Efavirenz Metabolism Research Articles

Activity Variations of CYP2B6 Determine the Metabolic Stratification of Efavirenz.

To investigate the effects of hepatic enzyme activity variations and CYP2B6 gene polymorphisms on the in vivo and in vitro metabolism of efavirenz. In vitro enzyme systems using rat and human liver microsomes (RLM/HLM) were established, with in vivo studies conducted on Sprague-Dawley rats. Metabolite detection was performed via LC-MS/MS. Human recombinant CYP2B6 microsomes were prepared using a baculovirus-insect cell system and ultracentrifugation, with efavirenz serving as the substrate to study enzyme kinetics. Isavuconazole exhibited an IC50 of 21.14 ± 0.57 μM in RLM, indicating a mixed competitive and noncompetitive mechanism, and an IC50 of 40.44 ± 4.23 μM in HLM, suggesting an anticompetitive mechanism. In rats, coadministration of efavirenz and isavuconazole significantly increased the AUC, Tmax, and Cmax of efavirenz. Co-administration of efavirenz and rifampicin significantly elevated the AUC, Tmax, and Cmax of 8-OH-efavirenz. The activity of CYP2B6.4, 6, and 7 increased significantly compared to CYP2B6.1, with relative clearance ranging from 158.34% to 212.72%. Conversely, the activity of CYP2B6.3, 8, 10, 11, 13-15, 18-21, 23-27, 31-33, and 37 was markedly reduced, ranging from 4.30% to 79.89%. Variations in liver enzyme activity and CYP2B6 genetic polymorphisms can significantly alter the metabolism of efavirenz. It provides laboratory-based data for the precise application of efavirenz and other CYP2B6 substrate drugs.

Quantification of Efavirenz Hydroxymetabolites in Human Plasma Using LC-HRMS/MS.

Efavirenz (EFV) is a drug used to treat HIV. Low plasma concentrations of EFV result in suboptimal viral suppression, whereas high concentrations can cause adverse neuropsychiatric side reactions. Some studies have identified a correlation between the plasma concentrations of EFV metabolites and neurotoxicity. To our knowledge, no studies have investigated the metabolism of EFV in young children and its effect on treatment outcomes. Therefore, the aim of this study was to develop and validate a method for quantifying EFV and its metabolites in human plasma derived from children. Sample preparation was performed using protein precipitation of 100 µL plasma. Thereafter, an aliquot of the supernatant was used to quantify EFV, 7-hydroxyefavirenz (7-OH-EFV), 8-hydroxyefavirenz (8-OH-EFV), and a newly discovered metabolite ("EFAdeg") associated with 8-OH-EFV. A second aliquot of the supernatant was hydrolyzed using β-glucuronidase/arylsulfatase and used with the first aliquot to quantify phase II metabolites. The analyses were performed using a Dionex Ultimate 3000RS LC-system coupled with a Q Exactive Orbitrap mass spectrometer. The method has a measuring range of 100-50,000 ng/mL (EFV, 8-OH-EFV), 125-25,000 ng/mL (7-OH-EFV), and 200-10,000 ng/mL ("EFAdeg"). All criteria of the European Medicines Agency guidelines regarding precision, accuracy, and selectivity were met. Of note, carryover must be considered for 8-OH-EFV. Overall, the validated method was successfully applied to plasma samples obtained from children and confirmed the presence of the newly discovered metabolite, "EFAdeg." An LC-HRMS/MS method for the quantification of EFV and its phase I and II metabolites was developed and validated. This method is suitable for analyzing plasma samples from children. Furthermore, studies using this method identified an additional metabolite that may influence the concentration of 8-OH-EFV in patient samples.

Biotransformation of Efavirenz and Proteomic Analysis of Cytochrome P450s and UDP-Glucuronosyltransferases in Mouse, Macaque, and Human Brain-Derived In Vitro Systems.

