CYP3A4 Variants Research Articles

Physiologically based pharmacokinetic combined BTK occupancy modeling for optimal dosing regimen prediction of acalabrutinib in patients alone, with different CYP3A4 variants, co-administered with CYP3A4 modulators and with hepatic impairment.

To develop a mathematical model combined between physiologically based pharmacokinetic and BTK occupancy (PBPK-BO) to simultaneously predict pharmacokinetic (PK) and pharmacodynamic (PD) changes of acalabrutinib (ACA) and active metabolite ACP-5862 in healthy humans as well as PD in patients. Next, to use the PBPK-BO to determine the optimal dosing regimens in patients alone, with different CYP3A4 variants, when co-administration with four CYP3A4 modulators and in patients with hepatic impairment, respectively. The PBPK-BO model was built using physicochemical and biochemical properties of ACA and ACP-5862 and then verified by observed PK and PD data from healthy humans and patients. Finally, the model was applied to determine optimal dosing regimens in various clinical situations. The simulations demonstrated that 100mg ACA twice daily (BID) was the optimal dosing regimen in patients alone. Additionally, dosage regimens might be reduced to 50mg BID in patients with five CYP3A4 variants. Moreover, the dosing regimen should be modified to 100mg (even to 50mg) once daily (QD) when co-administration with erythromycin or clarithromycin, and be increased to 200mg BID with rifampicin, and but be avoided co-administration with itraconazole. Furthermore, dosage regimen simulations showed that optimal dosing might be decreased to 50mg BID in patients with mild and moderate hepatic impairment, and be avoided taking ACA in severely hepatically impaired patients. This PBPK-BO model can predict PK and PD in healthy humans and patients and also predict the optimal dosing regimens in various clinical situations.

Prenatal organophosphorus pesticide exposure and executive function in preschool-aged children in the Norwegian Mother, Father and Child Cohort Study (MoBa)

IntroductionPrenatal exposure to organophosphorus pesticides (OPPs) has been associated with neurodevelopmental deficits in children, however evidence linking OPPs with specific cognitive mechanisms, such as executive function (EF), is limited. ObjectiveThis study aims to evaluate the association between prenatal exposure to OPPs with multiple measures of EF in preschool-aged children, while considering the role of variant alleles in OPP metabolism genes. MethodsWe included 262 children with preschool attention-deficit/hyperactivity disorder (ADHD), and 78 typically developing children, from the Preschool ADHD substudy of the Norwegian, Mother, Father, and Child Cohort Study. Participants who gave birth between 2004 and 2008 were invited to participate in an on-site clinical assessment when the child was approximately 3.5 years; measurements of EF included parent and teacher rating on Behavior Rating Inventory of Executive Function-Preschool (BRIEF-P), and three performance-based assessments. We measured OPP metabolites in maternal urines collected at ∼17 weeks' gestation to calculate total dimethyl- (ΣDMP) and diethyl phosphate (ΣDEP) metabolite concentrations. We estimated multivariable adjusted β′s and 95% confidence intervals (CIs) corresponding to a change in z-score per unit increase in log-ΣDMP/DEP. We further characterized gene-OPP interactions for maternal variants in PON1 (Q192R, M55L), CYP1A2 (1548T > C), CYP1A1 (IntG > A) and CYP2A6 (-47A > C). ResultsPrenatal OPP metabolite concentrations were associated with worse parent and teacher ratings of emotional control, inhibition, and working memory. A one log-∑DMP increase was associated with poorer teacher ratings of EF on the BRIEF-P (e.g. emotional control domain: β = 0.55, 95% CI: 0.35, 0.74), when weighted to account for sampling procedures. We found less consistent associations with performance-based EF assessments. We found some evidence of modification for PON1 Q192R and CYP2A6 -47A > C. Association with other variants were inconsistent. ConclusionsBiomarkers of prenatal OPP exposure were associated with more adverse teacher and parent ratings of EF in preschool-aged children.

Genetic diversity of CYP3A5 and ABCB1 variants in East-Central and South European populations

Background CYP3A5 enzyme encoded by CYP3A5 is important for drug metabolism in gut and liver, whereas P-glycoprotein by ABCB1, is an ATP-dependent drug efflux pump which exports endo- and exogenous substances outside the cell. Aim The study was to assess the prevalence of CYP3A5 alleles: *1, *2, *3, *4, *6 and *7, and C and T of ABCB1 in Poles, Belarusians and Bosnians and to compare it with the data reported from other European populations. Subjects and methods Overall, 511 unrelated healthy subjects from Poland (n = 239), Belarus (n = 104) and Bosnia and Herzegovina (n = 168) were included in this study. Allele frequencies and statistical parameters (AMOVA version 2.9.3) were determined. Results In Poles, Belarusians and Bosnians the *3 allele of CYP3A5 was the most common, and wild-type allele *1, were: 5.8%, 1.6% and 2.1%, respectively. Allele *2 was very rare, and alleles *4, *6 and *7 were not detected. For the populations mentioned above, the ABCB1 allele C was: 48.1%, 51.4%, 52.4%, respectively. Conclusion In compared populations, the distribution of CYP3A5 variants but not ABCB1, differed significantly. Alleles *4, *6 and *7 of CYP3A5 did not occur or occurred rarely.

