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Objectives: We evaluated whether dried blood spots (DBS) are suitable to monitor combined ART when samples are collected in rural Tanzania and transported over a long distance to a specialized bioanalytical laboratory., Methods: Plasma and DBS samples were collected in Tanzania from study patients treated with nevirapine, efavirenz or lopinavir. In addition, plasma, whole blood and DBS samples were obtained from a cohort of HIV patients at the site of the bioanalytical laboratory in Switzerland. DBS samples were analysed using a fully automated LC-MS/MS method., Results: Comparison of DBS versus plasma concentrations of samples obtained from the bridging study in Switzerland indicated an acceptable bias only for nevirapine (18.4%), whereas for efavirenz and lopinavir a pronounced difference of -47.4% and -48.1% was found, respectively. Adjusting the DBS concentrations by the haematocrit and the fraction of drug bound to plasma proteins removed this bias [efavirenz +9.4% (-6.9% to +25.7%), lopinavir +2.2% (-20.0% to +24.2%)]. Storage and transportation of samples from Tanzania to Switzerland did not affect the good agreement between plasma and DBS for nevirapine [-2.9% (-34.7% to +29.0%)] and efavirenz [-9.6% (-42.9% to +23.8%)]. For lopinavir, however, adjusted DBS concentrations remained considerably below [-32.8% (-70.4% to +4.8%)] corresponding plasma concentrations due to decay of lopinavir in DBS obtained under field conditions., Conclusions: Our field study shows that the DBS technique is a suitable tool for therapeutic drug monitoring in resource-poor regions; however, sample stability remains an issue for certain analytes and therefore needs special consideration.

Background: Newborns of HIV-infected mothers are given daily doses of nevirapine to prevent HIV-1 acquisition. Infants born to mothers with TB should also receive TB preventive therapy. TB preventive regimens include isoniazid for 6 months or rifampicin plus isoniazid for 3 months (RH preventive therapy). The effect of concomitant RH preventive therapy on nevirapine concentrations in infants is unknown., Patients and Methods: Tshepiso was a prospective case-control cohort study of pregnant HIV-infected women with and without TB whose newborn infants received standard doses of nevirapine for HIV prophylaxis. Infants born to mothers with TB also received RH preventive therapy. Infant plasma nevirapine concentrations were measured at 1 and 6 weeks. The effects of RH preventive therapy on nevirapine disposition were investigated in a population pharmacokinetic model., Results: Of 164 infants undergoing pharmacokinetic sampling, 46 received RH preventive therapy. After adjusting for weight using allometric scaling, the model estimated a 33% reduction in nevirapine trough concentrations with RH preventive therapy compared with TB-unexposed infants not receiving concomitant rifampicin and a 30% decline in trough concentrations in a typical infant between day 7 and 35 post-partum., Conclusions: Rifampicin-based TB preventative treatment reduces nevirapine concentrations significantly in HIV-exposed infants. Although the nevirapine exposures required to prevent HIV acquisition in breastfeeding infants are undefined, given the potential risks associated with underdosing nevirapine in this setting, it is prudent to avoid rifampicin-based preventive therapy in HIV-exposed children receiving prophylactic nevirapine., (© The Author 2017. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Objectives: Nevirapine is widely used for the treatment of HIV-1 infection; however, its chronic use has been associated with severe liver and skin toxicity. Women are at increased risk for these toxic events, but the reasons for the sex-related differences are unclear. Disparities in the biotransformation of nevirapine and the generation of toxic metabolites between men and women might be the underlying cause. The present work aimed to explore sex differences in nevirapine biotransformation as a potential factor in nevirapine-induced toxicity., Methods: All included subjects were adults who had been receiving 400 mg of nevirapine once daily for at least 1 month. Blood samples were collected and the levels of nevirapine and its phase I metabolites were quantified by HPLC. Anthropometric and clinical data, and nevirapine metabolite profiles, were assessed for sex-related differences., Results: A total of 52 patients were included (63% were men). Body weight was lower in women (P = 0.028) and female sex was associated with higher alkaline phosphatase (P = 0.036) and lactate dehydrogenase (P = 0.037) levels. The plasma concentrations of nevirapine (P = 0.030) and the metabolite 3-hydroxy-nevirapine (P = 0.035), as well as the proportions of the metabolites 12-hydroxy-nevirapine (P = 0.037) and 3-hydroxy-nevirapine (P = 0.001), were higher in women, when adjusted for body weight., Conclusions: There was a sex-dependent variation in nevirapine biotransformation, particularly in the generation of the 12-hydroxy-nevirapine and 3-hydroxy-nevirapine metabolites. These data are consistent with the sex-dependent formation of toxic reactive metabolites, which may contribute to the sex-dependent dimorphic profile of nevirapine toxicity.

