Potential Association of Cytochrome P450 Copy Number Alteration in Tumour with Chemotherapy Resistance in Lung Adenocarcinoma Patients.

Resistance to anticancer agents is a major obstacle to efficacious tumour therapy and responsible for high cancer-related mortality rates. Some resistance mechanisms are associated with pharmacokinetic variability in anticancer drug exposure due to genetic polymorphisms of drug-metabolizing cytochrome P450 (CYP) enzymes, whereas variations in tumoural metabolism as a consequence of CYP copy number alterations are assumed to contribute to the selection of resistant cells. A high-throughput quantitative polymerase chain reaction (qPCR)-based method was developed for detection of CYP copy number alterations in tumours, and a scoring system improved the identification of inappropriate reference genes that underwent deletion/multiplication in tumours. The copy numbers of both the target (CYP2C8, CYP3A4) and the reference genes (ALB, B2M, BCKDHA, F5, CD36, MPO, TBP, RPPH1) established in primary lung adenocarcinoma by the qPCR-based method were congruent with those determined by next-generation sequencing (for 10 genes, slope = 0.9498, r2 = 0.72). In treatment naïve adenocarcinoma samples, the copy number multiplication of paclitaxel-metabolizing CYP2C8 and/or CYP3A4 was more prevalent in non-responder patients with progressive disease/exit than in responders with complete remission. The high-throughput qPCR-based method can become an alternative approach to next-generation sequencing in routine clinical practice, and identification of altered CYP copy numbers may provide a promising biomarker for therapy-resistant tumours.

Association between CYP2B6 genetic variability and cyclophosphamide therapy in pediatric patients with neuroblastoma.

Cyclophosphamide, an oxazaphosphorine prodrug is frequently used in treatment of neuroblastoma, which is one of the most prevalent solid organ malignancies in infants and young children. Cytochrome P450 2B6 (CYP2B6) is the major catalyst and CYP2C19 is the minor enzyme in bioactivation and inactivation pathways of cyclophosphamide. CYP-mediated metabolism may contribute to the variable pharmacokinetics of cyclophosphamide and its toxic byproducts leading to insufficient response to the therapy and development of clinically significant side effects. The aim of the study was to reveal the contribution of pharmacogenetic variability in CYP2B6 and CYP2C19 to the treatment efficacy and cyclophosphamide-induced side effects in pediatric neuroblastoma patients under cyclophosphamide therapy (N = 50). Cyclophosphamide-induced hematologic toxicities were pivotal in all patients, whereas only moderate hepatorenal toxicity was developed. The patients' CYP2B6 metabolizer phenotypes were associated with the occurrence of lymphopenia, thrombocytopenia, and monocytopenia as well as of liver injury, but not with kidney or urinary bladder (hemorrhagic cystitis) toxicities. Furthermore, the patients' age (< 1.5 years, P = 0.03) and female gender (P ≤ 0.02), but not CYP2B6 or CYP2C19 metabolizer phenotypes appeared as significant prognostic factors in treatment outcomes. Our results may contribute to a better understanding of the impact of CYP2B6 variability on cyclophosphamide-induced side effects.

Possible genetical predictors of efficacy and safety of budesonide-MMX in patients with mild-to-moderate ulcerative colitis, and safety comparison with methylprednisolone.

BACKGROUND: Budesonide-MMX is a topically active corticosteroid degraded by cytochrome-P450 enzymes, resulting in favorable side-effect profile. We aimed to assess the effect of CYP genotypes on safety and efficacy, and make a direct comparison with systemic corticosteroids. RESEARCH DESIGN AND METHODS: We enrolled UC patients receiving budesonide-MMX and IBD patients on methylprednisolone in our prospective, observational-cohort study. Before and after treatment regimen clinical activity indexes, laboratory parameters (electrolytes, CRP, cholesterol, triglyceride, dehydroepiandrosterone, cortisol, beta-crosslaps, osteocalcin), and body composition measurements were assessed. CYP3A4 and CYP3A5 genotypes were determined in the budesonide-MMX group. RESULTS: 71 participants were enrolled (budesonide-MMX: 52; methylprednisolone: 19). CAI decreased (p<0.05) in both groups. Cortisol decreased (p<0.001), and the level of cholesterol was elevated in both groups (p<0.001). Body composition altered only following methylprednisolone. Bone homeostasis (osteocalcin; p<0.05) and DHEA (p<0.001) changed more prominently after methylprednisolone. Glucocorticoid-related adverse events were more common following methylprednisolone treatment (47.4% compared to 1.9%). CYP3A5(*1/*3) genotype positively influenced efficacy, but not safety. Only one patient's CYP3A4 genotype differed. CONCLUSIONS: CYP genotypes can affect the efficacy of budesonide-MMX; however, further studies would be needed with analyses of gene expression. Although budesonide-MMX is safer than methylprednisolone, due to glucocorticoid-related side effects, admission should require greater precaution.