Human genetic factors associated with pneumonia risk, a cue for COVID-19 susceptibility.

Pneumonia, an acute respiratory tract infection, is one of the major causes of mortality worldwide. Depending on the site of acquisition, pneumonia can be community acquired pneumonia (CAP) or nosocomial pneumonia (NP). The risk of pneumonia, is partially driven by host genetics. CYP1A1 is a widely studied pulmonary CYP family gene primarily expressed in peripheral airway epithelium. The CYP1A1 genetic variants, included in this study, alter the gene activity and are known to contribute in lung inflammation, which may cause pneumonia pathogenesis. In this study, we performed a meta-analysis to establish the possible contribution of CYP1A1 gene, and its three variants (rs2606345, rs1048943 and rs4646903) towards the genetic etiology of pneumonia risk. Using PRISMA guidelines, we systematically reviewed and meta-analysed case-control studies, evaluating risk of pneumonia in patients carrying the risk alleles of CYP1A1 variants. Heterogeneity across the studies was evaluated using I2 statistics. Based on heterogeneity, a random-effect (using maximum likelihood) or fixed-effect (using inverse variance) model was applied to estimate the effect size. Pooled odds ratio (OR) was calculated to estimate the overall effect of the risk allele association with pneumonia susceptibility. Egger's regression test and funnel plot were used to assess publication bias. Subgroup analysis was performed based on pneumonia type (CAP and NP), population, as well as age group. A total of ten articles were identified as eligible studies, which included 3049 cases and 2249 healthy controls. The meta-analysis findings revealed CYP1A1 variants, rs2606345 [T vs G; OR = 1.12 (0.75–1.50); p = 0.02; I2 = 84.89%], and rs1048943 [G vs T; OR = 1.19 (0.76–1.61); p = 0.02; I2 = 0.00%] as risk markers whereas rs4646903 showed no statistical significance for susceptibility to pneumonia. On subgroup analysis, both the genetic variants showed significant association with CAP but not with NP. We additionally performed a spatial analysis to identify the key factors possibly explaining the variability across countries in the prevalence of the coronavirus disease 2019 (COVID-19), a viral pneumonia. We observed a significant association between the risk allele of rs2606345 and rs1048943, with a higher COVID-19 prevalence worldwide, providing us important links in understanding the variability in COVID-19 prevalence.

Composite CYP3A phenotypes influence tacrolimus dose-adjusted concentration in lung transplant recipients.

Interpatient variability in tacrolimus pharmacokinetics is attributed to metabolism by cytochrome P-450 3A4/5 isoenzymes (encoded by CYP3A4 and CYP3A5). Guidelines for adjusting tacrolimus based on CYP3A5 test results are published; however, CYP3A4 variants also contribute to the variability in tacrolimus pharmacokinetics. The effects of composite phenotypes incorporating CYP3A5 and CYP3A4 increased (*1G, *1B) and decreased (*22) function variants have not been evaluated. The objective of this study is to investigate the impact of both increased and decreased function CYP3A variants on weight and dose-adjusted tacrolimus concentration (C0/D). We performed a single-center retrospective cohort study of lung transplant recipients to evaluate the median tacrolimus C0/D by composite CYP3A phenotype groups during the index transplant hospitalization. CYP3A4 and CYP3A5 alleles were used to classify patients into four CYP3A groups from least to most CYP3A activity. Exploratory analyses of ABCB1 and additional candidate genes were also assessed. Of the 92 included individuals, most (58) were CYP3A Group 2. The median tacrolimus C0/D differed significantly between CYP3A groups (P = 0.0001). CYP3A Group 2 median tacrolimus C0/D was 190.5 (interquartile range: 147.6-267.5) (ng/ml)/(mg/kg/d) and significantly higher than Group 4 [107.9 (90.4-116.1), P = 0.0001)]. Group 2 median tacrolimus C0/D did not significantly differ from Group 1 and Group 3 [373.5 (149.2-490.3) and 81.4 (62.6-184.1), respectively]. No significant differences in tacrolimus C0/D were found for the ABCB1 diplotypes. These data indicate that a composite CYP3A phenotype incorporating both increase and decrease variant information from CYP3A4 in addition to CYP3A5 may significantly influence tacrolimus C0/D during the early postoperative period.