Background: Nevirapine is an important component of paediatric combination HIV therapy. Adequate drug exposure is necessary in order to achieve long-lasting viral suppression., Objectives: To study the influence of age, drug dose and formulation type, nutritional status and CYP2B6 516G>T polymorphism on blood concentrations of nevirapine in children treated with generic antiretroviral drugs., Methods: A multicentre study was conducted at four sites in India. HIV-infected children receiving generic nevirapine-based fixed-dose combinations were recruited. Trough and 2 h nevirapine plasma concentrations were determined by HPLC. Characterization of the CYP2B6 gene polymorphism was performed using direct sequencing. Clinical and nutritional status was recorded. Groups were compared using the Mann-Whitney U-test and multivariable logistic regression analysis was performed to identify factors contributing to low drug levels., Results: Ninety-four children of median age 78 months were studied; 60% were undernourished or stunted. Stunted children had a significantly lower 2 h nevirapine concentration compared with non-stunted children (P < 0.05); there were no significant differences in trough concentrations between different nutritional groups. Nevirapine levels were significantly higher in children with TT compared with GG and GT CYP2B6 genotypes (P < 0.01). Children ≤ 3 years had a 3.2 (95% confidence interval 1.07-9.45) times higher risk of having sub-therapeutic nevirapine concentrations., Conclusions: Nevirapine blood concentrations are affected by many factors, most notably age ≤ 3 years; a combination of young age, stunting and CYP2B6 GG or GT genotype could potentially result in sub-therapeutic nevirapine concentrations. Dosing recommendations for children should be reviewed in the light of these findings.

Here we describe a simple, stable, and specific gas chromatography-mass spectrometry (GC-MS) method for the determination of nevirapine in plasma. After precipitation of proteins, the non-nucleoside reverse transcriptase inhibitor nevirapine was extracted with dichloromethane. For the determination and quantification of nevirapine, 1 microL of the organic layer was injected onto the GC-MS system. Linear calibration curves were obtained with BIRH 0414BS as internal standard in a range from 0.01 to 15 microg/mL. Intra- and inter-day accuracy and precision of this method were good with an accuracy between 96-109% and a precision between 2-8% across the therapeutic range of nevirapine. GC-MS proved to be a valid alternative to high-performance liquid chromatography and liquid chromatography-MS.