The Prognostic Role of CYP Enzyme in Kidney Transplantation: A Single Centre Experience.

BACKGROUND: The main goal of immunosuppressive agents is to reach a balance of preserving allograft function while minimizing adverse effects. The purpose of our research is to corroborate the role of CYP3A enzyme in developing individual medication therapy via measuring medicine levels in patients' blood samples. METHODS: This retrospective analysis studies 15 kidney transplant recipients. We carried out genotyping (CYP3A5, CYP3A4) after isolating DNA and RNA in patient and donor blood samples; we also determined CYP3A4 messenger RNA expression in case of recipients. Tacrolimus blood levels, dosage, and tacrolimus concentration normalized by dose and the body weight (C0/D ratio) were evaluated. RESULTS: In this research, recipients were divided into 2 groups based on their CYP3A5 genotype. Those who carry CYP3A5*1 allele (*1/*1 or *1/*3) are CYP3A5 expressors, whereas those who are homozygous for the nonfunctional CYP3A5*3 allele are CYP3A5 nonexpressors. There were 3 patients with functioning CYP3A5 enzyme (patients with CYP3A5*1/*3 genotype) where increased tacrolimus metabolism was expected. Our data show that C0/D ratio of CYP3A5 nonexpressors was around 3 times higher than of CYP3A5 expressors. Looking at CYP3A4 enzyme, we found 1 patient carried CYP3A4*22/*22 genotype where we expected decreased CYP3A4 expression. It is clear that this patient had adequate therapy medication levels (9.50 µg/L) despite having received very low dosage of tacrolimus (0.03 mg/weight/d). CONCLUSIONS: Our results confirmed the importance of determining CYP status of recipients after a transplant because individual differences were observed in tacrolimus treatment that were partly influenced by CYP status of recipients.

CYP1A2 mRNA Expression Rather than Genetic Variants Indicate Hepatic CYP1A2 Activity.

CYP1A2, one of the most abundant hepatic cytochrome P450 enzymes, is involved in metabolism of several drugs and carcinogenic compounds. Data on the significance of CYP1A2 genetic polymorphisms in enzyme activity are highly inconsistent; therefore, the impact of CYP1A2 genetic variants (−3860G>A, −2467delT, −739T>G, −163C>A, 2159G>A) on mRNA expression and phenacetin O-dealkylation selective for CYP1A2 was investigated in human liver tissues and in psychiatric patients belonging to Caucasian populations. CYP1A2*1F, considered to be associated with high CYP1A2 inducibility, is generally identified by the presence of −163C>A polymorphism; however, we demonstrated that −163C>A existed in several haplotypes (CYP1A2*1F, CYP1A2*1L, CYP1A2*1M, CYP1A2*1V, CYP1A2*1W), and consequently, CYP1A2*1F was a much rarer allelic variant (0.4%) than reported in Caucasian populations. Of note, −163C>A polymorphism was found to result in an increase of neither mRNA nor the activity of CYP1A2. Moreover, hepatic CYP1A2 activity was associated with hepatic or leukocyte mRNA expression rather than genetic polymorphisms of CYP1A2. Consideration of non-genetic phenoconverting factors (co-medication with CYP1A2-specific inhibitors/inducers, tobacco smoking and non-specific factors, including amoxicillin+clavulanic acid therapy or chronic alcohol consumption) did not much improve genotype−phenotype estimation. In conclusion, CYP1A2-genotyping is inappropriate for the prediction of CYP1A2 function; however, CYP1A2 mRNA expression in leukocytes can inform about patients' CYP1A2-metabolizing capacity.

CYP2B6 allelic variants and non-genetic factors influence CYP2B6 enzyme function.