The Influence of CYP3A4 Genetic Polymorphism and Proton Pump Inhibitors on Osimertinib Metabolism.

The aim of this study was to 1) investigate the effects of 27 CYP3A4 variants on the metabolism of osimertinib and 2) study the interactions between osimertinib and others as well as the underlying mechanism. A recombinant human CYP3A4 enzymatic incubation system was developed and employed to determine the kinetic profile of CYP3A4 variants. Ultra-performance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS) was applied to detect the concentration of the main metabolite, AZ5104. The results demonstrated that the relative clearance rates of CYP3A4.19, 10, 18, 5, 16, 14, 11, 2, 13, 12, 7, 8, and 17 in catalyzing osimertinib were significantly reduced to a minimum of 25.68% compared to CYP3A4.1, while those of CYP3A4.29, 32, 33, 28, 15, 34, and 3 were obviously enhanced, ranging from 114.14% to 284.52%. The activities of the remaining variants were almost equal to those of CYP3A4.1. In addition, 114 drugs were screened to determine the potential interaction with osimertinib based on the rat liver microsome (RLM) reaction system. Sixteen of them inhibited the production of AZ5104 to 20% or less, especially proton pump inhibitors, among which the IC50 of rabeprazole was 6.49 ± 1.17 μM in RLM and 20.39 ± 2.32 μM in human liver microsome (HLM), with both following competitive and non-competitive mixed mechanism. In an in vivo study, Sprague–Dawley (SD) rats were randomly divided into groups, with six animals per group, receiving osimertinib with or without rabeprazole, omeprazole, and lansoprazole. We found that the AUC(0–t), AUC(0–∞), and Cmax of osimertinib decreased significantly after co-administration with rabeprazole orally, but they increased remarkably when osimertinib was administered through intraperitoneal injection. Taken together, our data demonstrate that the genetic polymorphism and proton pump inhibitors remarkably influence the disposition of osimertinib, thereby providing basic data for the precise application of osimertinib.

Accuracy and applications of sequencing and genotyping approaches for CYP2A6 and homologous genes.

We evaluated multiple genotyping/sequencing approaches in a homologous region of chromosome 19, and investigated associations of two common 3'-UTR CYP2A6 variants with activity in vivo. Individuals (n = 1704) of European and African ancestry were phenotyped for the nicotine metabolite ratio (NMR), an index of CYP2A6 activity, and genotyped/sequenced using deep amplicon exon sequencing, SNP array, genotype imputation and targeted capture sequencing. Amplicon exon sequencing was the gold standard to which other methods were compared within-individual for CYP2A6, CYP2A7, CYP2A13, and CYP2B6 exons to identify highly discordant positions. Linear regression models evaluated the association of CYP2A6*1B and rs8192733 genotypes (coded additively) with logNMR. All approaches were ≤2.6% discordant with the gold standard; discordant calls were concentrated at few positions. Fifteen positions were discordant in >10% of individuals, with 12 appearing in regions of high identity between homologous genes (e.g. CYP2A6 and CYP2A7). For six, allele frequencies in our study and online databases were discrepant, suggesting errors in online sources. In the European-ancestry group (n = 935), CYP2A6*1B and rs8192733 were associated with logNMR (P < 0.001). A combined model found main effects of both variants on increasing logNMR. Similar trends were found in those of African ancestry (n = 506). Multiple genotyping/sequencing approaches used in this chromosome 19 region contain genotyping/sequencing errors, as do online databases. Gene-specific primers and SNP array probes must consider gene homology; short-read sequencing of related genes in a single reaction should be avoided. Using improved sequencing approaches, we characterized two gain-of-function 3'-UTR variants, including the relatively understudied rs8192733.

Effect of CYP3A5 and CYP3A4 Genetic Variants on Fentanyl Pharmacokinetics in a Pediatric Population.

