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  • protoporphyrin ix Management of HIV treatment with HIV PIs

    2021-10-12

    Management of HIV-1 treatment with HIV PIs can be complicated due to wide inter-patient variability in protoporphyrin ix levels. In certain scenarios, therapeutic drug monitoring (TDM) has been recommended as a useful approach to individualize therapy for HIV-infected patients [[9], [10], [11], [12]]. Several analytical methods have been reported for measuring HIV PIs in human plasma, and these primarily involve liquid chromatography coupled with tandem mass spectrometry (LC–MS/MS) [[13], [14], [15]]. Although those state of the art methods has numerous advantages, specifically excellent selectivity and sensitivity, major limitations include long turn-around times, expensive equipment, and technically trained lab staff, thus this approach is often too complex for routine clinical laboratories. Colloidal gold nanoparticles have been utilized for centuries due to the vibrant colors produced when the particles interact with the visible light. These unique optoelectronic properties have been applied in high technology applications, for example, sensory probes, therapeutic agents, drug delivery in biological and medical applications, electronic conductors and catalysis. Gold nanoparticles can be widely used to adsorb biomolecules such as proteins, antibodies, and aptamers onto their surfaces [[16], [17], [18]]. The remarkable physicochemical properties of gold nanoparticles have prompted developments in the exploration of biomolecular interactions with gold particle-containing systems, in particular for biomedical applications in diagnostics. As gold particles can help visualize biomolecule complexes by the naked they has been widely applied to simplify diagnostic tests [19]. Previously we reported the development of a novel immunochromatographic (IC) strip test that utilized colloidal gold tracers allowing interpretation of the results by the naked eye. This high-throughput assay is for detecting the enzymatic activity of HIV-PR and the level of HIV PI spiked in human plasma. This novel immunochromatographic (IC) strip test, named “HIV protease inhibitors-based immunochromatographic strip” or “PIs-IC strip” was established by combining proteolysis activity of HIV-PR and an immunochromatographic reaction [20]. In this present study, we report the assessment of this IC strip for the semi-quantification of HIV PIs in plasma samples from HIV-infected patients on ART. The method is sufficiently sensitive to measure therapeutic concentrations of PIs and could be adaptable to laboratories for drug monitoring in resource limited-countries. Moreover, this novel strip could also be useful to detect and discover new drugs for combating with HIV-1 PR drug resistance strains.
    Methods
    Results
    Discussion Simplified assays for detecting antiretroviral drugs concentrations in people receiving antiretroviral therapy can help with the treatment management. The use of IC strip test to detect the levels of the antiretroviral drugs in patient specimens have been developed to overcome the limitation of technical-demanding, time-consuming and complex-expensive of the quantitative analytical methods such as HPLC, LC-MS or GC-MS [25,31,32]. Recently, we developed a high throughput assay to determine the enzymatic activity of HIV-PRH6, which was able to apply to detect HIV PIs in spiked-human plasma samples. Our previous data show that the PIs-IC strip could detect LPV concentration as low as 100 ng mL-1, while LPV concentration over 1,000 ng mL-1 fully inhibited HIV-PRH6 activity. Thus it was thought that the PIs-IC strip is a semi-quantitative method because it could give information that the LPV concentration was less than or greater than 1,000 ng mL-1. In addition, this concentration point is matched with the proposed efficacy “cut off” concentration for LPV, HIV/AIDS Treatment Guideline/AIDSinfo 2014. The production of the H6MA-CA substrate and recombinant HIV-1 PR were verified by specific monoclonal antibody testing (Figs. 1 and 2). In order to detect HIV-PR activity, we used an ELISA-based assay and western blotting analysis using specific binding of both anti-MA G53 and anti-mA HB-8975 mAb were able to detect HIV-PR activity protoporphyrin ix by both methods (Figs. S1 and S2).