Antiretroviral drugs such as efavirenz (EFV) are essential to combat human immunodeficiency virus (HIV) infection in the brain, but little is known about how these drugs are metabolized locally. In this study, the cytochrome P450 (P450) and UDP-glucuronosyltransferase (UGT)-dependent metabolism of EFV was probed in brain microsomes from mice, cynomolgus macaques, and humans as well as primary neural cells from C57BL/6N mice. Utilizing ultra high performance liquid chromatography high-resolution mass spectrometry (uHPLC-HRMS), the formation of 8-hydroxyefavirenz (8-OHEFV) from EFV and the glucuronidation of P450-dependent metabolites 8-OHEFV and 8,14-dihydroxyefavirenz (8,14-diOHEFV) were observed in brain microsomes from all three species. The direct glucuronidation of EFV, however, was only detected in cynomolgus macaque brain microsomes. In primary neural cells treated with EFV, microglia were the only cell type to exhibit metabolism, forming 8-OHEFV only. In cells treated with the P450-dependent metabolites of EFV, glucuronidation was detected only in cortical neurons and astrocytes, revealing that certain aspects of EFV metabolism are cell type specific. Untargeted and targeted proteomics experiments were used to identify the P450s and UGTs present in brain microsomes. Eleven P450s and 11 UGTs were detected in human brain microsomes, whereas seven P450s and 14 UGTs were identified in mouse brain microsomes and 15 P450s and four UGTs, respectively, were observed in macaque brain microsomes. This was the first time many of these enzymes have been noted in brain microsomes at the protein level. This study indicates the potential for brain metabolism to contribute to pharmacological and toxicological outcomes of EFV in the brain. SIGNIFICANCE STATEMENT: Metabolism in the brain is understudied, and the persistence of human immunodeficiency virus (HIV) infection in the brain warrants the evaluation of how antiretroviral drugs such as efavirenz are metabolized in the brain. Using brain microsomes, the metabolism of efavirenz by both cytochrome P450s (P450s) and UDP-glucuronosyltransferases (UGTs) is established. Additionally, proteomics of brain microsomes characterizes P450s and UGTs in the brain, many of which have not yet been noted in the literature at the protein level.

Effects of rifampicin, CYP2B6 and ABCB1 polymorphisms on efavirenz plasma concentration in Chinese patients living with HIV and tuberculosis.

Tuberculosis (TB) is the leading opportunistic infection of people living with human immunodeficiency virus (HIV; PLWH). Cytochrome P450 (CYP) 2B6 and ATP-binding cassette sub-family B member 1 (ABCB1) are involved in the metabolism and transportation of efavirenz. The study was aimed to investigate the effects of rifampicin, CYP2B6 and ABCB1 polymorphisms on efavirenz exposure in Chinese PLWH co-infected with TB. PLWH were screened according to inclusion and exclusion criteria and divided into HIV group and HIV/TB group. Efavirenz plasma concentration (C0) was determined, dose-adjusted concentration (C0/D) was calculated, and genotypes of CYP2B6 516G>T, 785A>G, and ABCB1 2677G>T, 3435C>T were analyzed. 252 PLWH were enrolled, including 75 co-infected with TB and concomitant with rifampicin. Efavirenz C0 and C0/D were both higher in HIV group (1.94μg/mL, 0.2007 (μg/ml)/(mg/kg/d)) compared with HIV/TB group (1.52μg/mL, 0.1557 (μg/ml)/(mg/kg/d)) (p = .001). Efavirenz C0/D was significantly higher in patients with variant genotypes of CYP2B6 516G>T and 785A>G (p<.001), and was significantly lower in HIV/TB group compared with HIV group among patients with CYP2B6 516GG, TT, and 785 AA, AG genotypes (p < .05). Efavirenz exposure is reduced by co-administration with rifampicin, and related to genetic polymorphisms of CYP2B6.

Improvement in Bioavailability and Pharmacokinetic Characteristics of Efavirenz with Booster Dose of Ritonavir in PEGylated PAMAM G4 Dendrimers.

Efavirenz (EFV) with a booster dose of ritonavir (RTV) (EFV-RTV) inhibits the metabolism of EFV and improves its bioavailability. However, inadequate organ perfusion with surface permeability glycoprotein (P-gp) efflux sustains the viable HIV. Hence, the present investigations were aimed to evaluate the pharmacokinetics and tissue distribution efficiency of EFV by encapsulating it into PEGyalated PAMAM (polyamidoamine) G4 dendrimers with a booster dose of RTV (PPG4ER). The entrapment efficiency of PEGylated PAMAM G4 dendrimers was found to be 94% and 92.12% for EFV and RTV respectively with a zeta potential of 0.277mV. The pharmacokinetics and tissue distribution behavior of EFV within PPG4ER was determined by developing and validating a simple, sensitive, and reliable bioanalytical method of RP-HPLC. The developed bioanalytical method was very sensitive with a quantification limit of 18.5ng/ml and 139.2ng/ml for EFV and RTV, respectively. The comparative noncompartmental pharmacokinetic parameters of EFV were determined by administrating a single intraperitoneal dose of EFV, EFV-RTV, and PPG4ER to Wistar rats. The PPG4ER produced prolonged release of EFV with a mean residential time (MRT) of 24h with Cmax 7.68µg/ml in plasma against EFV-RTV with MRT 11h and Cmax 3.633µg/ml. The PPG4ER was also detected in viral reservoir tissues (lymph node and spleen) for 3-4days, whereas free EFV and EFV-RTV were cleared within 72h. The pharmacokinetic data including Cmax, t1/2, AUCtot, and MRT were significantly improved in PPG4ER as compared with single EFV and EFV-RTV. This reveals that the PPG4ER has great potential to target the virus harbors tissues and improve bioavailability.