Fentanyl is an anesthetic/analgesic commonly used in surgical and recovery settings. CYP3A4 and CYP3A5 encode enzymes, which metabolize fentanyl; genetic variants in these genes impact fentanyl pharmacokinetics in adults. Pharmacokinetic (PK) studies are difficult to replicate in children due to the burden of additional blood taken solely for research purposes. The aim of this study is to test the effect of CYP3A5 and CYP3A4 genetic variants on fentanyl PKs in children using opportunistically collected samples. Fentanyl concentrations were measured from remnant blood specimens and dosing data were extracted from electronic health records. Variant data defining CYP3A4*1G and CYP3A5*3 and *6 alleles were available from prior genotyping; alleles with no variant were defined as *1. The study cohort included 434 individuals (median age 9months, 52% male subjects) and 1,937 fentanyl concentrations were available. A two-compartment model was selected as the base model, and the final covariate model included age, weight, and surgical severity score. Clearance was significantly associated with either CYP3A5*3 or CYP3A5*6 alleles, but not the CYP3A4*1G allele. A genotype of CYP3A5*1/*3 or CYP3A5*1/*6 (i.e., intermediate metabolizer status) was associated with a 0.84-fold (95% confidence interval (CI): 0.71-1.00) reduction in clearance vs. CYP3A5*1/*1 (i.e., normal metabolizer status). CYP3A5*3/*3, CYP3A5*3/*6, or CYP3A5*6/*6 (i.e., poor metabolizer status) was associated with a 0.76-fold (95% CI: 0.58-0.99) reduction in clearance. In the final model, expected clearance was 8.9 and 6.8 L/hour for a normal and poor metabolizer, respectively, with median population covariates (9months old, 7.7kg, low surgical severity).

Effect of CYP3A4, CYP3A5, MDR1 and POR Genetic Polymorphisms in Immunosuppressive Treatment in Chilean Kidney Transplanted Patients.

Cyclosporine (CsA) and tacrolimus (TAC) are immunosuppressant drugs characterized by a narrow therapeutic range and high pharmacokinetic variability. The effect of polymorphisms in genes related to the metabolism and transport of these drugs, namely CYP3A4, CYP3A5, MDR1 and POR genes, has been evaluated in diverse populations. However, the impact of these polymorphisms on drug disposition is not well established in Latin American populations. Using TaqMan® probes, we determined the allelic frequency of seven variants in CYP3A4, CYP3A5, MDR1 and POR in 139 Chilean renal transplant recipients, of which 89 were treated with CsA and 50 with TAC. We tested associations between variants and trough and/or 2-hour concentrations, normalized by dose (C0/D and C2/D) at specific time points post-transplant. We found that CYP3A5*3/*3 carriers required lower doses of TAC. In TAC treated patients, most CYP3A5*3/*3 carriers presented higher C0/D and a high proportion of patients with C0 levels outside the therapeutic range relative to other genotypes. These results reinforce the value of considering CYP3A5 genotypes alongside therapeutic drug monitoring for TAC treated Chilean kidney recipients.

Predictors of Efavirenz Plasma Exposure, Auto-Induction Profile, and Effect of Pharmacogenetic Variations among HIV-Infected Children in Ethiopia: A Prospective Cohort Study

(1) Background: Efavirenz plasma concentration displays wide between-patient variability partly due to pharmacogenetic variation and autoinduction. Pediatric data on efavirenz pharmacokinetics and the relevance of pharmacogenetic variation are scarce, particularly from sub-Saharan Africa, where >90% of HIV-infected children live and population genetic diversity is extensive. We prospectively investigated the short- and long-term effects of efavirenz auto-induction on plasma drug exposure and the influence of pharmacogenetics among HIV-infected Ethiopian children. (2) Method: Treatment-naïve HIV-infected children aged 3–16 years old (n = 111) were enrolled prospectively to initiate efavirenz-based combination antiretroviral therapy (cART). Plasma efavirenz concentrations were quantified at 4, 8, 12, 24, and 48 weeks of cART. Genotyping for CYP2B6, CYP3A5, UGT2B7, ABCB1, and SLCO1B1 common functional variant alleles was performed. (3) Results: The efavirenz plasma concentration reached a peak at two months, declined by the 3rd month, and stabilized thereafter, with no significant difference in geometric mean over time. On average, one-fourth of the children had plasma efavirenz concentrations ≥4 µg/mL. On multivariate analysis, CYP2B6*6 and ABCB1c.3435 C > T genotypes and low pre-treatment low-density lipoprotein (LDL) were significantly associated with higher plasma efavirenz concentration regardless of treatment duration. Duration of cART, sex, age, nutritional status, weight, and SLCO1B, CYP3A5, UGT2B7, and ABCB1 rs3842 genotypes were not significant predictors of efavirenz plasma exposure. (4) Conclusion: Pre-treatment LDL cholesterol and CYP2B6*6 and ABCB1c.3435 C > T genotypes predict efavirenz plasma exposure among HIV-infected children, but treatment-duration-dependent changes in plasma efavirenz exposure due to auto-induction are not statistically significant.

Preliminary Pharmacogenomic-Based Predictive Models of Tamoxifen Response in Hormone-dependent Chilean Breast Cancer Patients.

