HIV pre-exposure prophylaxis adherence test using reverse transcription isothermal amplification inhibition assay

Current HIV antiretroviral therapy (ART) or pre-exposure prophylaxis (PrEP) therapy adherence monitoring relies on either patient self-reported adherence or monitored drug dispensing, which are not reliable. We report a proof-of-concept adherence monitoring assay which directly measures nucleotide r...

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Bibliographic Details
Published in:Analytical methods 2022-03, Vol.14 (13), p.1361-1370
Main Authors: Zhang, Jane Y, Zhang, Yu, Bender, Andrew T, Sullivan, Benjamin P, Olanrewaju, Ayokunle O, Lillis, Lorraine, Boyle, David, Drain, Paul K, Posner, Jonathan D
Format: Article
Language:English
Subjects:
Amplification
Anti-HIV Agents - therapeutic use
Antiretroviral agents
Antiretroviral therapy
Assaying
Deoxyadenosine
Design optimization
Disease prevention
HIV
HIV Infections - diagnosis
HIV Infections - drug therapy
HIV Infections - prevention & control
Human immunodeficiency virus
Humans
Instrumentation
Monitoring
Nucleic acids
Nucleotides
Pre-Exposure Prophylaxis
Prophylaxis
Recombinase
Reverse Transcription
Ribonucleic acid
RNA
RNA-directed DNA polymerase
Statistical analysis
Tenofovir
Tenofovir - therapeutic use
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Summary:Current HIV antiretroviral therapy (ART) or pre-exposure prophylaxis (PrEP) therapy adherence monitoring relies on either patient self-reported adherence or monitored drug dispensing, which are not reliable. We report a proof-of-concept adherence monitoring assay which directly measures nucleotide reverse transcriptase inhibitor (NRTI) concentration using a reverse transcription isothermal amplification inhibition assay. We measure the concentration of Tenofovir diphosphate (TFV-DP) - an NRTI that functions as a deoxyadenosine triphosphate (dATP) analog and long-term adherence marker for PrEP - by measuring the inhibition of the reverse transcription of an RNA template. The completion or inhibition of reverse transcription is evaluated by recombinase polymerase amplification (RPA), an isothermal nucleic acid amplification assay commonly used for point-of-care diagnostics. We present and validate a model that predicts the amplification probability as a function of dATP and TFV-DP concentrations, nucleotide insertion sites on the RNA template, and RNA template concentration. The model can be used to rationally design and optimize the assay to operate at clinically relevant TFV-DP concentrations. We provide statistical analysis that demonstrates how the assay may be used as a qualitative or semi-quantitative tool for measuring adherence to NRTI drugs and used to support patient compliance. Due to its simple instrumentation and short runtime (
ISSN:1759-9660
1759-9679
DOI:10.1039/d2ay00008c