Pharmacogenetic predictors of variability in efavirenz pharmacokinetics in an admixed Brazilian HIV cohort

To investigate the influence of pharmacogenetic polymorphisms on efavirenz (EFV) exposure and metabolism in HIV-infected Brazilians under treatment with EFV-containing antiretroviral (ART) regimens. HIV-positive adults (n =82) on stable ART regimens containing 600 mg EFV once daily for at least 6 months were recruited at 2 university hospitals. Blood samples collected at mid-dose interval were used to quantify the plasma concentrations of EFV (denoted [EFV]), its major metabolite 8-OH-EFV ([8-OH-EFV]) and [8-OH-EFV]/[EFV] metabolic ratio, and to genotype single nucleotide polymorphisms in CYP2B6 (rs3745274, c.516G > T; rs28399499, c.983 T > C) and ABCB1 (rs3842, c.4036G > A). CYP2B6 metabolic phenotypes were inferred from the CYP2B6 diplotypes. Linear regression modelling was applied to identify sociodemographic, clinical and pharmacogenetic predictors of [EFV] and [8-OH-EFV]/[EFV] metabolic ratio. Wide (50-fold) interindividual variation in [EFV], [8-OH-EFV] and [8-OH-EFV]/[EFV] was observed; 69.5% of participants had [EFV] within the nominal therapeutic range (1000-4000 ng/mL), while 19.5 and 11.0% had [EFV] below and above this range, respectively. Multiple regression modelling retained only CYP2B6 metabolic phenotypes or the combined rs3745274 and rs28399499 genotypes, as significant predictors of [EFV] and [8-OH-EFV]/[EFV]. EFV exposure and disposition varied widely among HIV-infected Brazilians under stable treatment with EFV-containing ART regimens. About 1/10 of the participants had [EFV] exceeding nominal supratherapeutic concentration (4000 ng/mL), but reported tolerance to the ARV regimens, while 1/5 of participants had nominal subtherapeutic [EFV] (<1000 ng/mL) but adequate virological response. Genotype for the 2 CYP2B6 single nucleotide polymorphisms studied explained 48% of variation in [EFV] and 35% of variation in [8-OH-EFV]/[EFV].

Efavirenz Biotransformation in Brain Microsomes and Neural Cells

Efavirenz (EFV) is a non‐nucleoside reverse transcriptase inhibitor that is metabolized by both the cytochromes P450 and UDP‐glucuronosyltransferases (UGTs). EFV is one of several anti‐HIV medications that can cross the blood‐brain barrier and is essential to combatting the viral reservoir in the brain that can eventually lead to HIV‐associated neurocognitive disorder (HAND) in nearly 50% of patients. EFV and 8‐hydroxyefavirenz (8‐OHEFV), the primary P450‐dependent metabolite of EFV, have both been linked to neurological adverse events and neurotoxicity. However, very little is known about the fate of EFV in the brain, particularly how it’s metabolized or the precise mechanism of neurotoxicity. To investigate EFV biotransformation in the brain, we first performed microsomal metabolism assays with brain microsomes obtained from mice, cynomolgus macaques, and humans. EFV was incubated with the brain microsomes and NADPH to characterize P450‐dependent metabolism, while EFV or one of its P450‐dependent metabolites, 8‐OHEFV or 8‐,14‐dihydroxyefavirenz (8,14‐diOHEFV), was incubated with brain microsomes and UDPGA to investigate UGT‐catalyzed metabolism. Ultra‐high‐performance liquid chromatography tandem high resolution mass spectrometry (UHPLC‐HRMS) was employed to detect metabolites. Formation of 8‐OHEFV was observed in all three species, as was glucuronidation of 8‐OHEFV and 8,14‐diOHEFV. Interestingly, direct glucuronidation of EFV was only found in the cynomolgus macaque brain microsomes. When comparing brain microsome metabolism to liver microsome metabolism, we see that the liver has a greater capacity to transform EFV and its P450‐dependent metabolites. We tested cell‐type specific metabolism using primary cortical neurons, striatal neurons, astrocytes, and microglia from mice. Cells were cultured and treated with EFV, 8‐OHEFV, or 8,14‐diOHEFV for approximately 24 hours, with DMSO as a vehicle control. In the microglia and astrocytes, we observed P450‐mediated formation of 8‐OHEFV in the EFV‐treated cells. In the cortical neurons, striatal neurons, and astrocytes, both 8‐OHEFV and 8,14‐diOHEFV were glucuronidated. These results shed additional light on the microsome data, indicating which cell‐types may be contributing to EFV metabolism. In male mice dosed with EFV for 5 days in drinking water, UHPLC‐HRMS analysis of brain tissue homogenate and serum revealed 8‐OHEFV in brain tissue in conjunction with serum concentrations in the range of 15‐20 ng/mL. EFV and other metabolites were below the limit of detection in both brain tissue and serum. These data indicate that 8‐OHEFV has the potential to be formed and/or further metabolized in mouse brain. Additionally, inter‐species differences in EFV glucuronidation were observed in brain microsomes. The characterization of EFV metabolism in the brain improves our understanding of tissue‐specific EFV disposition as well as brain glucuronidation more broadly.

The role of CYP2B6 516G>T polymorphism on efavirenz/nevirapine toxicity. Implications on treatment outcomes: Lessons from Botswana.

The two non-nucleoside reverse transcriptase inhibitors (NNRTIs), efavirenz (EFV) and nevirapine (NVP), are currently the core antiretroviral drugs for treatment of HIV in sub-Saharan Africa including Botswana. The drugs are metabolized by Cytochrome P450 2B6 (CYP2B6) liver enzyme. The CYP2B6 gene that encodes for metabolism of these drugs is known to be highly polymorphic. One of the polymorphism in the CYP2B6 gene, 516G>T, particularly the 516T allele, is known to confer poor metabolism of EFV and NVP. This may lead to high levels of plasma drug concentrations and development of treatment toxicities, like central nervous system toxicities, and cutaneous and hepatic toxicities, for EFV and NVP, respectively. The CYP2B6 516G allele on the other hand is associated with an extensive metabolism of the two NNRTIs drugs. We sought to establish association between possible developments of NNRTIs toxicities with CYP2B6 516G>T variation in Botswana.A total of 316 peripheral blood mononuclear cells samples were used in a retrospective view. All the samples were from participants on EFV/NVP-containing regimen with known toxicity output. TaqMan Real-Time PCR approach was applied for assessing CYP2B6 516 allele variation in cases with treatment toxicity and those without. Analysis was performed by chi-square statistics and logistic regression analysis.The rate of poor metabolizers among participants with toxicity and those without toxicity was 18.4% and 15.1%, respectively. The CYP2B6 516 genotype distribution comparisons between the participants with toxicity and those without were not statistically different (chi-square = .326; P = .568).CYP2B6 516 variation was not associated with NNRTI toxicity. No other factors were associated with toxicity when considering age, baseline body mass index, baseline CD4, baseline HIV viral load and adherence. The results were discussed in the context of all the studies done in Botswana to date.

Influence of SULT1A1*2 Polymorphism on Plasma Efavirenz Concentration in Thai HIV-1 Patients.

PurposePlasma efavirenz (EFV) concentrations within therapeutic levels are essential to successfully treat patients suffering from human immunodeficiency virus (HIV) type 1. In addition to the drug-metabolizing enzyme CYP2B6, other phase II drug-metabolizing enzymes and transporters may have an important role in the pharmacokinetics of EFV. Thus, the influence of phase II drug-metabolizing enzymes and drug transporters on plasma EFV levels was investigated in Thai HIV patients receiving EFV.Patients and MethodsGenotyping was performed by TaqMan® real-time PCR in 149 HIV-infected Thai adults, and plasma efavirenz concentration was measured by a validated high-performance liquid chromatography in 12 hours after dosing steady-state plasma samples at week 12 and 24.ResultsPatients with three or more copies of SULT1A1 had significantly lower median plasma EFV concentrations than those carrying two copies at week 12 (p=0.046) and SULT1A1*2 (c.638G>A) carriers had significantly lower median plasma EFV concentrations compared to those not carrying the variant at week 24 (p=0.048). However, no significant association was found after adjusting for CYP2B6 genotype.ConclusionGenetic variation in a combination of SULT1A1*2 and SULT1A1 copy number may contribute to variability in EFV metabolism and thereby may impact drug response. The influence of a combination between the SULT1A1 and CYP2B6 genotype on EFV pharmacokinetics should be further investigated in a larger study population.

Efavirenz Pharmacogenetics and Weight Gain Following Switch to Integrase Inhibitor-Containing Regimens.

Unwanted weight gain affects some people living with human immunodeficiency virus (HIV) who are prescribed integrase strand transfer inhibitors (INSTIs). Mechanisms and risk factors are incompletely understood. We utilized 2 cohorts to study pharmacogenetics of weight gain following switch from efavirenz- to INSTI-based regimens. In an observational cohort, we studied weight gain at 48 weeks following switch from efavirenz- to INSTI-based regimens among patients who had been virologically suppressed for at least 2 years at a clinic in the United States. Associations were characterized with CYP2B6 and UGT1A1 genotypes that affect efavirenz and INSTI metabolism, respectively. In a clinical trials cohort, we studied weight gain at 48 weeks among treatment-naive participants who were randomized to receive efavirenz-containing regimens in AIDS Clinical Trials Group studies A5095, A5142, and A5202 and did not receive INSTIs. In the observational cohort (n = 61), CYP2B6 slow metabolizers had greater weight gain after switch (P = .01). This was seen following switch to elvitegravir or raltegravir, but not dolutegravir. UGT1A1 genotype was not associated with weight gain. In the clinical trials cohort (n = 462), CYP2B6 slow metabolizers had lesser weight gain at week 48 among participants receiving efavirenz with tenofovir disoproxil fumarate (P = .001), but not those receiving efavirenz with abacavir (P = .65). Findings were consistent when stratified by race/ethnicity and by sex. Among patients who switched from efavirenz- to INSTI-based therapy, CYP2B6 genotype was associated with weight gain, possibly reflecting withdrawal of the inhibitory effect of higher efavirenz concentrations on weight gain. The difference by concomitant nucleoside analogue is unexplained.

Endogenous Hormones and Antiretroviral Exposure in Plasma, Cervicovaginal Fluid, and Upper-Layer Packed Cells of Malawian Women Living with HIV.

Overlap in metabolism pathways of endogenous female sex hormones and antiretroviral drugs may lead to altered exposure to these compounds. In a family planning clinic in Lilongwe, Malawi, blood, blood cell, and cervicovaginal fluid (CVF) samples from seventy-three HIV positive Malawian women taken in follicular and luteal menstrual phases were assessed for estradiol and progesterone by chemiluminescent immunoassay, and for antiretroviral concentration by liquid chromatography-mass spectrometry. In both follicular and luteal phases, estradiol concentrations were lower in women receiving efavirenz compared with women on non-efavirenz regimens or no antiretroviral therapy (p < .01). Serum estradiol was moderately and negatively correlated with efavirenz plasma (r = −0.36, p < .001) and CVF (r = −0.50, p < .001) concentrations. Serum estradiol was a significant predictor of efavirenz CVF concentrations even after adjusting for efavirenz plasma concentrations (p = .02). In upper-layer packed cells (ULPCs), tenofovir diphosphate (TFVdp) concentrations were similar between follicular and luteal phases and were not correlated with estradiol or progesterone concentrations. Tenofovir concentrations in CVF were not associated with menstrual cycle or serum hormone concentrations. In CVF and plasma, efavirenz concentrations were negatively correlated with serum estradiol concentrations, suggesting a modulatory effect of estradiol on efavirenz metabolism and/or transport processes, and/or an effect of efavirenz on the metabolism of estradiol. Differences in CVF persisted even after adjusting for plasma concentrations, suggesting a mechanism specific to the female genital compartment separate from absorption or hepatic metabolism. In contrast, TFVdp concentrations in ULPC were not influenced by endogenous estradiol or progesterone concentrations.

The Pharmacogenetics of Efavirenz Metabolism in Children: The Potential Genetic and Medical Contributions to Child Development in the Context of Long-Term ARV Treatment.

Efavirenz (EFV) is a well-known, effective anti-retroviral drug long used in first-line treatment for children and adults with HIV and HIV/AIDS. Due to its narrow window of effective concentrations, between 1 and 4 μg/mL, and neurological side effects at supratherapeutic levels, several investigations into the pharmacokinetics of the drug and its genetic underpinnings have been carried out, primarily with adult samples. A number of studies, however, have examined the genetic influences on the metabolism of EFV in children. Their primary goal has been to shed light on issues of appropriate pediatric dosing, as well as the manifestation of neurotoxic effects of EFV in some children. Although EFV is currently being phased out of use for the treatment of both adults and children, we share this line of research to highlight an important aspect of medical treatment that is relevant to understanding the development of children diagnosed with HIV.

Efavirenz Metabolism: Influence of Polymorphic CYP2B6 Variants and Stereochemistry.