Tamoxifen (TAM), a selective oestrogen receptor modulator, is one of the most used treatments in oestrogen receptor-positive (ER+) early and metastatic breast cancer (BC) patients. The response to TAM has a high degree of inter-individual variability. This is mainly due to genetic variants in CYP2D6 gene, as well as other genes encoding proteins involved in the TAM pharmacokinetic and/or pharmacodynamic. Therefore, prediction of the TAM response using these genetic factors together with other non-genetic variables may be relevant to improve breast cancer treatment. Thus, in this work, we used genetic polymorphisms and clinical variables for TAM response modelling. One hundred sixty-two ER + BC patients with 2 years of TAM treatment were retrospectively recruited, and the genetic polymorphisms CYP2D6*4, CYP3A4*1B (CYP3A4*1.001), CYP3A5*3, UGT2B7*2, UGT2B15*2, SULT1A1*2, and ESRA V364E were analyzed by PCR-RFLP. Concomitantly, the therapeutic response was obtained from clinical records for association with genotypes using univariate and multivariate biostatistical models. Our results show that UGT2B15*1/*2 genotype protects against relapse (OR = 0.09; p = 0.02), CYP3A5*3/*3 genotype avoids endometrial hyperplasia (OR = 0.07; p = 0.01), SULT1A1*1/*2 genotype avoids vaginal bleeding (OR = 0.09; p = 0.03) and ESRA 364E/364E genotype increases the probability of vaginal bleeding (OR = 5.68; p = 0.02). Logistic regression models, including genomic and non-genomic variables, allowed us to obtain preliminary predictive models to explain relapse (p = 0.010), endometrial hyperplasia (p = 0.002) and vaginal bleeding (p = 0.014). Our results suggest that the response to TAM treatment in ER + BC patients might be associated with the presence of the studied genetic variants in UGT2B15, CYP3A5, SULT1A1 and ESRA genes. After clinical validation protocols, these models might be used to help to predict a percentage of BC relapse and adverse reactions, improving the individual response to TAM-based treatment.

The Relationship of ABCB1/MDR1 and CYP1A1 Variants with the Risk of Disease Development and Shortening of Overall Survival in Patients with Multiple Myeloma.

(1) Background: The aim of our study was to analyze the possible relationship of ABCB1 and CYP1A1 gene variants with susceptibility and outcome of multiple myeloma (MM); (2) Methods: Genomic DNA samples from 110 newly-diagnosed MM patients and 100 healthy blood donors were analyzed by methods-PCR-RFLP (for ABCB1 3435C > T, CYP1A1 6235T > C—m1), automated DNA sequencing (for ABCB1 1236C > T, 2677G > T/A) and allele-specific PCR (for CYP1A1 4889A > G—m2); (3) Results: The genotypic frequencies of CYP1A1 4889A > G variant were not in Hardy-Weinberg equilibrium for MM patients. The presence of m1 and m2 CYP1A1 alleles decreased the risk of MM—OR = 0.49 (p = 0.011) and OR = 0.27 (p = 0.0003), respectively. In turn, TT genotype (ABCB1 2677G > T/A) increased the risk of this disease (p = 0.007). In the multivariate Cox analysis CT + TT genotypes (ABCB1 3435C > T) were associated with decreased risk of death (HR = 0.29, p = 0.04). In log-rank test in patients with CT genotype (ABCB1 3435C > T) was observed association of overall survival with the type of treatment; (4) Conclusions: Our findings suggest that T-alleles of ABCB1 2677G > T/A and m1/m2 alleles of CYP1A1 affected the susceptibility of MM. Moreover, T-allele of ABCB1 3435C > T might be independent positive prognostic factor in MM.

127 Impact of CYP2D6, CYP3A4, and CYP2C9 pharmacogenomic profile on pain relief and adverse events in Canadian children treated with oxycodone and ibuprofen