A bioanalytical method for determination of lamivudine (3TC), zidovudine (AZT), and nevirapine (NVP) in 100 microL capillary blood applied onto sampling paper has been developed and validated. The antiretroviral drugs (ARV) were analyzed by reversed phase gradient liquid chromatography with UV detection. Separation was performed on a Zorbax SB C(8) (250 x 4.6 mm) column with a two-step gradient: (i) methanol-0.05 mol/L acetic acid-sodium acetate buffer (pH 3.95, 15:85 v/v) and (ii) methanol-0.05 mol/L acetic acid-sodium acetate buffer (pH 3.95, 50:50 v/v) with a flow rate of 1.0 mL/min. UV detection was performed at 260 nm. Total assay precisions were 6.3, 4.7, and 4.9% for 3TC at 0.34, 0.69, and 3.9 microg/mL, and 5.1, 5.5, and 3.2% for AZT at 0.40, 0.80, and 4.5 microg/mL. For NVP, total assay precisions were 5.2, 8.3, and 3.5% at 2.6, 4.5, and 8.8 microg/mL. Lower limit of quantifications (LLOQ) were 0.11 and 0.13 microg/mL for 3TC and AZT where the precisions were 2.0% for both the analytes. For NVP, LLOQ was 1.3 microg/mL where precision was 2.6%. Concentrations were determined for 10 h for two subjects receiving standard twice daily antiretroviral therapy containing 3TC, AZT, and NVP. Maximum 3TC concentrations were 2.5 and 2.8 microg/mL for subject 1 and 2, respectively. For AZT, maximum concentrations were 1.8 and 1.1 microg/mL while being 15 and 9.6 microg/mL for NVP. Pre-dose trough concentration of NVP was 11 microg/mL for subject 1 and 9.6 microg/mL for subject 2.

Background: Polymorphisms in CYP2B6 affect the steady-state plasma concentrations of nevirapine and efavirenz. In many resource-limited countries, a single dose of nevirapine has been widely prescribed to pregnant women at delivery, to reduce mother-to-child transmission of human immunodeficiency virus type 1 (HIV-1). We characterized associations between genetic polymorphisms and the pharmacokinetics of single doses of nevirapine and efavirenz., Methods: Plasma drug concentrations were determined over the 13-day period after administration of a 200-mg oral dose of nevirapine to nonpregnant, HIV-negative African Americans. A 600-mg oral dose of efavirenz was subsequently administered, and pharmacokinetic sampling was repeated. Pharmacokinetic parameters were estimated using a noncompartmental approach. Primary analyses involved 2 CYP2B6 polymorphisms (516G --> T and 983T --> C) known to predict increased steady-state plasma nevirapine and efavirenz exposure. Exploratory analyses involved another 51 polymorphisms in CYP2B6, ABCB1, CYP3A4, and CYP3A5., Results: On the basis of the composite CYP2B6 516/983 genotype, the 34 participants comprised 10 extensive, 17 intermediate, and 7 slow metabolizer genotypes. The composite CYP2B6 516/983 genotype was significantly associated with plasma drug exposure and clearance for efavirenz but not nevirapine. Exploratory analyses suggested possible associations between additional CYP2B6 polymorphisms and the pharmacokinetics of nevirapine and efavirenz., Conclusions: Selective pressure for drug-resistant HIV-1 after administration of single-dose nevirapine may not differ substantially according to CYP2B6 516/983 genotype. Additional polymorphisms, genes, and populations warrant further study.

Background: Most HPLC-UV methods for therapeutic drug monitoring of anti-HIV drugs have long run times, which reduce their applicability for high-throughput analysis. We developed an ultra-performance liquid chromatography (UPLC)-diode array detection method for the simultaneous quantification of the HIV-protease inhibitors (PIs) amprenavir, atazanavir, indinavir, lopinavir, nelfinavir, ritonavir, saquinavir, and tipranavir (TPV), and the nonnucleoside reverse transcriptase inhibitors (NNRTIs) efavirenz and nevirapine., Methods: Solid-phase extraction of 1 mL plasma was performed with Waters HLB cartridges. After 3 wash steps, we eluted the drugs with methanol, evaporated the alcohol, and reconstituted the residue with 50 microL methanol. We injected a 4-microL volume into the UPLC system (Waters ACQUITY UPLC BEH C8 column maintained at 60 degrees C) and used a linear gradient of 50 mmol/L ammonium acetate and 50 mmol/L formic acid in water versus acetonitrile to achieve chromatographic separation of the drugs and internal standard (A-86093). Three wavelengths (215, 240, and 260 nm) were monitored., Results: All drugs were eluted within 15 min. Calibration curves with concentrations of 0.025-10 mg/L (1.875-75 mg/L for TPV) showed coefficients of determination (r(2)) between 0.993 and 0.999. The lower limits of quantification were well below the trough concentrations reported in the literature. Inter- and intraassay CVs and the deviations between the nominal and measured concentrations were <15%. The method was validated by successful participation in an international interlaboratory QC program., Conclusions: This method allows fast and simultaneous quantification of all commercially available PIs and NNRTIs for therapeutic drug monitoring.

