Association of plasma docetaxel levels with ABCB1 gene polymorphisms and tumour response in locally advanced breast cancer patients of South India on neo-adjuvant chemotherapy.

BACKGROUND: Genetic factors could be attributed to the variability in docetaxel plasma levels and its subsequent therapeutic response. The objectives of this study were to assess the effect of ABCB1 gene polymorphisms [SNPs rs1045642 (C3435T) and rs1128503 (C1236T)] on docetaxel plasma levels and also to analyze the influence of docetaxel plasma levels on tumour response in the ethnically distinct South Indian population. METHODS: 104 locally advanced breast cancer (LABC) patients on docetaxel-based neo-adjuvant chemotherapy (NACT) were included. The plasma docetaxel levels were estimated using the validated reverse phase liquid chromatography with mass spectrometry (LC-MS/MS). DNA was extracted (phenol-chloroform extraction method) and the real-time PCR system using validated TaqMan® SNP genotyping assay method was used for genotyping. Tumour response was assessed by RECIST criteria based on the MRI images. RESULTS: Patients with "CT/TT" genotype of the SNP C1236T had a C0/Ct ratio of 1.6 times higher than those with "CC" genotype (13.5 ± 6.5 vs 8.3 ± 3.1, p = 0.002). Though not significant, patients with "CT/TT" genotype had greater initial plasma concentration (C0) and area under the plasma concentration-time curve (AUC0-t). Conversely, the SNP C3435T was not associated with the plasma docetaxel levels. Furthermore, the C0 and normalized C0 were found to be higher in tumour responders compared to non-responders (p < 0.05). CONCLUSIONS: The plasma levels of docetaxel were significantly influenced by the SNP C1236T of ABCB1 gene coding for the MDR1 transporter (P-glycoprotein). The plasma levels of docetaxel were also found to influence its therapeutic effect.

Genetic influence of DPYD*9A polymorphism on plasma levels of 5-fluorouracil and subsequent toxicity after oral administration of capecitabine in colorectal cancer patients of South Indian origin.

OBJECTIVES: High interindividual variability was reported with capecitabine toxicities among colorectal cancer (CRC) patients. DPYD*9A polymorphism was reported responsible for grade 3 or 4 toxicities. Finding the phenotypic association between DPYD*9A polymorphism and 5-fluorouracil (5-FU) plasma levels will give a better prediction for toxicity susceptibility. METHODS: A total of 145 CRC patients were included in the final analysis. Each patient received capecitabine of 1,000 mg/m2 twice daily for the first 14 days of a 21 day cycle. 5-FU levels were measured at two-time points 2 and 3 h post capecitabine administration across the 1st and 4th cycles of chemotherapy. 5-FU levels were measured using liquid chromatography and tandem mass spectrometry (LC-MS/MS). Genotyping analysis was done by real-time PCR (RT-PCR). RESULTS: The mean 5-FU drug levels measured during the 1st cycle at time points 2 and 3 h were found to be 267 ng/mL ± (29) and 124 ng/mL ± (22) respectively. Whereas, the observed 5-FU levels in the 4th cycle were 275 ng/mL ± (28) and 130 ng/mL ± (26) respectively. Patients with 5-FU levels in the range of 281-320 and 141-160 ng/mL at 2 and 3 h respectively showed a higher risk for the hand-foot syndrome (HFS) and thrombocytopenia. No association was found between DPYD*9A polymorphism and 5-FU drug levels measured at time point 2 h across both the cycles. However, the drug levels measured at 3 h were found to be significantly different across the DPYD*9A genotypes. Individuals with GG genotype showed significantly higher 5-FU levels when compared to AA genotype. CONCLUSIONS: DPYD*9A polymorphism had a significant influence on the plasma levels of 5-FU after capecitabine administration. The 5-FU levels measured at 3 h corresponding to elimination t1/2 was significantly higher in patients with GG genotype compared AA genotype.

Association of NUDT15*3 and FPGS 2572C>T Variants with the Risk of Early Hematologic Toxicity During 6-MP and Low-Dose Methotrexate-Based Maintenance Therapy in Indian Patients with Acute Lymphoblastic Leukemia.

Genetic variants influencing the pharmacokinetics and/or pharmacodynamics of the chemotherapeutic drugs used in Acute Lymphoblastic Leukemia (ALL) therapy often contribute to the occurrence of treatment related toxicity (TRT). In this study, we explored the association of candidate genetic variants with early hematological TRT (grade 3-4) occurring within the first 100 days of low-dose methotrexate and 6-mercaptopurine based maintenance therapy (n = 73). Fourteen variants in the following candidate genes were genotyped using allele discrimination assay by real-time PCR: ABCB1, DHFR, GGH, FPGS, MTHFR, RFC1, SLCO1B1, TPMT, and NUDT15. Methotrexate polyglutamate (MTXPG3-5) levels in red blood cells were measured by LC-MS/MS. Early hematological TRT (grade 3-4) was seen in 54.9% of patients. The NUDT15*3 allele was associated with early TRT occurrence [HR: 3.04 (95% CI: 1.5-6.1); p = 0.007]. Sensitivity of early TRT prediction improved (from 30.7% to 89.7%) by considering FPGS variant (rs1544105) carrier status along with NUDT15*3 allele [HR = 2.7 (1.5-4.7, p = 0.008)]. None of the considered genetic variants were associated with MTXPG3-5 levels, which in turn were not associated with early TRT. NUDT15*3 allele carrier status could be used as a stratifying marker for Indian ALL patients to distinguish patients at high or low risk of developing early hematological TRT.

Folyl polyglutamate synthethase (FPGS) gene polymorphisms may influence methotrexate adverse events in South Indian Tamil Rheumatoid Arthritis patients.

The aim of the study was to look for the association of FPGS 2752 G > A (rs1544105), FPGS 1994 A > G (rs10106), and GGH 452 C > T (rs 11545078), GGH -401C > T (rs 3758149) gene polymorphisms with methotrexate (MTX) treatment response and MTX-induced adverse events in South Indian Tamil patients with rheumatoid arthritis (RA). Further the influence of these gene polymorphisms on MTX polyglutamate levels was analyzed. A total of 330 patients with RA were investigated. FPGS gene polymorphisms were analyzed by TaqMan 5'nuclease assay and GGH gene polymorphisms were analyzed by PCR-RFLP. Methotrexate polyglutamates (nmol/L of packed erythrocytes) were measured by liquid chromatography mass spectrometry (LCMS/MS) method. We found that the heterozygous genotype of FPGS rs1544105 [p = 0.02, OR 1.93, 95% CI (1.15-3.35)] and FPGS rs10106 AG genotype [p = 0.01, OR 2.11, 95% CI (1.20-3.71)] were associated with MTX adverse events. FPGS rs1544105 and GGH -401C > T SNPs influenced the polyglutamate levels. None of the investigated SNPs seems to be associated with MTX treatment outcome.

Effect of antituberculosis treatment on CYP2C19 enzyme activity in genetically polymorphic South Indian Tamilian population.

Patients on antituberculosis therapy (ATT) are more prone to drug interactions in the presence of coexisting illnesses which require drug therapy. Rifampicin is a pleiotropic inducer of CYP enzymes, and isoniazid is an enzyme inhibitor. Genetic variations are common in the gene coding for CYP2C19 enzyme. These variations would be important in predicting the individual variations in CYP2C19 activity. The objectives of the study were to find the net effect of 1-month ATT on CYP2C19 enzyme activity and its association with CYP2C19 genetic polymorphisms. Newly diagnosed tuberculosis patients (n = 125) were included in the study. Before commencing ATT, they were given a single dose of omeprazole 20 mg as a probe drug for CYP2C19. Blood sample was collected after 3 h to carry out phenotyping for CYP2C19 enzyme by measuring omeprazole hydroxylation index (OHI) using LC-MS/MS. The phenotyping procedure was repeated after 1 month of ATT. CYP2C19 genotyping was carried out by PCR-RFLP method. Significant reduction in OHI was observed after 1 month of ATT in all the metabolizer groups. The percentage reduction in OHI was maximum with poor metabolizers, 84.1 (IQR - 74.6, 86.6), and minimum with ultra-rapid metabolizers, 39.6 (IQR - 12.7, 54.7). CYP2C19 enzyme induction is predominant in patients after 1 month of antituberculosis treatment (ATT). Genetic variations in the enzyme could not clearly explain the interindividual differences in induction. There is a potential risk of drug failure/adverse effect in poor metabolizers regardless of their genotype after ATT.

Evaluation of interleukin-6 and serotonin as biomarkers to predict response to fluoxetine.

OBJECTIVE: Only 30% of major depressive disorder (MDD) patients achieve complete remission with a serotonergic antidepressant (selective serotonin reuptake inhibitor). We investigated the potential of serotonin (5-HT) and interleukin-6 (IL-6) to serve as functional biomarkers of fluoxetine response. METHODS: Serum IL-6 and 5-HT were measured in 73 MDD patients (39 responders and 34 non-responders) pre- and 6 weeks post-treatment and in 44 normal controls with ELISA. Fluoxetine and norfluoxetine were measured using LC MS/MS. RESULTS: IL-6 levels were significantly higher in MDD patients when compared with controls (p < 0.01), and 5-HT levels were significantly lower in non-responders compared with controls (p = 0.0131). Pre- and post-treatment levels of both biomarkers individually and in combination did not significantly differ between responders and non-responders. Area under the receiver operating characteristics curve for the biomarkers was 0.5. Significant correlation was seen between the percentage change in IL-6 and percentage change in Hamilton Rating Scale for Depression score in responders. Fluoxetine and norfluoxetine concentrations were not significantly different in responders and non-responders, and there was no correlation between fluoxetine concentrations and percentage reduction in 5-HT from week 0 to 6. CONCLUSION: 5-HT and IL-6 may not serve as useful markers of response to fluoxetine because of inconsistent results across different studies. Copyright © 2016 John Wiley & Sons, Ltd.

A study to explore the correlation of ABCB1, ABCG2, OCT1 genetic polymorphisms and trough level concentration with imatinib mesylate-induced thrombocytopenia in chronic myeloid leukemia patients.

PURPOSE: Imatinib mesylate is presently the first-line treatment for chronic myeloid leukemia (CML). Therapeutic drug monitoring (TDM) and pharmacogenetic screening is warranted for better management of imatinib therapy. The present study was framed to explore the influence of common drug transporter gene polymorphisms of ABCB1, ABCG2, OCT1 and trough level concentration on commonly occurring adverse events in CML patients treated with imatinib mesylate. METHODS: A total number of 111 patients in chronic phase (Philadelphia chromosome +ve) were included in the study. The plasma drug concentration of imatinib was estimated using LC-MS/MS method. RESULTS: The mean ± SD trough level concentration of imatinib mesylate was found to be 1430.7 ± 438.7 ng/ml. The trough level concentration at steady state (Cmin.ss) was significantly higher in patients with grade 2-4 thrombocytopenia compared with patients without the adverse event (P value 0.033). CONCLUSION: The drug level of imatinib in plasma correlates with the severity of thrombocytopenia, which adds to the utility of TDM in the management of CML patients.

Influence of Sokal, Hasford, EUTOS scores and pharmacogenetic factors on the complete cytogenetic response at 1 year in chronic myeloid leukemia patients treated with imatinib.

Imatinib mesylate is currently considered the first-line treatment for chronic myeloid leukemia (CML). Sokal, Hasford and EUTOS are the three major risk categorization scores available for CML patients. The present study aimed to explore the influence of three risk score, genetic polymorphisms of ABCB1, OCT1, ABCG2 and trough level concentration on complete cytogenetic response at 1 year and overall survival. The mean time period of follow-up was 53.05 months, and the overall survival was 94.6%. The Sokal score (P 0.014), Hasford score (P 0.016) and MDR1 C3435T (P 0.001) tend to influence the overall survival in the patients. The patients who had better overall survival had early complete cytogenetic response (P 0.0003). The ABCG2 C421A was the covariate which had correlation with the complete cytogenetic response. A perceptive approach incorporating pharmacogenetic evaluation with major risk categorization score at the initial stage will help in ensuring better treatment success in CML patients treated with imatinib.

Relative Copy Number Variations of CYP2C19 in South Indian Population.

CYP2C19 is a polymorphic enzyme involved in the metabolism of clinically important drugs. Genotype-phenotype association studies of CYP2C19 have reported wide ranges in the metabolic ratios of its substrates. These discrepancies could be attributed to the variations in the promoter region and this aspect has been reported recently. The observations in the recent reports on the influence of promoter region variants on the metabolism of CYP2C19 substrates might also have been influenced by the copy number variations of CYP2C19. In this paper, we describe copy number variations of CYP2C19 using real-time polymerase chain reaction by comparative Ct method. No copy number variations were observed in the south Indian population indicating the observed discrepancies in genotype-phenotype association studies might be due to the regulatory region polymorphisms as reported earlier.

Genetic variations and haplotypes of the 5' regulatory region of CYP2C19 in South Indian population.

CYP2C19 is expressed polymorphically with about 21 variant alleles. Genotype-phenotype association studies of CYP2C19 have shown marked deviations, suggesting the presence of other variations in the intronic and 5' regulatory region affecting its expression. This study aims to identify the genetic polymorphisms and construction of haplotypes of variations in 5' regulatory region of CYP2C19 among the South Indian population. CYP2C19 5' regulatory region was amplified and sequenced from the DNA of 58 healthy volunteers of South Indian origin. Genetic analysis revealed the existence of 14 variations including eight novel ones in the 5' regulatory region. Identified novel variations and their percentage frequencies were: -779A>C (16.4), -828T>A (2.6), -934del>T (3.5), -1051T>C (1.72), -1289T>G (3.4), -1442T>C (12.1), -1498T>G (25.0) and -1558T>G (2.6). The reported variations found in the study population and their frequencies were: -98T>C (28.4), -806C>T (2.6), -833del>T (9.5), 889T>G (10.3), -1041A>G (100.0) and -1418C>T (1.7). The two known non synonymous single nucleotide polymorphisms, 681G>A ((*)2 allele) and 636G>A ((*)3 allele) were detected at 0.371 and 0.025 frequencies, respectively. Forty three haplotypes were constructed and linkage disequilibrium analysis showed strong linkage between several variations identified in the gene. Fourteen polymorphisms including 8 novel ones in CYP2C19 5' flanking region are reported for the first time in an Indian population from South India. Results from this study provide additional information for genotyping of CYP2C19 in the South Indian population and probably in the Indian population.

Lack of association between Glu(298) asp polymorphism of endothelial nitric oxide synthase (eNOS) gene and coronary artery disease in Tamilian population.

OBJECTIVE: Glu298 Asp polymorphism of endothelial nitric oxide synthase (eNOS) gene has been recently implicated as a genetic marker for coronary artery disease (CAD) in some studies. There is no information on the prevalence of this polymorphism and its relationship with CAD in south Indian population. METHODS: A case control study was performed for the determination of the influence of Glu298 Asp polymorphism of eNOS gene in Tamilian population of south India. The study subjects comprised of 100 angiographically proven CAD patients and 100 age- and sex-matched volunteers asymptomatic for CAD, with a low coronary risk score. Genotyping of the eNOS gene was done by the polymerase chain reaction-restriction fragment length polymorphism (PCR RFLP) method. RESULTS: The genotype distribution was not significantly different between CAD (GG; 72, GT; 26, TT; 2) and control subjects (GG; 79, GT; 18, TT; 3). The corresponding allele frequencies were G 0.85, T 0.15 and G 0.88, T 0.12, respectively. The odds ratio for the association of CAD with the Asp variant failed to achieve statistical significance (OR = 0.66; 95% CI: 0.11-4.04, P = 1.0). CONCLUSION: No significant association was observed between the Glu298 Asp polymorphism and CAD in this population group.