CLINICAL PHAP-dMACOLO(iY & THERAPEUTICS

P 5 8 American Society for Clinical Pharmacology and Therapeutics FEBRUARY2003

PDI-B-2 EFFECT OF ITRACONAZOLE ON THE PHARMACOKINET-

ICS AND PHARMACODYNAMICS OF FEXOFENADINE IN SUBJECTS WITH KNOWN GENOTYPE OF MDRI 3435C>T ALLELE. J. SShon, MD, l. Lee, MS, M. Kim, MS, H. Chun, MS, J. Lee, MS, Y. Yoon, MD, PhD, S. Cha, MD, H. Kim, MD, J. Shin, MD, PhD, Inje University College of Medicine, Busan Paik Hospital, Maritime Medical Center, Korean Navy, Busan, Korea.

The effect of itraconazole on the disposition of fexofenadine, a known P-glycoprotein substrate, was evaluated in 10 subjects whose genotype of MDR1 3435C>T allele was predetermined. All subjects were given single oral dose of 180mg fexofenadine at 1 hour after single oral dose of 200mg itraconazole or placebo as placebo based double blind randomized crossover study. Blood samples were serL ally drawn and urine was collected up to 24 hours. Histamine induced wheal and flare sizes were also measured for 4 hours after fexofena- dine dose to compare the pharmacodynamic effects.

In the phase of placebo pretreatment, the AUC and Cmax of subjects with MDRI 3435 T/T allele tend to be higher than those in subjects with C/C allele. The pretreatment of itraconazole signifi- cantly increased the AUC and Cmax (2.1 - 2.7 fold) and decreased C1/F (2.7 fold) of fexofenadine compared to pretreatment of placebo in subjects with both genotypes as summarized in the following table.

3435C/C 3453T/T

Cmax (ng/ml) Placebo 560.9_ + 173.3 798.0_ + 186.0 Itraconazole 1396.5_+184.1 1919.3_+192.5

AUC (ng'hr/mI) Placebo 3857.0_ + 1059.4 5406.4_+ 1227.2 itraconazole 9145.0_+1215.3 14397.0-+1000.7

CI/F (L/kg/h) Placebo 794.7- + 134.6 475.1 +84.6 Itraconazole 292.3_+25.0 158.7_ + l 1.0

The size of histamine induced skin flare tend to be decreased after itraconazole pretreatment compared to placebo treatment, especially at 1 and 2 hours after fexofenadine dose.

These results suggest that the disposition and antihistamine effect of fexofenadine were markedly influenced by co-administered itra- conazole, a P-gp inhibitor, in addition to the moderate effect of MDR1 3435C>T genotype.

PDI-B-3 ROLE OF P-GLYCOPROTEIN IN THE INTESTINAL AB-

SORPTION OF MORPHINE, FENTANYL AND METHADONE. E. D. K h a ~ MD PhD, D. Whittington, BS, C. J. Hoffer, CCRC, T. G. Altuntas, PhD, P. Sheffels, BS, University of Washington, Seattle, WA.

Background: There is much variability in opioid dose-effect relationships. Morphine, fentanyl & methadone may be P-glycoprotein (P-gp) substrates in vitro or in animals, but the role of P-gp in vivo in humans is unknown. We tested the hypothesis that P-gp influences oral morphine, fentanyl & methadone absorption, using quinidine (Q) as a P-gp inhibitor and in vivo probe.

Methods: Three IRB-approved, randomized, double-blind, placebo-controlled crossover studies were conducted in i 0-12 healthy volunteers. Plasma (LC- or GC-MS) morphine & 3- & 6-glucu- ronides; fentanyl; and methadone & metabolite (EDDP) were mea- sured for 8 hr after oral morphine, fentanyl or methadone, 1 hr after oral Q or placebo.

Results: Q incr. morphine Cmax (2-l.uld) & AUC (50%) without changing Tmax or elimination. M3G & M6G were unchanged; M3G/ morphine & M6G/morphine ratios were decr by 40%. Q decr fentanyl Tmax & increased Cmax and AUC (2.5 to 3-fold) without changing elimination. Q had no effect on methadone Cmax, AUC, or EDDP/ methadone ratio, however Tmax was decreased by 30%.