Background: The optimum strategy for stopping treatment with drugs that have different half-lives in a combination regimen to minimize the risk of selecting drug-resistant viruses remains unknown. We evaluated drug concentrations in plasma, human immunodeficiency virus (HIV) load, and development of drug resistance after a planned treatment interruption of a nonnucleoside reverse-transcriptase inhibitor (NNRTI)-containing regimen in HIV type 1-infected children., Methods: Children with viral loads <50 copies/mL and CD4 cell percentages > or =30% (for children aged 2-6 years) or CD4 cell percentages > or =25% and CD4 cell counts > or =500 cells/microL (for children aged 7-15 years) were randomized to either a planned treatment interruption or to continuous therapy. In the planned treatment interruption arm, either (1) treatment with nevirapine or efavirenz was stopped, and treatment with the remaining drugs was continued for 7-14 days, or (2) nevirapine or efavirenz were replaced by a protease inhibitor, and all drugs were stopped after 7-14 days. Sampling for determination of plasma drug concentrations, measurement of viral load, and drug resistance testing was scheduled at day 0, day 7 (drug concentrations only), day 14, and day 28 after interruption of treatment with an NNRTI., Results: Treatment with an NNRTI was interrupted for 35 children (20 were receiving nevirapine, and 15 were receiving efavirenz). Median time from NNRTI cessation to stopping all drugs was 9 days (range, 6-15 days) for nevirapine and 14 days (range, 6-18 days) for efavirenz. At 7 days, 1 (5%) of 19 and 4 (50%) of 8 children had detectable nevirapine and efavirenz concentrations, respectively; efavirenz remained detectable in 3 (25%) of 12 children at 14 days. At 14 days, viral load was > or =50 copies/mL in 6 of 16 children interrupting treatment with nevirapine (range, 52-7000 copies/mL) and in 2 of 12 children interrupting treatment with efavirenz (range, 120-1600 copies/mL). No new NNRTI mutations were observed., Conclusions: In children with virological suppression who experienced interruption of treatment with an NNRTI, staggered or replacement stopping strategies for a median of 9 days for nevirapine and 14 days for efavirenz were not associated with the selection of NNRTI resistance mutations.

Objectives: The aim of this study was to investigate the frequency of CYP2B6 polymorphisms (according to ethnicity) and the influence of heterozygosity and homozygosity on plasma concentrations of efavirenz and nevirapine., Methods: Following written informed consent, 225 Caucasians and 146 Blacks were recruited from the German Competence Network for HIV/AIDS. Plasma concentrations of efavirenz and nevirapine were assessed by HPLC, and genotyping for 516G>T, 983T>C and 1459T>C polymorphisms in CYP2B6 was conducted by real-time PCR-based allelic discrimination., Results: The minor allele frequency for 516G>T, 983T>C and 1459T>C was 0.29, 0 and 0.08 in Caucasians and 0.34, 0.07 and 0.02 in Blacks, respectively. Two Black patients with the 983C allele receiving efavirenz were identified and both were withdrawn from therapy within 1 week of sampling due to toxicity. In multivariate analyses, efavirenz and nevirapine plasma concentrations were significantly associated with 983T>C (P < 0.0001 and P = 0.02, respectively), 516G>T (P < 0.0001 and P = 0.002, respectively) and time of drug analysis post-dose (P < 0.0001 for both). Body mass index was independently related to efavirenz (P = 0.04) but not nevirapine concentrations, and age was related to nevirapine (P = 0.05) but not efavirenz concentrations. Consistent with other studies, 1459C>T was not associated with plasma concentrations of either drug (P > 0.05 for both drugs)., Conclusions: This is the first report that the 983T>C genotype (part of the CYP2B6*18 haplotype) impacts on nevirapine plasma concentrations and the first study to assess the impact of 983C homozygosity on efavirenz concentrations. These data have implications for administration of non-nucleoside reverse transcriptase inhibitors to Black patients.

