Deep analysis of CD4 T cells in the rhesus CNS during SIV infection.

Virologic suppression with antiretroviral therapy (ART) has significantly improved health outcomes for people living with HIV, yet challenges related to chronic inflammation in the central nervous system (CNS)-known as Neuro-HIV- persist. As primary targets for HIV-1 with the ability to survey and populate the CNS and interact with myeloid cells to co-ordinate neuroinflammation, CD4 T cells are pivotal in Neuro-HIV. Despite their importance, our understanding of CD4 T cell distribution in virus-targeted CNS tissues, their response to infection, and potential recovery following initiation of ART remain limited. To address these gaps, we studied ten SIVmac251-infected rhesus macaques using an ART regimen simulating suboptimal adherence. We evaluated four macaques during the acute phase pre-ART and six during the chronic phase. Our data revealed that HIV target CCR5+ CD4 T cells inhabit both the brain parenchyma and adjacent CNS tissues, encompassing choroid plexus stroma, dura mater, and the skull bone marrow. Aligning with the known susceptibility of CCR5+ CD4 T cells to viral infection and their presence within the CNS, high levels of viral RNA were detected in the brain parenchyma and its border tissues during acute SIV infection. Single-cell RNA sequencing of CD45+ cells from the brain revealed colocalization of viral transcripts within CD4 clusters and significant activation of antiviral molecules and specific effector programs within T cells, indicating CNS CD4 T cell engagement during infection. Acute infection led to marked imbalance in the CNS CD4/CD8 ratio which persisted into the chronic phase. These observations underscore the functional involvement of CD4 T cells within the CNS during SIV infection, enhancing our understanding of their role in establishing CNS viral presence. Our findings offer insights for potential T cell-focused interventions while underscoring the challenges in eradicating HIV from the CNS, particularly in the context of sub-optimal ART.

Application of infrared matrix-assisted laser desorption electrospray ionization mass spectrometry for morphine imaging in brain tissue.

Here, we present a method developed for the analysis of spatial distributions of morphine in mouse brain tissue using infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) coupled to a Q Exactive Plus mass spectrometer. The method is also capable of evaluating spatial distributions of the antiretroviral drug abacavir. To maximize sensitivity to morphine, we analyze various Orbitrap mass spectrometry acquisition modes utilizing signal abundance and frequency of detection as evaluation criteria. We demonstrate detection of morphine in mouse brain and establish that the selected ion monitoring mode provides 2.5 times higher sensitivity than the full-scan mode. We find that distributions of morphine and abacavir are highly correlated with the Pearson correlation coefficient R = 0.87. Calibration showed that instrument response is linear up to 40 pg/mm2 (3.8 µg/g of tissue).

Doravirine Concentrations and Human Immunodeficiency Virus Type 1 RNA in the Genital Fluids of Virologically Suppressed Adults Switching to Doravirine Plus Emtricitabine/Tenofovir Alafenamide.

Doravirine (DOR) concentrations and HIV-1 RNA were evaluated in genital fluids from adults with HIV on stable therapy who switched to DOR + FTC/TAF. High protein-unbound DOR concentrations were observed in both seminal plasma and cervicovaginal fluid. DOR + FTC/TAF maintained viral suppression in genital fluids in all but 1 participant.

Mass Spectroscopy Imaging of Hair Strands Captures Short-Term and Long-Term Changes in Emtricitabine Adherence.

Most measures of adherence to antiretroviral therapy require a blood sample, and none capture longitudinal daily adherence. A new noninvasive method for measuring daily adherence to antiretroviral regimens containing emtricitabine (FTC) was developed for intact hair strands using infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) mass spectrometry imaging (MSI). A directly observed therapy study of daily and intermittent (3, 1, and 0 doses/week) FTC dosing (n = 12) benchmarked adherence in hair, revealing distinct accumulation patterns and median FTC signal abundance (1,702, 495, 352, and 0, respectively) with each dosing frequency. A threshold value of FTCsignal abundance of 500 differentiated daily dosing from 3 or fewer doses/week (specificity, 100%; sensitivity, 100% over 30 days and 80% over 60 days). Using these criteria, daily FTC hair adherence was classified in young men (n = 8) who have sex with men (YMSM) engaged in or initiating preexposure prophylaxis (PrEP). Four types of adherence profiles were observed in sequential 30-day periods: consistently high, occasional missed doses, improvement following study initiation, and intermittent. Discrete days of nonadherence were identified across the 60-day window, with the average number of consecutive days classified as nonadherent increasing across the four profile types (1, 2, 19, and 58 days, respectively). Additionally, cumulative FTC response in hair (60-day average) significantly correlated with dried blood spot tenofovir diphosphate concentrations collected simultaneously (rs = 0.79, P = 0.03). Based on these data, IR-MALDESI FTC adherence classification in hair strands can better delineate short-term changes in adherence behaviors over a long retrospective window, offering great potential for noninvasive adherence monitoring and quick supportive interventions.

Quantitative Imaging Analysis of the Spatial Relationship between Antiretrovirals, Reverse Transcriptase Simian-Human Immunodeficiency Virus RNA, and Fibrosis in the Spleens of Nonhuman Primates.

Although current antiretroviral therapy (ART) has increased life expectancy, a cure for human immunodeficiency virus (HIV) remains elusive due to the persistence of the virus in tissue reservoirs. In the present study, we sought to elucidate the relationship between antiretrovirals (ARVs) and viral expression in the spleen. We performed mass spectrometry imaging (MSI) of 6 different ARVs, RNAscope in situ hybridization of viral RNA, and immunohistochemistry of three different fibrosis markers in the spleens of 8 uninfected and 10 reverse transcriptase simian-human immunodeficiency virus (RT-SHIV)-infected rhesus macaques (infected for 6 weeks) that had been dosed for 10 days with combination ART. Using MATLAB, computational quantitative imaging analysis was performed to evaluate the spatial and pharmacological relationships between the 6 ARVs, viral RNA, and fibrotic deposition. In these spleens, >50% of the spleen tissue area was not covered by any detectable ARV response (any concentration above the limits of detection for individual ARVs). The median spatial ARV coverage across all tissues was driven by maraviroc followed by efavirenz. Yet >50% of RNA-positive cells were not exposed to any detectable ARV. