Current HIV antiretroviral (ART) or pre-exposure prophylaxis (PrEP) therapy adherence monitoring relies on either patient self-reported adherence or monitoring drug dispensing, which are not reliable. We propose an objective 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), a deoxyadenosine triphosphate (dATP) analog and an active NRTI compound 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. We present and validate a model that predicts the amplification probability as function of dATP and TFV-DP concentrations, nucleotide binding sites on the RNA template, and the RNA template concentration. The model helps to rationally design and optimize the assay to operate at clinically relevant TFV-DP concentrations. We provide statistical analysis that demonstrates how the assay can be used as a qualitative or semi-quantitative tool for measuring adherence to NRTI drugs and used to support patient compliance.