Background and Objectives: Nevirapine-containing antiretroviral therapy (ART) and rifampicin-based antitubercular therapy are commonly co-administered in Africa, where nevirapine is often the only available non-nucleoside reverse transcriptase inhibitor. Rifampicin induces the metabolism of nevirapine, but the extent of the reduction in nevirapine concentrations has varied widely in previous studies. We describe the steady-state pharmacokinetics of nevirapine during and after antitubercular therapy in South African patients., Methods: Sixteen patients receiving ART including standard doses of nevirapine were admitted twice for intensive pharmacokinetic sampling: during and after rifampicin-based antitubercular therapy., Results: Geometric mean ratios for nevirapine pharmacokinetic parameters during versus after antitubercular therapy were 0.61 [90% confidence interval (CI) 0.49-0.79] for Cmax, 0.64 (90% CI 0.52-0.80) for area under the curve up to 12 h (AUC(0-12)) and 0.68 (90% CI 0.53-0.86) for Cmin. Nevirapine Cmin was subtherapeutic (<3 mg/L) in six patients during antitubercular therapy (one of whom developed virological failure) and in none afterwards. There was no correlation between rifampicin concentrations and the degree of nevirapine induction assessed by the proportional change in nevirapine concentrations between the two admissions. The ratio of nevirapine AUC(0-12) to the AUC(0-12) of its 12-hydroxy metabolite was significantly lower in the presence of antitubercular therapy, consistent with induced metabolism., Conclusions: Nevirapine concentrations were significantly decreased by concomitant rifampicin-based antitubercular therapy and a high proportion of patients had subtherapeutic plasma concentrations. Further study in African patients is required to determine the implications for treatment outcomes.

Background: Most antiretrovirals are metabolized in the liver, and lower dosing could be advisable in patients with severe liver insufficiency., Methods: Plasma drug levels were measured in hepatitis C virus (HCV)/human immunodeficiency virus (HIV)-coinfected patients receiving nevirapine (NVP), efavirenz (EFV), lopinavir/ritonavir (LPV/r), or atazanavir (ATV) with or without ritonavir. Liver fibrosis was measured using elastometry., Results: A total of 268 coinfected patients with compensated liver disease were analyzed. Mean plasma levels were 6.1 micro g/mL for NVP (35 patients), 2.8 micro g/mL for EFV (46 patients), 5.8 micro g/mL for LPV (56 patients), 0.4 micro g/mL for ATV (58 patients), and 0.7 micro g/mL for ATV/r (73 patients). Overall, drug levels were higher in patients with cirrhosis than in those without cirrhosis for EFV (median, 3.4 vs. 1.9 micro g/mL; P<.01) and NVP (median, 6.6 vs. 5.8 micro g/mL; P=.33). EFV plasma levels above the toxic threshold (>4 micro g/mL) were more frequent in patients with cirrhosis than in those without (31% vs. 3%; P<.001). The same trend was seen for NVP levels >8 micro g/mL (50% vs. 27%; P=.27). By contrast, plasma levels of protease inhibitors (PIs) did not differ significantly between patients with and those without cirrhosis., Conclusions: Liver clearance of nonnucleoside reverse-transcriptase inhibitors, particularly EFV, is impaired in patients with cirrhosis. No similar effect is seen for PIs. Assessment of liver fibrosis by noninvasive tools may identify HCV/HIV-coinfected patients who might benefit from therapeutic drug monitoring to avoid drug overexposure.