Abstract Primary Subject area Clinical Pharmacology and Toxicology Background Genomic variation impacts drug pharmacokinetics for commonly used children’s pain medications such as oxycodone and ibuprofen. In order to personalize and best treat children’s pain, how cytochrome enzyme polymorphisms for CYP2D6, CYP3A4, and CYP2C9 impact clinical effectiveness and safety for oxycodone and ibuprofen were studied. Objectives Primary objectives were to evaluate if allelic variations of CYP2D6, CYP2C9, and CYP3A4 would a) alter clinical effectiveness of ibuprofen and oxycodone for pain relief, and b) impact the occurrence of adverse events. Secondary objectives were to determine the degree to which these genetic and other clinical factors influence analgesic effectiveness and safety. Design/Methods This prospective, observational cohort included children aged 4-16 years who were seen in a pediatric emergency department (between June 2010 - July 2014) with an acute fracture and used ibuprofen OR oxycodone for at-home pain management. Saliva samples were obtained prior to discharge, and daily telephone follow-up collected self-reported pain scores, medication use, adverse events, and functional limitations for 3 days. Genotyping identified allelic variants of CYP2D6, CYP3A4, and CYP2C9. Pain was measured using the Faces Pain Scale-Revised. CYP2D6 metabolic phenotypes were determined based on identified variants. Regression analyses were employed to determine relationships between clinical and genomic patient characteristics, pain relief, and adverse events. Results We included 210 children (n=140 ibuprofen, n=70 oxycodone); mean age was 11.1 (±3.5) years, 66.2% were male, and 79.5% self-identified as Caucasian. The median pain reduction in the ibuprofen group was 4 (±2.0) and 4 (±3.5) in the oxycodone group on Day 1 (p = 0.69). Adverse events were experienced by 53.2% of the ibuprofen group and 78.3% of the oxycodone group (p &amp;lt; 0.001). CYP2D6 Intermediate Metabolizers had significantly less pain relief using oxycodone than Extensive Metabolizers (p = 0.04). CYP3A4 variants did not significantly impact pain relief or adverse events. Those with the decreased functioning CYP2C9*2 allele experienced less adverse events compared to the normal functioning allele CYP2C9*1 (p = 0.003) when using ibuprofen. Males (p = 0.035) and all children using non-pharmacological pain strategies (p = 0.02) experienced less pain relief with oxycodone. Conclusion A better understanding of pharmacogenomic variation could help personalize medication choice. Sex and non-pharmacologic pain management impact pain relief with oxycodone, warranting further study to better understand their relationship with opioid pain relief in children.

Unraveling the Genomic Architecture of the CYP3A Locus and ADME Genes for Personalized Tacrolimus Dosing.

Tacrolimus (TAC) is an immunosuppressant widely prescribed following an allogenic organ transplant. Due to wide interindividual pharmacokinetic (PK) variability, optimizing TAC dosing based on genetic factors is required to minimize nephrotoxicity and acute rejections. We enrolled 1133 participants receiving TAC from 4 cohorts, consisting of 3 with kidney transplant recipients and 1 with healthy males from clinical trials. The effects of clinical factors were estimated to appropriately control confounding variables. A genome-wide association study, haplotype analysis, and a gene-based association test were conducted using the Korea Biobank Array or targeted sequencing for 114 pharmacogenes. Genome-wide association study verified that CYP3A5*3 is the only common variant associated with TAC PK variability in Koreans. We detected several CYP3A5 and CYP3A4 rare variants that could potentially affect TAC metabolism. The haplotype structure of CYP3A5 stratified by CYP3A5*3 was a significant factor for CYP3A5 rare variant interpretation. CYP3A4 rare variant carriers among CYP3A5 intermediate metabolizers displayed higher TAC trough levels. Gene-based association tests in the 61 absorption, distribution, metabolism, and excretion genes revealed that CYP1A1 are associated with additional TAC PK variability: CYP1A1 rare variant carriers among CYP3A5 poor metabolizers showed lower TAC trough levels than the noncarrier controls. Our study demonstrates that rare variant profiling of CYP3A5 and CYP3A4, combined with the haplotype structures of CYP3A locus, provide additive value for personalized TAC dosing. We also identified a novel association between CYP1A1 rare variants and TAC PK variability in the CYP3A5 nonexpressers that needs to be further investigated.

Functional characterization of novel rare CYP2A6 variants and potential implications for clinical outcomes.

CYP2A6 activity, phenotyped by the nicotine metabolite ratio (NMR), is a predictor of several smoking behaviors, including cessation and smoking‐related disease risk. The heritability of the NMR is 60–80%, yet weighted genetic risk scores (wGRSs) based on common variants explain only 30–35%. Rare variants (minor allele frequency <1%) are hypothesized to explain some of this missing heritability. We present two targeted sequencing studies where rare protein‐coding variants are functionally characterized in vivo, in silico, and in vitro to examine this hypothesis. In a smoking cessation trial, 1687 individuals were sequenced; characterization measures included the in vivo NMR, in vitro protein expression, and metabolic activity measured from recombinant proteins. In a human liver bank, 312 human liver samples were sequenced; measures included RNA expression, protein expression, and metabolic activity from extracted liver tissue. In total, 38 of 47 rare coding variants identified were novel; characterizations ranged from gain‐of‐function to loss‐of‐function. On a population level, the portion of NMR variation explained by the rare coding variants was small (~1%). However, upon incorporation, the accuracy of the wGRS was improved for individuals with rare protein‐coding variants (i.e., the residuals were reduced), and approximately one‐third of these individuals (12/39) were re‐assigned from normal to slow metabolizer status. Rare coding variants can alter an individual’s CYP2A6 activity; their integration into wGRSs through precise functional characterization is necessary to accurately assess clinical outcomes and achieve precision medicine for all. Investigation into noncoding variants is warranted to further explain the missing heritability in the NMR.

