Determination of irinotecan, SN-38 and SN-38 glucuronide using HPLC/MS/MS: Application in a clinical pharmacokinetic and personalized medicine in colorectal cancer patients.

BACKGROUND: Irinotecan (CPT-11) is chemotherapy used mainly in the metastatic colorectal cancer. The purpose of this study was to develop and validate the LC-MS/MS for the simultaneous determination of CPT-11, SN-38, and SN-38G. METHODS: A 100 µL of plasma was prepared after protein precipitation and analyzed on a C18 column using 0.1% acetic acid in water and 0.1% acetic acid in acetonitrile as mobile phases. The mass spectrometer worked with multiple reaction monitoring (MRM) in positive scan mode. The standard curves were linear on a concentration range of 5-10 000 ng/mL for CPT-11, 5-1000 ng/mL for SN-38, and 8-1000 ng/mL for SN-38G. RESULTS: In this assay, the intra and interday precision consisted of ≤9.11% and ≤11.29% for CPT-11, ≤8.70% and 8.31% for SN-38, and ≤9.90 and 9.64% for SN-38G. CONCLUSION: This method was successfully used to quantify CPT-11, SN-38, and SN-38G and applied to a pharmacokinetic study.

Development of Pyrosequencing Method for Detection of UGT1A1 Polymorphisms in Thai Colorectal Cancers.

BACKGROUND: UGT1A1 is a polymorphic enzyme that has been associated with irinotecan drug metabolisms. We developed a pyrosequencing method to detect allele frequency and genotype of UGT1A1 polymorphisms (UGT1A1*28 and UGT1A1*6) in Thai colorectal cancer patients. METHOD: A pyrosequencing method was designed to determine UGT1A1 genetic polymorphisms including UGT1A1*28 (A[TA]7TAA) and UGT1A1*6 (211G>A) in 91 Thai colorectal cancers. RESULT: Genotyping by the pyrosequencing technique was 100% concordant with capillary electrophoresis sequencing. The allele frequencies for UGT1A1 genetic polymorphisms were *1/*1 (54.95%), *1/*6 (13.19%), *1/*28 (25.27%), *28/*6 (4.40%), and *28/*28 (2.20%). No homozygous mutation UGT1A1*6 was found in our population. CONCLUSIONS: We developed a rapid, reliable, more cost-effective, and simple assay to detect UGT1A1 genetic polymorphisms in routine practice before initiating irinotecan therapy. The UGT1A1*28 and UGT1A1*6 alleles were found to be similar in the Asian populations.

Correlation of UGT1A1(*)28 and (*)6 polymorphisms with irinotecan-induced neutropenia in Thai colorectal cancer patients.

UDP-glucuronosyltransferase1A1 (UGT1A1) polymorphisms have been related with irinotecan toxicity. The purpose of this study was to determine the associations between UGT1A1(*)28 and (*)6 polymorphisms and irinotecan toxicity in Thai patients with metastatic colorectal cancer. 44 metastatic colorectal cancer patients received irinotecan-based chemotherapy. Hematologic toxicities were determined in the first and second cycles of treatment. The genotypes of UGT1A1(*)28 and (*)6 were analyzed by pyrosequencing technique. The frequencies of genetic testing for UGT1A1(*)28 and (*)6 polymorphisms were 22.8% (TA6/TA7; 20.5%, TA7/TA7; 2.3%) and 15.9% (GA), respectively. No patients had the homozygous UGT1A1(*)6 (AA). Neither UGT1A1(*)28 nor UGT1A1(*)6 polymorphisms were significantly associated with severe hematologic toxicities. However, analysis of UGT1A1(*)28 and (*)6 in combination revealed an association with severe neutropenia in the first and second cycles (P = 0.044, P = 0.017, respectively). Both UGT1A1(*)28 and (*)6 polymorphisms may have an increased risk of irinotecan-induced neutropenia in Thai colorectal cancer patients.

Association of CYP2D6 and CYP2C19 polymorphisms and disease-free survival of Thai post-menopausal breast cancer patients who received adjuvant tamoxifen.

PURPOSE: To investigate the impact of CYP2D6 and CYP2C19 polymorphisms in predicting tamoxifen efficacy and clinical outcomes in Thai breast cancer patients. METHODS: Polymorphisms of CYP2D6 and CYP2C19 were genotyped by the AmpliChip™ CYP450 Test (Roche Molecular Diagnostics, Branchburg, NJ, USA) for 57 patients, who were matched as recurrent versus non-recurrent breast cancers (n = 33 versus n = 24, respectively, with a 5-year follow-up). RESULTS: Based on the genotype data, five CYP2D6 predicted phenotype groups were identified in this study including homozygous extensive metabolizer (13 of 57, 22.80%), extensive/intermediate metabolizer (23 of 57, 40.40%), extensive/poor metabolizer (3 of 57, 5.30%), homozygous intermediate metabolizer (14 of 57, 24.50%), and intermediate/poor metabolizer (4 of 57, 7.00%), and three CYP2C19 genotype groups including homozygous extensive metabolizer (27 of 57, 47.40%), extensive/intermediate metabolizer (27 of 57, 47.40%), and homozygous poor metabolizer (3 of 57, 5.30%). The CYP2D6 variant alleles were *10 (52 of 114, 45.60%), *5 (5 of 114, 4.40%), *41 (2 of 114, 1.80%), *4 (1 of 114, 0.90%), and *36 (1 of 114, 0.90%); the CYP2C19 variant alleles were *2 (27 of 114, 23.70%) and *3 (6 of 114, 5.30%). Kaplan-Meier estimates showed significantly shorter disease-free survival in patients with homozygous TT when compared to those with heterozygous CT or homozygous CC at nucleotides 100C>T and 1039C>T (CYP2D6*10) post-menopausal (log-rank test; P = 0.046). They also had increased risk of recurrence, but no statistically significant association was observed (hazard ratio 3.48; 95% confidence interval 0.86-14.07; P = 0.080). CONCLUSION: The CYP2D6 and CYP2C19 polymorphisms were not involved in tamoxifen efficacy. However, in the subgroup of post-menopausal women, the polymorphisms in CYP2D6 and CYP2C19 might be useful in predicting tamoxifen efficacy and clinical outcomes in breast cancer patients receiving adjuvant tamoxifen treatment. As the number of breast cancer patients was relatively small in this study, results should be confirmed in a larger group of prospective patients.