Pharmacogenetic studies of paclitaxel in ovarian cancer – focus on interindividual differences in pharmacodynamics and pharmacokinetics

.................................................................................................... 9 POPULÄRVETENSKAPLIG SAMMANFATTNING............................. 11 PAPERS IN THE PRESENT THESIS ....................................................... 13 ABBREVIATIONS ....................................................................................... 14 INTRODUCTION......................................................................................... 15 INDIVIDUALIZATION OF CANCER CHEMOTHERAPY....................................... 15 GENETIC VARIATIONS IN DRUG METABOLIZING ENZYMES AND TRANSPORTERS ........................................................................................ 17 PHENOTYPING AND GENOTYPING................................................................. 19 PACLITAXEL ................................................................................................. 20 The Clinical Use of Paclitaxel and Ovarian Cancer................................................... 21 Paclitaxel Pharmacokinetics....................................................................................... 24 Metabolism of Paclitaxel............................................................................................ 24 Resistance to Paclitaxel.............................................................................................. 28 Decreased sensitivity to apoptosis-inducing stimuli ......................................................... 29 Tubulin Alterations ........................................................................................................... 29 P-glycoprotein and other drug transporters....................................................................... 30 AIMS OF THE THESIS............................................................................... 37 MATERIAL AND METHODS.................................................................... 39 PATIENTS...................................................................................................... 39 METHODOLOGICAL OVERVIEW .................................................................... 41 Single Strand Conformation Analysis........................................................................ 41 DNA-sequencing........................................................................................................ 42 Solid Phase Extraction ............................................................................................... 44 High Performance Liquid Chromatography............................................................... 45 UV detectors ..................................................................................................................... 46 Fluorescence detectors ...................................................................................................... 47 Mass spectrometry ..................................................................................................... 47 STATISTICS ................................................................................................... 50 RESULTS AND DISCUSSION ................................................................... 51 THE SCARCITY OF β-TUBULIN MUTATIONS IN OVARIAN TUMORS................ 51 ALLELE FREQUENCIES OF SNPS IN ABCB1, CYP2C8 AND CYP3A4 IN A SWEDISH POPULATION AND IN OVARIAN CANCER PATIENTS ................... 52 CORRELATIONS OF ABCB1 GENOTYPES WITH THE RESPONSE TO PACLITAXEL............................................................................................. 54 QUANTIFICATION OF PACLITAXEL AND ITS HYDROXYMETABOLITES IN HUMAN PLASMA ...................................................................................... 59 Extraction of paclitaxel from human plasma ............................................................. 59 Quantification of paclitaxel and metabolites using HPLC......................................... 60 Quantification of paclitaxel and metabolites using LC/MS....................................... 63 CORRELATIONS OF PACLITAXEL PHARMACOKINETICS TO ABCB1, CYP2C8, CYP3A4 GENOTYPES AND CYP3A4 PHENOTYPE ................... 67 CORRELATIONS BETWEEN PACLITAXEL EXPOSURE, RESPONSE AND GENOTYPES OF ABCB1, CYP2C8 AND CYP3A4.................................... 71 CONCLUSIONS............................................................................................ 75 FUTURE ASPECTS ..................................................................................... 77 ACKNOWLEDGEMENTS.......................................................................... 79 REFERENCES .............................................................................................. 82