Deciphering Structural Alterations Associated with Activity Reductions of Genetic Polymorphisms in Cytochrome P450 2A6 Using Molecular Dynamics Simulations.

Cytochrome P450 (CYP) 2A6 is a monooxygenase involved in the metabolism of various endogenous and exogenous chemicals, such as nicotine and therapeutic drugs. The genetic polymorphisms in CYP2A6 are a cause of individual variation in smoking behavior and drug toxicities. The enzymatic activities of the allelic variants of CYP2A6 were analyzed in previous studies. However, the three-dimensional structures of the mutants were not investigated, and the mechanisms underlying activity reduction remain unknown. In this study, to investigate the structural changes involved in the reduction in enzymatic activities, we performed molecular dynamics simulations for ten allelic mutants of CYP2A6. For the calculated wild type structure, no significant structural changes were observed in comparison with the experimental structure. On the other hand, the mutations affected the interaction with heme, substrates, and the redox partner. In CYP2A6.44, a structural change in the substrate access channel was also observed. Those structural effects could explain the alteration of enzymatic activity caused by the mutations. The results of simulations provide useful information regarding the relationship between genotype and phenotype.

Nicotine metabolism and its association with CYP2A6 genotype among Indigenous people in Alaska who smoke.

Prevalence of smoking is higher in Alaska Native and American Indian (ANAI) populations living in Alaska than the general US population. Genetic factors contribute to smoking and cessation rates. The objective of this study was to compare CYP2A6 genetic variation and CYP2A6 enzyme activity toward nicotine in an ANAI population. ANAI (N = 151) people trying to quit smoking were recruited. DNA samples were genotyped for CYP2A6 variants *1X2A, *1B, *2, *4, *9, *10, *12, and *35. Multiple nicotine metabolites were measured in plasma and urine samples, including cotinine and 3′‐hydroxycotinine used to determine CYP2A6 activity (e.g., nicotine metabolite ratio [NMR]). We calculated summary statistics for all of the genotypes and metabolites and assigned CYP2A6 activity scores based on known information. We studied the association of CYP2A6 variants with the NMR and smoking histories. The overall frequency of the CYP2A6*1B gain of function allele was high in the ANAI versus non‐ANAI populations in other studies. Both *4 null and *9 decrease of function alleles had frequencies similar to previous studies of ANAI populations. In a multivariate analysis, the genotype‐inferred CYP2A6 activity score was associated with both plasma and urine NMR (p value = 8.56E‐08 and 4.08E‐13, respectively). Plasma NMR was also associated with duration of smoking (p value < 0.01) but not urinary total nicotine equivalents uncorrected for creatinine (TNE9uc) or biological sex. Urine NMR was significantly associated (p value < 0.01) with TNE9uc. Variation in NMR in this ANAI population is explained in part by CYP2A6 genetic variation.

Effect of Genetic Variation in CYP450 on Gonadal Impairment in a European Cohort of Female Childhood Cancer Survivors, Based on a Candidate Gene Approach: Results from the PanCareLIFE Study.

Simple SummaryChildhood cancer patients receiving treatment containing alkylating agents are at risk of infertility, yet inter-individual variability in treatment-related ovarian damage is observed. Alkylating agents are metabolized by cytochrome P450 (CYP450) enzymes and polymorphisms in these CYP450 enzymes may explain this variability in ovarian damage. This study on genetic variation in CYP450 enzymes of chemotherapy-induced gonadotoxicity, using anti-Müllerian hormone (AMH) levels as a proxy for ovarian reserve, in female childhood cancer survivors (CCSs) may identify patients at risk of infertility. This unique global collaboration of two large CCS studies shows the significant gonadotoxic effect of enzyme CYP3A4*3 and significant protective effect of CYP2B6*2 on gonadal function in CCSs receiving alkylating agents. Genetic variation in CYP3A4 and CYP2B6 have previously been associated with gonadotoxicity after cancer treatment. These findings could guide risk prediction models determining patients at risk of chemotherapy-induced gonadal impairment.Background: Female childhood cancer survivors (CCSs) carry a risk of therapy-related gonadal dysfunction. Alkylating agents (AA) are well-established risk factors, yet inter-individual variability in ovarian function is observed. Polymorphisms in CYP450 enzymes may explain this variability in AA-induced ovarian damage. We aimed to evaluate associations between previously identified genetic polymorphisms in CYP450 enzymes and AA-related ovarian function among adult CCSs. Methods: Anti-Müllerian hormone (AMH) levels served as a proxy for ovarian function in a discovery cohort of adult female CCSs, from the pan-European PanCareLIFE cohort (n = 743; age (years): median 25.8, interquartile range (IQR) 22.1–30.6). Using two additive genetic models in linear and logistic regression, nine genetic variants in three CYP450 enzymes were analyzed in relation to cyclophosphamide equivalent dose (CED) score and their impact on AMH levels. The main model evaluated the effect of the variant on AMH and the interaction model evaluated the modifying effect of the variant on the impact of CED score on log-transformed AMH levels. Results were validated, and meta-analysis performed, using the USA-based St. Jude Lifetime Cohort (n = 391; age (years): median 31.3, IQR 26.6–37.4). Results: CYP3A4*3 was significantly associated with AMH levels in the discovery and replication cohort. Meta-analysis revealed a significant main deleterious effect (Beta (95% CI): −0.706 (−1.11–−0.298), p-value = 7 × 10−4) of CYP3A4*3 (rs4986910) on log-transformed AMH levels. CYP2B6*2 (rs8192709) showed a significant protective interaction effect (Beta (95% CI): 0.527 (0.126–0.928), p-value = 0.01) on log-transformed AMH levels in CCSs receiving more than 8000 mg/m2 CED. Conclusions: Female CCSs CYP3A4*3 carriers had significantly lower AMH levels, and CYP2B6*2 may have a protective effect on AMH levels. Identification of risk-contributing variants may improve individualized counselling regarding the treatment-related risk of infertility and fertility preservation options.