Efavirenz (more specifically the S-enantiomer) is a cornerstone antiretroviral therapy for treatment of HIV infection. The major primary metabolite is S-8-hydroxyefavirenz, which does not have antiretroviral activity but is neurotoxic. Cytochrome P450 2B6 (CYP2B6) is the major enzyme catalyzing S-8-hydroxyefavirenz formation. CYP2B6 genetics and drug interactions are major determinants of clinical efavirenz disposition and dose adjustment. In addition, as a prototypic CYP2B6 substrate, S-efavirenz and analogs can inform on the structure, activity, catalytic mechanisms, and stereoselectivity of CYP2B6. Metabolism of R-efavirenz by CYP2B6 remains unexplored. This investigation assessed S-efavirenz metabolism by clinically relevant CYP2B6 genetic variants. This investigation also evaluated R-efavirenz hydroxylation by wild-type CYP2B6.1 and CYP2B6 variants. S-Efavirenz 8-hydroxylation by wild-type CYP2B6.1 and variants exhibited positive cooperativity and apparent cooperative substrate inhibition. On the basis of Clmax values, relative activities for S-efavirenz 8-hydroxylation were in the order CYP2B6.4 > CYP2B6.1 ≈ CYP2B6.5 ≈ CYP2B6.17 > CYP2B6.6 ≈ CYP2B6.7 ≈ CYP2B6.9 ≈ CYP2B6.19 ≈ CYP2B6.26; CYP2B6.16 and CYP2B6.18 showed minimal activity. Rates of R-efavirenz metabolism were approximately 1/10 those of S-efavirenz for wild-type CYP2B6.1 and variants. On the basis of Clmax values, there was 14-fold enantioselectivity (S > R-efavirenz) for wild-type CYP2B6.1, and 5- to 22-fold differences for other CYP2B6 variants. These results show that both CYP2B6 516G > T (CYP2B6*6 and CYP2B6*9) and 983T > C (CYP2B6*16 and CYP2B6*18) polymorphisms cause canonical diminishment or loss-of-function variants for S-efavirenz 8-hydroxylation, provide a mechanistic basis for known clinical pharmacogenetic differences in efavirenz disposition, and may predict additional clinically important variant alleles. Efavirenz is the most stereoselective CYP2B6 drug substrate yet identified and may be a useful probe for the CYP2B6 active site and catalytic mechanisms. SIGNIFICANCE STATEMENT: Clinical disposition of the antiretroviral S-efavirenz is affected by CYP2B6 polymorphisms. Expressed CYP2B6 with 516G>T (CYP2B6*6 and CYP2B6*9), and 983T>C (CYP2B6*16 and CYP2B6*18) polymorphisms had a diminishment or loss of function for efavirenz 8-hydroxylation. This provides a mechanistic basis for efavirenz clinical pharmacogenetics and may predict additional clinically important variant alleles. Efavirenz metabolism showed both cooperativity and cooperative substrate inhibition. With greater than 10-fold enantioselectivity (S- vs. R- metabolism), efavirenz is the most stereoselective CYP2B6 drug substrate yet identified. These findings may provide mechanistic insights.

Impact of Efavirenz Metabolism on Loss to Care in Older HIV+ Africans.

We conducted a prospective cohort study of 914 treatment-naïve HIV+ adults initiating efavirenz-based antiretroviral treatment at public HIV clinics in Gaborone, Botswana between 2009 and 2013. Older age, defined as age ≥ 50years, was the primary exposure and loss to care at 6months was the primary outcome. Interaction between age and CYP2B6 516G>T and 983T>C polymorphisms, defined as extensive, intermediate, and slow metabolism, was assessed. Neurocognitive toxicity was measured using a symptom questionnaire. Age-stratified logistic regression was performed to identify factors associated with loss to care. Older age was associated with loss to care (OR 1.95, 95% CI 1.30-2.92). Age modified the effect of CYP2B6 genotype on loss to care with older, slow metabolizers at over four-fold higher risk when compared to older, intermediate metabolizers (OR 4.06 95% CI 1.38-11.89); neurocognitive toxicity did not mediate this risk. CYP2B6 metabolism genotype did not increase risk of loss to care in younger participants. Older age was associated with loss to care, especially among those with slow efavirenz metabolism. Understanding the relationship between older age and CYP2B6 genotype will be important to improving outcomes in an aging population initiating efavirenz-based ART in similar settings.

CYP2B6 Genetic Polymorphisms, Depression, and Viral Suppression in Adults Living with HIV Initiating Efavirenz-Containing Antiretroviral Therapy Regimens in Uganda: Pooled Analysis of Two Prospective Studies.