Discussion: Q incr absorption & apparent bioavailability of mor- phine & fentanyl, and less so methadone. Tho potential Q inhibition of CYP3A first-pass fentanyl or methadone metabolism might ex- plain this, the unchanged EDDP/methadone ratio does not, nor can it explain the morphine results. Rather, results suggest that intestinal P-gp aflects absorption and bioavailability of morphine, fentanyl, and to a lesser extent methadone.

PDI-B-4 MDR-1 C3435T GENE POLYMORPHISM DOES NOT COR-

RELATE WITH P-GLYCOPROTEIN EXPRESSION AND FUNC- TION IN ACUTE MYELOID LEUKEMIA. I .P. van der Heiden M. M. van den Heuvel, MD, PbD, E. Wiemer, PbD, R. Pieters, MD, PhD, J. Lindemans, PhD, J. van den Anker, MD, PhD, R. van Schaik, Ptd), Erasmus MC, Erasmus MC-Sophia Children's Hospital, Eras- mus MC-Sophia Childrens' Hospital, Childrens' National Medical Center, Rotterdam, Netherlands.

Introduction: One of the major reasons for treatment failure in acute myeloid leukemia (AML) is clinical resistance to chemother- apy, which is associated with the expression of multi drug resistance (MDR) proteins. MDR-1 encodes the drug efflux pump P-glycoprotein (P-gp). The MDR-J genetic polymorphism C3435T has shown to be associated with decreased expression and function of P-gp in duodenum.

Purpose: To determine the relation of the M D R - I polymorphism C3435T to P-gp expression and function in blast cells from patients with AML.

Methods: Mononuclear bone marrow cells were isolated and purified (>85% blasts) from 23 AML patients (8 children, 15 adults) prior to chemotherapy. The C3435T polymorphism was detected using a PCR-RFLP assay on genomic DNA, isolated from the mono- nuclear cells. Protein expression was determined by FACS analysis using antibodies UIC2 and MRK16. P-gp function was analysed using a Rhodamin 123 retention assay, with the modifier PSC833.

Results: We identified 6 wild type, 8 heterozygote and 9 hoinozy- gote subjects for the C3435T variant allele. Both protein expression (medians MRK16:2.4 (CC), 2.1 (CT) and 2.0 (TT); medians UIC2: 2.6 (CC), 2.2 (CT) and 2.4 (TT)) and P-gp function (medians: 1.11 (CC), 1.12 (CT), 1.11 (TT) did not differ significantly between groups.

Conclusion: Our data do not support the importance of pharmao cogenetic testing for the MDR-] C3535T variant allele in patients with AML.

PDI-B-5 ALTERED P-GLYCOPROTE1N IMMUNOREACTIV1TY BUT

NORMAL FUNCTION IN HUMAN PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMCS). L. W. Chiun, D. L. Kroetz, PhD, University of California San Francisco, San Francisco, CA.

The M D R I gene encodes P-gp, a xenobiotic efflux pump that plays an important role in multi-drug resistance and in drug absorption and distribution. Recent analysis of P-gp expression and function in the natural killer (NK) cell subpopulation of human PBMCs identified only 70 and 80 kDa immunoreactive proteins that did not transport two established P-gp substrates, calcein acetoxymethyl ester (calcein AM) or daunorubicin. The goal of our study was to test the hypoth- esis that human NK ceils express a truncated P-gp with altered /'unction. Flow cytometry, Western blot analysis, and reverse tran- scription polymerase chain reaction (RT-PCR) were used to charac- terize MDRt and P-gp in PBMCs. The P-gp expressed in CD56 + PBMCs transported daunorubicin and calcein AM, and this transport was significantly inhibited by verapamil and/or GF120918. RT-PCR of total RNA from human PBMCs gave a 4 kB product consistent with the well characterized MDR1 transcript in other tissues. This PCR product was sequenced at both ends and there was complete identity to MDR1. However, Western blots stained with the C494 antibody detected only 80 kDa immunoreactive proteins and the C219 antibody gave no signal. Cell surface expression of P-gp in PBMCs can be detected with both the UIC2 and MRK-16 antibodies. Further characterization of P-gp in PBMCs is important since func- tional P-gp activity in these cells is commonly used to reflect sys- temic P-gp activity. Supported by NIGMS 61390.