Nicotine Metabolism Predicted by CYP2A6 Genotypes in Relation to Smoking Cessation: A Systematic Review.

Identifying genetic factors associated with smoking cessation could inform precision cessation interventions. Of major interest is genetic variation in nicotine metabolism, largely predicted by CYP2A6 variations. We conducted a systematic literature review to summarize the population-based evidence of the association between CYP2A6 and smoking cessation. In the 12 studies meeting the inclusion criteria, the known functional metabolic effect of CYP2A6 variants was used to classify nicotine metabolism as normal (>75% metabolic activity), intermediate (50.1%-75% activity), slow (25%-50% activity), and poor (<25% activity). Summary odds ratios of smoking cessation were calculated across metabolic groups, stratified by ancestry and whether participants received pharmacotherapy or placebo/no treatment. Among untreated people of European ancestry (n = 4 studies), those with CYP2A6 reduced metabolism were more likely to quit smoking than those with normal metabolism (Summary OR = 2.05, 95% CI 1.23 to 3.42) and the likelihood of cessation increased as nicotine metabolism decreased. Nicotine replacement therapy attenuated the association at end-of-treatment, while bupropion modified the association such that intermediate/slow metabolizers were less likely to quit than normal metabolizers (Summary OR = 0.86, 95% CI 0.79 to 0.94). Among untreated Asian people (n = 3 studies), results differed compared with those with European ancestry: those with slow metabolism were less likely to have quit smoking than normal metabolizers (Summary OR = 0.52, 95% CI 0.38 to 0.71). Evidence for people of African ancestry (n = 1 study) suggested the CYP2A6 association with cessation may differ compared with those of European ancestry. Most studies included in this review were of European ancestry populations; these showed slower nicotine metabolism was associated with increased likelihood of smoking cessation in a dose-related manner. Pharmacotherapy appeared to attenuate or modify this association among people of European ancestry, but it is unclear whether the change in the association remains consistent after treatment ceases. This finding has implications for precision medicine cessation interventions. Based on only a few studies of people of Asian or African ancestry, the association between CYP2A6 variants and cessation may differ from that observed among those of European ancestry, but more evidence is needed.

The influence of CYP2C19*2 and CYP3A5*3 variants on the development of depression and effectiveness of therapy: A preliminary study

The most common mental illness is depression; however, its pathogenesis is not fully understood. One of the factors that may influence its development and the effectiveness of therapy are the cytochromes of the p450 complex. CYP3A5 and CYP2C19 are involved in the metabolism of drugs used in the treatment of depression. These cytochromes can also generate reactive oxygen species, which are known to participate in the pathogenesis of depression. The aim of the study was to determine the frequency of CYP3A5*3 and CYP2C19*2 variants among a group of patients with depression to identify any potential association with disease development and progression, and the effectiveness of pharmacotherapy. A group of 103 patients suffering from recurrent depressive disorder and another of 93 healthy individuals were investigated using RFLP. It was found that the CYP3A5*3 allele may have a protective role in the development of depression (p = 0.0036). Heterozygous CYP3A5*1/*3 was more common in controls than the patients (p = 0.0300). Homozygotes were associated with an earlier onset than heterozygotes (p = 0.0292). For CYP2C19, patients with at least one CYP2C19*2 allele revealed better treatment results expressed as percentage change in Hamilton Depression Rating Scale (p = 0.0239). The identification of CYP3A5 and CYP2C19 allelic variants may be useful when assessing the effectiveness of pharmacotherapy.