Single-nucleotide polymorphisms (SNPs) in CYP2B6 have been shown to predict variation in plasma efavirenz concentrations, but associations between these SNPs and efavirenz-mediated depression and viral suppression are less well described. We evaluated three SNPs in CYP2B6 (rs3745274, rs28399499, and rs4803419) in Ugandan persons living with HIV. To define exposure, we used previously published pharmacokinetic modeling data to categorize participants as normal, intermediate, and poor efavirenz metabolizers. Our outcomes were probable depression in the first 2 years after antiretroviral therapy (ART) initiation (mean score of >1.75 on the Hopkins Symptom Depression Checklist) and viral suppression 6 months after ART initiation. We fit generalized estimating equation and modified Poisson regression models adjusted for demographic, clinical, and psychosocial characteristics with or without individuals with depression at the time of ART initiation. Among 242 participants, there were no differences in the pre-ART depression or viral load by efavirenz metabolism strata (p > .05). Participants were classified as normal (32%), intermediate (50%), and poor (18%) metabolizers. Seven percent (56/242) of follow-up visits met criteria for depression. Eighty-five percent (167/202) of participants who completed a 6-month visit achieved viral suppression. CYP2B6 metabolizer strata did not have a statistically significant association with either depression [adjusted risk ratio (aRR) comparing intermediate or poor vs. normal, 1.46; 95% confidence interval (CI), 0.72-2.95] or 6-month viral suppression (aRR, 1.01; 95% CI, 0.88-1.15). However, in analyses restricted to participants without pre-ART depression, poorer CYP2B6 metabolism was associated with increased odds of depression (adjusted odds ratio, 4.11; 95% CI, 1.04-16.20). Efavirenz-metabolizing allele patterns are strongly associated with risk of incident depression. Future work should elucidate further region-specific gene-environment interactions and whether alternate polymorphisms may be associated with efavirenz metabolism.

Drug metabolism and transport gene polymorphisms and efavirenz adverse effects in Brazilian HIV-positive individuals.

There are limited data regarding efavirenz pharmacogenetics in admixed populations. The Brazilian population is highly admixed. In a Brazilian cohort, we sought to characterize associations between efavirenz adverse effects (all-cause and CNS) and polymorphisms in seven genes known or suspected to affect efavirenz metabolism and transport. We studied 225 HIV-positive individuals who had been prescribed efavirenz-containing regimens at a hospital in Rio de Janeiro, Brazil. Eighty-nine cases had efavirenz adverse effects, including 43 with CNS adverse effects, while 136 controls had no adverse effect of any antiretroviral after treatment for at least 6 months. A total of 67 candidate polymorphisms in ABCB1, CYP2A6, CYP2B6, CYP3A4, CYP3A5, NR1I2 and NR1I3 genes were selected for association analysis. Admixture was assessed using 28 ancestry-informative polymorphisms previously validated for the Brazilian population. Associations were evaluated with logistic regression models adjusted for sex and genetic ancestry. There was extensive African, European and Native American admixture in the cohort. Increased all-cause adverse effects were associated with the CYP2B6 genotype combination 15582CC-516TT-983TT (OR = 7.26, P = 0.003) and with the CYP2B6 slow metabolizer group 516TT or 516GT-983CT (OR = 3.10, P = 0.04). CNS adverse effects were nominally associated with CYP3A4 rs4646437 (OR = 4.63, P = 0.014), but not after adjusting for multiple comparisons. In a highly admixed Brazilian cohort, the CYP2B6 slow metabolizer genotype was associated with an increased risk of efavirenz adverse effects.

Polymorphisms in cytochrome P450 are associated with extensive efavirenz pharmacokinetics and CNS toxicities in an HIV cohort in Botswana.

Inter-individual variability in efavirenz (EFV) pharmacokinetics and dynamics are dominantly driven by the polymorphism in cytochrome P450 (CYP) isoenzyme 2B6 516G>T. We hypothesized that additional CYP polymorphisms mediate the relationship between CYP2B6 516G>T, EFV metabolism, and clinical events. We investigated 21 SNPs in 814 HIV-infected adults initiating EFV-based therapy in Botswana for population pharmacokinetics, CNS toxicities, and treatment outcomes. Two SNPs (rs28399499 and rs28399433) showed reduced apparent oral EFV clearance. Four SNPs (rs2279345, rs4803417, rs4802101, and rs61663607) showed extensive clearance. Composite CYP2B-mediated EFV metabolism was significantly associated with CNS toxicity (p=0.04), with extensive metabolizers reporting more and slow and very slow metabolizers reporting less toxicity after one month compared to intermediate metabolizers. Composite CYP2B6 metabolism was not associated with composite early treatment failure. In conclusion, our data suggest that CNS-related toxicities might not be solely the result of super-therapeutic parent EFV concentrations in HIV-infected individuals in patients of African ancestry.