Seventy human immunodeficiency virus (HIV)-infected patients receiving rifampicin and 70 HIV-infected patients not receiving rifampicin were enrolled to receive 400 mg of nevirapine-based highly active antiretroviral therapy per day. Mean plasma nevirapine levels at 8 and 12 weeks were lower in patients receiving rifampicin (P=.048). However, virological and immunological outcomes at 24 weeks were not different between the 2 groups (P>.05).

Background: The magnitude of infant antiretroviral (ARV) exposure from breast milk is unknown., Methods: We measured concentrations of nevirapine, lamivudine, and zidovudine in serum and whole breast milk from human immunodeficiency virus type 1 (HIV-1)-infected women in Botswana receiving ARV treatment and serum from their uninfected, breast-feeding infants., Results: Twenty mother-infant pairs were enrolled. Maternal serum concentrations of nevirapine were high (median, 9534 ng/mL at a median of 4 h after nevirapine ingestion). Median breast-milk concentrations of nevirapine, lamivudine, and zidovudine were 0.67, 3.34, and 3.21 times, respectively, those in maternal serum. The median infant serum concentration of nevirapine was 971 ng/mL, at least 40 times the 50% inhibitory concentration and similar to peak concentrations after a single 2-mg/kg dose of nevirapine. The median infant serum concentration of lamivudine was 28 ng/mL, and the median infant serum concentration of zidovudine was 123 ng/mL, but infants were also receiving zidovudine prophylaxis., Conclusions: HIV-1 inhibitory concentrations of nevirapine are achieved in breast-feeding infants of mothers receiving these ARVs, exposing infants to the potential for beneficial and adverse effects of nevirapine ingestion. Further study is needed to understand the impact of maternal ARV treatment on breast-feeding HIV-1 transmission, infant toxicity, and HIV-1 resistance mutations among infected infants.

We conducted a 12-week, multicenter, randomized, double-blind, placebo-controlled trial of cetirizine to assess the ability of antihistamines to prevent nevirapine-associated rash in patients infected with human immunodeficiency virus type 1. Patients initiating treatment with nevirapine were randomized to receive either cetirizine, 10 mg q.d. (104 patients), or placebo (96 patients) during the first 6 weeks of therapy. Rash occurred in 22 (11%) of 200 patients; 10 (9.6%) were in the cetirizine group and 12 (12.5%) were in the placebo group (odds ratio [OR], 0.75; 95% confidence interval [CI], 0.31-1.81; P=.5). Five of 22 rashes were cases of hypersensitivity syndrome. The rate of nevirapine discontinuation due to rash was similar in the 2 groups (7.7% and 6.25% in the cetirizine and placebo groups, respectively; P=.4). Multivariate analysis showed no treatment-group effect but indicated that age >40 years (OR, 3.83; 95% CI, 1.4-10.46; P=.008) was associated with an increased risk of rash. Cetirizine has no preventive effect on nevirapine-associated rash.

We compared use of a 3-class regimen (nevirapine [Nvp], stavudine [d4T], and indinavir [Idv; 1000 mg 3 times daily]) with use of a 2-class regimen (lamivudine [3TC], d4T, and Idv [800 mg 3 times daily]) for 145 patients infected with human immunodeficiency virus type 1 (HIV-1). At week 72, the plasma HIV-1 RNA level was undetectable in 52% of Nvp recipients versus 79% of 3TC recipients (P<.001). Idv trough levels were 81 ng/mL in the Nvp group and 99 ng/mL in the 3TC group (P=.012). In the Nvp group, 42.5% of patients discontinued the study regimen; in the 3TC group, 22.5% of patients discontinued therapy (P=.013). The rate of resistance to nonnucleoside analogue reverse-transcriptase inhibitors among patients in the Nvp group with virological failure was not different from the rate of resistance to 3TC among patients in the 3TC group with virological failure. These results do not support the use of a 3-class regimen that includes Nvp for patients with no or limited exposure to nucleoside analogues.