 h. The statistical characteristics of the pharmacokinetic parameters were assessed from the first courses of treatment performed in 23/34 patients; then the following three-sample protocol was designed: the end of the infusion and 5 and 24 h after the start of the infusion. For validation of the model the main pharmacokinetic parameters (clearance, half-lives, volume of distribution) were estimated in the 11 remaining patients by maximum-likelihood estimation (ML) and by Bayesian estimation (BE) using the three sampling times designed. Statistical comparison showed a good concordance between ML and BE estimates (the bias for clearance was -1.72%). The limited-sampling strategy presented herein can thus be used for accurate estimation of VP16 pharmacokinetic parameters.

Comparison of the pharmacokinetics and efficacy of irinotecan after administration by the intravenous versus intraperitoneal route in mice.

Irinotecan (CPT-11) is a new drug active in colorectal cancer. A comparison was made of the efficacy and pharmacokinetics of CPT-11 after i.p. versus i.v. administration to mice. We found that i.p. administration of CPT-11 to mice bearing C26 colon cancer was more efficient and less toxic than i.v. administration; a 100-mg i.p. dose induced an increase in life span equivalent to that produced by a 300-mg i.v. dose, and toxic deaths appeared after doses of 400 mg/kg given i.v. and 800 mg/kg given i.p. Pharmacokinetic parameters of CPT-11 and SN-38 were compared after i.v. or i.p. administration in mice bearing P388 leukemia ascites. Peritoneal CPT-11 and SN-38 AUC values were higher after i.p. administration than after i.v. injection. Plasmatic AUC values remained equivalent. Moreover, peritoneal CPT-11 clearance was 10-fold lower after i.p. versus i.v. administration. If the survival and pharmacologic advantage of i.p. CPT-11 in the murine model considered can be translated to a safe and practical mode of therapy in patients and if local toxicity does not prove to be a major adverse effect, then a potentially useful agent could be added to the drugs known to be active when given by the i.p. route for adjuvant therapy in colon cancer.

Cytotoxic effect of interferon-alpha2a in combination with all-trans retinoic acid or cisplatin in human ovarian carcinoma cell lines.

Ovarian cancer has a poor prognosis due to the frequent appearance of a drug-resistant state. An alternative therapeutic approach may lie in combinations of conventional chemotherapeutic agents with new classes of drug, such as interferons (IFN) and differentiation-inducing agents. There is clinical evidence that both IFN-alpha2a-all-trans retinoic acid (ATRA) and IFN-alpha2a-cisplatin have significant activities on growth of malignant cells, cell differentiation or programmed cell death in solid tumors. In order to throw more light on the cellular basis of these findings and to optimize a schedule of such drug combinations, we examined the cytotoxic effects of various combinations on five human ovarian carcinoma cell lines. The experiments were based on a clonogenic assay on plastic. The different cell lines exhibited different sensitivities to the three drugs tested. Using the cell line most sensitive to these drugs, we then examined the effect of different sequences of two drug combinations. We observed a potentiation after pretreatment with ATRA followed by IFN-alpha2a and ATRA or after pretreatment with IFN-alpha2a followed by IFN-alpha2a and cisplatin. Using this schedule of administration, cytotoxic interactions between the two drugs were investigated by median effect analysis. Synergism or antagonism were observed depending on the intrinsic sensitivity of the cell line to the first drug and the concentrations used. The magnitude of these interactions was found to be influenced by the cellular sensitivity to the second drug. These results show that schedules of drug combinations are not easy to design and may help account for the various failures and the discrepant effects observed in clinical trials.

Population pharmacokinetics of total and unbound etoposide.

A population pharmacokinetics study using the NONMEM program was undertaken to determine the effects of different covariates on the pharmacokinetic parameters of etoposide. A total of 1,044 plasma etoposide concentrations were determined by high-performance liquid chromatography (HPLC) in 100 patients (pts; 75 men and 25 women aged 25-85 years) treated for various tumor types with i.v. (57 pts) or oral (43 pts) etoposide. For 67 pts, etoposide plasma protein binding was determined by equilibrium dialysis; the unbound fraction ranged from 4% to 24%. A linear two-compartment model with first-order absorption (for oral dosing) accurately described the concentration versus time data. The central and peripheral volumes of distribution were significantly correlated with the body surface area [Vc (L) = 5.5 x BSA (m2) and Vp = 4.1 x BSA], but even after BSA had been taken into account, the interindividual variability of the two volumes remained high (34% and 57%, respectively). The clearance (CL) was not correlated with the following covariates: age, BSA, sex, height, and levels of serum bilirubin and liver enzymes. The final regression model for CL was CL (ml/min) = 49.8 x (1 - 0.009 x PRO) x WT/Scr + 33.8 x (1 - 0.29 x META) x (1 - 0.012 x ALB), where ALB, PRO, WT, and Scr, respectively, were albuminemia, proteinemia (g/l), weight (kg), and serum creatinine (microM) and META = 1 if the patient had liver metastases (otherwise, META = 0). The interindividual variability in CL (mean value 30 ml/min) decreased only from 32% to 26% when these covariates were taken into account. The mean oral bioavailability was 66%, showing an interindividual variability of 37%. The plasma clearance of the unbound fraction was strongly and negatively correlated with Scr but was not dependent on either PRO or ALB. These data show that modifications in PRO levels do not directly affect plasma exposure to unbound etoposide. This analysis makes possible the rational consideration of modifications of covariates such as Scr in etoposide dosing. This population data base will constitute the prerequisite for adaptative control with feedback dosing for continuous oral administration of etoposide.