Efavirenz-metabolizing polymorphisms, viral suppression, and depression in HIV-infected individuals initiating antiretroviral therapy in southwestern Uganda

BackgroundSingle-nucleotide polymorphisms (SNPs) in CYP2B6 have previously been associated with a 10-fold range in trough plasma efavirenz concentrations, but associations between these SNPs and efavirenz (EFV)-mediated viral suppression and tolerability remain unclear.MethodsWe evaluated three SNPs in CYP2B6 (rs3745274, rs28399499, and rs4803419, Illumina OmniExpress) among HIV-infected Ugandans observed in a cohort study every 3–4 months from 2005–2015. Genotypes from these SNPs were used to group participants into previously described pharmacokinetic strata: extensive (EXT), intermediate (INT), and slow metabolizers (Figure 1). The primary outcomes were viral suppression, defined by an undetectable viral load in the first measurement a minimum of three months after ART initiation, and incident depression in the first two years, defined by a mean score >1.75 on the Hopkins Symptom Checklist. We fitted standard and generalized estimating equations (GEE) logistic regression models for viral suppression and depression, respectively. Models were adjusted for clinical and demographic covariates that reached a significance of P < 0.25 in unadjusted models.ResultsAmong 103 participants with genotyping, there were no differences in pre-ART viral load or depression by metabolism strata (P > 0.5). Minor allele frequencies for rs3745274, rs28399499, and rs4803419 were 33%, 7%, and 4%, respectively. Approximately 79%, 78%, and 94% of participants were suppressed at their first viral load measurement in the extensive, intermediate, and slow metabolizer strata, respectively (Figure 2; P = 0.35). In adjusted models, metabolism strata were not associated with viral suppression (AORINT 0.81, 95% CI 0.26–2.56; AORSLOW 3.92, 95% CI 0.39–39.40) or with depression (AORINT 1.95, 95% CI 0.75–5.09; AORSLOW 0.72, 95% CI 0.17–3.02; Table).ConclusionWe did not identify an association between efavirenz-metabolizing polymorphisms and viral suppression or depression in a cohort of HIV-infected individuals initiating ART in southwestern Uganda. Future work should reassess these relationships with larger samples and longer-term outcomes and explore additional polymorphisms that may be associated with efavirenz metabolism in this population.Disclosures P. Hunt, Merck: Consultant, Consulting fee; Gilead: Consultant, Consulting fee; Viiv: Consultant, Consulting fee

CYP2B6 genotypes and early efavirenz-based HIV treatment outcomes in Botswana.

To determine the association between cytochrome p450 2B6 genotypes and efavirenz-based HIV treatment outcomes. Observational cohort study of HIV-infected adults initiating efavirenz-based regimens in Botswana. The primary endpoint was a composite of death or loss to care or HIV RNA more than 25 copies/ml at 6 months. CYP2B6 516G>T and 983T>C genotyping was done with Taqman Open Array platform. Adverse experiences were measured by using the Subject Experience Questionnaire. Metabolism alleles were included in logistic regression models of the composite endpoint. A total of 801 individuals included 406 (51%) men, median age 37 years, median baseline CD4 cell count 195 cells/μl, and plasma HIV RNA 4.9 log10 copies/ml. 288 (36%) reached the endpoint, including 34 (4%) deaths, 151 (19%) lost to care, 11 (1%) lost to the study, but alive and in care, and 92 (11%) with plasma HIV RNA more than 25 copies/ml. Metabolism variant alleles were common with 396 (49%) intermediate and 192 (24%) slow metabolizers. There were no statistically significant associations between metabolism and treatment endpoints. However, slower metabolism was associated with fewer adverse experiences. Slow metabolism alleles were associated with lower efavirenz clearance but not any of the treatment endpoints. Slow efavirenz metabolism did not exacerbate central nervous system toxicity. These results should allay concern that slow efavirenz metabolism adversely impacts individuals in sub-Saharan African settings in which these alleles are common.

Population Approach to Efavirenz Therapy

Efavirenz (EFV) is a nonnucleoside reverse transcriptase inhibitor commonly used as first-line therapy in the treatment of human immunodeficiency virus (HIV), with a narrow therapeutic range and a high between-subject variability which can lead to central nervous system toxicity or therapeutic failure. To characterize the sources of variability and better predict EFV steady-state plasma concentrations, a population pharmacokinetic model was developed from 96 HIV-positive individuals, using a nonlinear mixed-effect method with Monolix® software. A one-compartment with first-order absorption and elimination model adequately described the data. To explain between-subject variability, demographic characteristics, biochemical parameters, hepatitis C virus–HIV coinfection, and genetic polymorphisms were tested. A combination of the single-nucleotide polymorphisms rs2279343 and rs3745274, both in the CYP2B6 gene, were the only covariates influencing clearance, included in the final model. Oral clearance was estimated to be 19.6 L/h, 14.15 L/h, and 6.08 L/h for wild-type, heterozygous mutated and homozygous mutated individuals, respectively. These results are in accordance with the current knowledge of EFV metabolism and also suggest that in homozygous mutated individuals, a dose adjustment is necessary. Hepatitis C virus–HIV coinfection does not seem to be a predictive indicator of EFV pharmacokinetic disposition.