Short Communication: A Low-Cost Method for Analyzing Nevirapine Levels in Hair as a Marker of Adherence in Resource-Limited Settings

Abstract

The measurement of antiretroviral concentrations in hair is emerging as an important technology to objectively quantify adherence to combination antiretroviral therapy. Hair levels of antiretrovirals are the strongest independent predictor of virologic success in large prospective cohorts of HIV-infected patients and surpass self-report in predicting outcomes. Hair is easy to collect and store, but validated methods to analyze antiretroviral levels in hair using liquid chromatography tandem mass spectrometry (LC-MS/MS) are expensive. We report here on the development of a thin-layer chromatography (TLC) assay for the semiquantitative analysis of nevirapine in hair. TLC assay results from 11 samples were consistent with results using LC-MS/MS [Spearman correlation coefficient 0.99 (95% CI 0.95-0.996)]. This simple, low-cost method of analyzing nevirapine concentrations in hair may provide a novel monitoring tool for antiretroviral adherence in resource-limited settings and merits further study in clinical settings.A dherence to antiretroviral (ARV) therapy is essential for the realization of virologic, immunologic, and clinical benefits, but the limitations of self-report and other commonly used adherence measures are well-described.1 The measurement of ARV concentrations in hair as a biological measure of adherence is emerging as an important technology to objectively quantify adherence to combination antiretroviral therapy (cART).2 Hair levels of ARVs are the strongest independent predictor of virologic success in large prospective cohorts of HIV-infected patients, 3-6 and surpass self-report in predicting outcomes.3 Moreover, hair levels can serve as a measure of exposure to maternal ARVs in infants during pregnancy and breastfeeding. Since drugs accumulate in hair over prolonged periods, 8, 8a a single measurement of an ARV in a hair sample provides a longer-term measure of adherence than a single plasma level, which captures only short-term use.9-13 Nevirapine (NVP) is widely prescribed in cART regimens for the treatment of HIVinfected adults and children in global settings, 14 and plays a role in worldwide perinatal transmission prevention. A lowcost testing method for analyzing NVP levels in small hair samples that could be performed in local laboratories would therefore be useful for monitoring adherence to NVP-based regimens in resource-limited settings.Hair samples are simple and inexpensive to collect and can be stored at room temperature prior to analysis. Unlike phlebotomy, hair collection is noninvasive and does not require specific skills or sterile equipment. These features provide obvious cost and feasibility advantages for hair collection and storage, over plasma but validated methods of analyzing ARV levels in hair samples use liquid chromatography/tandem mass spectrometry (LC-MS/MS). 15,16 The LC-MS/MS method for analyzing NVP in hair samples is highly sensitive, 16 but the equipment required is expensive, limiting its suitability in resource-limited settings. In contrast, thin-laye...

Using Lopinavir Concentrations in Hair Samples to Assess Treatment Outcomes on Second-Line Regimens Among Asian Children

“…Widespread applicability of hair assays in clinical settings will rely on the availability of low-cost lowertechnology hair assays, currently under development. 26 In conclusion, this is the first study to demonstrate an association between ARV concentrations in hair and virologic outcomes among HIV-infected children. In our cohort, selfreported adherence and pill counts were not associated with virologic outcomes, reflecting well-described limitations of these measures.…”

Antiretroviral Concentrations in Small Hair Samples as a Feasible Marker of Adherence in Rural Kenya

“…Measurement of NVP concentrations via LC-MS/MS is approximately $30/hair sample, although a newly-described method using thin-layer chromatography would cost significantly less (~$5/hair sample). 31 Of note, the cost of an HIV viral load in the region is ~$50.…”

“…Although the costs to analyze the samples using LC/MS-MS were higher, we have developed a low-cost assay for analyzing NVP in hair semiquantitatively that will significantly lower costs 31 and similar low-cost assays for other ARVs (including efavirenz) are in development.…”

“…The wide variability in drug exposure in hair at similar levels of self-reported adherence reinforces the limited utility of self-report. 34 Hair ARV measures, especially those using lower-cost methods, 31 may supplement assessment of ART adherence in remote, resource-limited settings and provide an objective tool to predict impending virologic failure and improve outcomes.…”

Strong Correlation Between Concentrations of Tenofovir (TFV) Emtricitabine (FTC) in Hair and TFV Diphosphate and FTC Triphosphate in Dried Blood Spots in the iPrEx Open Label Extension: Implications for Pre-exposure Prophylaxis Adherence Monitoring

“…However, it should be noted that current assays to quantify drug levels for PrEP, including those in both DBS and hair, use LC/MS-MS, which requires expensive equipment. Widespread applicability of biologic adherence measures in the context of PrEP rollout will rely on the availability of lower-cost methods to analyze drug levels, currently in development for hair assays [13].…”