Relation between unbound plasma concentrations and toxicity in a prolonged oral etoposide schedule.

OBJECTIVE: This study was undertaken in order to evaluate the impact of pharmacokinetics on the toxicity of oral etoposide administered daily for 21 days. METHODS: The daily dose was 50 mg/m2. Thirty-two patients 24 males and eight females, 36 76 years old, treated for various tumour types), were evaluated. Blood samples were obtained on day 1 for all patients, and on day 21 for 16 patients. Plasma etoposide concentrations were determined by high-performance liquid chromatography, and etoposide plasma protein binding by equilibrium dialysis. RESULTS: On day 1, the mean value (with coefficient of variation for interindividual variability) for the unbound fraction (fu), area under the concentration versus time curve (AUC), and unbound AUC was 9.8% (59%), 34 mg x h/l (39%), and 3.5 mg x h/l (92%), respectively. The ratio between AUC on day 1 and day 21 ranged between 0.5 and 1.8 (mean 0.9, with CV 33%). The plasma trough unbound concentrations and the unbound AUCs both corresponding to the first administration were significantly higher in the 11 patients who had a severe neutropenia than in the 21 patients who had no or moderate toxicity. However, total etoposide concentrations did not differ between these two groups. A limited sampling strategy using the NONMEM program and a database of 89 patients previously studied was performed. The optimal sampling schedule (i.e. 1, 4, and 24 h after oral etoposide administration) allowed to obtain the AUC accurately on day 1. CONCLUSION: Individual adjustment of oral etoposide based on unbound pharmacokinetics after the first administration appears relevant and feasible.

[Bayesian estimation of pharmacokinetic parameters of etoposide]. / Estimation bayésienne des paramètres pharmacocinétiques de l'étoposide.

Studies of the relationships between the pharmacokinetics of a drug and its pharmacodynamics could significantly improve chemotherapy efficacy. However, despite their proven value, pharmacokinetic studies sometimes appear as cumbersome and difficult procedures. The bayesian approach associated with an optimal sampling time strategy (OST) allows the determination of the pharmacokinetic parameters of a drug with a smaller number of blood samples compared with that required by the classic maximum likelihood estimation (MLE). Therefore, the bayesian approach may lead to a less discomfort to the patients and less work for the medical staff. Such a method was developed to determine the individual pharmacokinetic parameters of etoposide (VP16). First, the statistical characteristics of the pharmacokinetic parameters were evaluated in 14 courses from 14 patients. Then, based on these results, a three-sample strategy was developed. Validation of this methodology was performed in 7 new patients and evaluated by computing bias and precision. The performance of the developed methodology shows that it could successfully be applied for the determination of VP16 pharmacokinetic parameters.

Multifactorial mechanism for the potentiation of cisplatin (CDDP) cytotoxicity by all-trans retinoic acid (ATRA) in human ovarian carcinoma cell lines.

All-trans retinoic acid (ATRA) has been previously shown to inhibit the proliferation of some human ovarian carcinoma cell lines, and this inhibition was accompanied by cellular changes that were indicative of differentiation (Caliaro et al, 1994). In this work, a pretreatment of these adenocarcinoma cells with ATRA, for their respective doubling time, enhanced cisplatin (CDDP) cytotoxicity in the cell ines that were sensitive to its antiproliferative effect, but not in the ATRA-resistant ones. Results were assessed using median effect analysis in two ATRA-sensitive cell lines (OVCCR1 and NIHOVCAR3 cells) and in one ATRA-insensitive cell line (IGROV1 cells). Synergy between these two agents was observed only in cells sensitive to ATRA, regardless of their relative sensitivity to CDDP. Potential mechanisms for this synergy were investigated. ATRA did not increase the cellular platinum content, did not decrease the cellular glutathione and had no influence on the metallothionein IIA mRNA levels in NIHOVCAR3 cells. Moreover, the protein kinase C (PKC) activity was modulated by this differentiating agent in all cell lines tested, indicating that this activity was not directly involved in this potentiation. However, an ATRA inhibition of glutathione-S-transferase activity associated with an increase in the total DNA adducts formation could explain the potentiation of the CDDP cytotoxicity observed in NIHOVCAR3 cells. Finally, the ATRA modulation of the epidermal growth factor (EGF) receptor mRNA level could also be implicated in this synergy.

[Relations between hematological toxicity and total and free plasma levels of etoposide in daily oral administration]. / Relation entre la toxicité hématologique et les concentrations plasmatiques totales et libres d'étoposide administré quotidiennement par voie orale.

This study was undertaken in order to evaluate the inter- and intra-individual pharmacokinetic variabilities and their impact on the toxicity of oral Vépéside Sandoz administered daily for 21 days. The pharmacokinetic results confirmed the low bioavailability of this formulation (14% +/- 10%) and its large interindividual variability. Moreover, a great intraindividual variability was shown between day 1 and day 21. This fact can explain that the relationship between the relative decrease in neutrophil count and the pharmacokinetic parameters was observed only with either the mean area under the curve of concentrations versus time (AUC) or the mean residual concentrations (Cr). The determination of the fraction of plasma etoposide unbound, which ranged from 4.6 to 24.8%, improved the pharmacokinetic-pharmacodynamy relationship for AUC but not for Cr. This study showed the potential interest of etoposide drug monitoring. However, no dosage adjustment could be performed with this oral etoposide formulation because of its large intraindividual bioavailability. Since this study was performed, the Sandoz company withdrew this formulation, and replaced it by one identical with that available in other countries.