Human CYP2B6 enzyme although constitutes relatively low proportion (1-4%) of hepatic cytochrome P450 content, it is the major catalyst of metabolism of several clinically important drugs (efavirenz, cyclophosphamide, bupropion, methadone). High interindividual variability in CYP2B6 function, contributing to impaired drug-response and/or adverse reactions, is partly elucidated by genetic polymorphisms, whereas non-genetic factors can significantly modify the CYP2B6 phenotype. The influence of genetic and phenoconverting non-genetic factors on CYP2B6-selective activity and CYP2B6 expression was investigated in liver tissues from Caucasian subjects (N = 119). Strong association was observed between hepatic S-mephenytoin N-demethylase activity and CYP2B6 mRNA expression (P < 0.0001). In less than one third of the tissue donors, the CYP2B6 phenotype characterized by S-mephenytoin N-demethylase activity and/or CYP2B6 expression was concordant with CYP2B6 genotype, whereas in more than 35% of the subjects, an altered CYP2B6 phenotype was attributed to phenoconverting non-genetic factors (to CYP2B6-specific inhibitors and inducers, non-specific amoxicillin + clavulanic acid treatment and chronic alcohol consumption, but not to the gender). Furthermore, CYP2B6 genotype-phenotype mismatch still existed in one third of tissue donors. In conclusion, identifying potential sources of CYP2B6 variability and considering both genetic variations and non-genetic factors is a pressing requirement for appropriate elucidation of CYP2B6 genotype-phenotype mismatch.

CYP3A-status is associated with blood concentration and dose-requirement of tacrolimus in heart transplant recipients.

High inter-individual variability in tacrolimus clearance is attributed to genetic polymorphisms of CYP3A enzymes. However, due to CYP3A phenoconversion induced by non-genetic factors, continuous changes in tacrolimus-metabolizing capacity entail frequent dose-refinement for optimal immunosuppression. In heart transplant recipients, the contribution of patients' CYP3A-status (CYP3A5 genotype and CYP3A4 expression) to tacrolimus blood concentration and dose-requirement was evaluated in the early and late post-operative period. In low CYP3A4 expressers carrying CYP3A5*3/*3, the dose-corrected tacrolimus level was significantly higher than in normal CYP3A4 expressers or in those with CYP3A5*1. Modification of the initial tacrolimus dose was required for all patients: dose reduction by 20% for low CYP3A4 expressers, a 40% increase for normal expressers and a 2.4-fold increase for CYP3A5*1 carriers. The perioperative high-dose corticosteroid therapy was assumed to ameliorate the low initial tacrolimus-metabolizing capacity during the first month. The fluctuation of CYP3A4 expression and tacrolimus blood concentration (C0/D) was found to be associated with tapering and cessation of corticosteroid in CYP3A5 non-expressers, but not in those carrying CYP3A5*1. Although monitoring of tacrolimus blood concentration cannot be omitted, assaying recipients' CYP3A-status can guide optimization of the initial tacrolimus dose, and can facilitate personalized tacrolimus therapy during steroid withdrawal in the late post-operative period.

Impact of genetic and non-genetic factors on hepatic CYP2C9 expression and activity in Hungarian subjects.

CYP2C9, one of the most abundant hepatic cytochrome P450 enzymes, is involved in metabolism of 15-20% of clinically important drugs (warfarin, sulfonylureas, phenytoin, non-steroid anti-inflammatory drugs). To avoid adverse events and/or impaired drug-response, CYP2C9 pharmacogenetic testing is recommended. The impact of CYP2C9 polymorphic alleles (CYP2C9*2, CYP2C9*3) and phenoconverting non-genetic factors on CYP2C9 function and expression was investigated in liver tissues from Caucasian subjects (N = 164). The presence of CYP2C9*3 allele was associated with CYP2C9 functional impairment, and CYP2C9*2 influenced tolbutamide 4'-hydroxylase activity only in subjects with two polymorphic alleles, whereas the contribution of CYP2C8*3 was not confirmed. In addition to CYP2C9 genetic polymorphisms, non-genetic factors (co-medication with CYP2C9-specific inhibitors/inducers and non-specific factors including amoxicillin + clavulanic acid therapy or chronic alcohol consumption) contributed to the prediction of hepatic CYP2C9 activity; however, a CYP2C9 genotype-phenotype mismatch still existed in 32.6% of the subjects. Substantial variability in CYP2C9 mRNA levels, irrespective of CYP2C9 genotype, was demonstrated; however, CYP2C9 induction and non-specific non-genetic factors potentially resulting in liver injury appeared to modify CYP2C9 expression. In conclusion, complex implementation of CYP2C9 genotype and non-genetic factors for the most accurate estimation of hepatic CYP2C9 activity may improve efficiency and safety of medication with CYP2C9 substrate drugs in clinical practice.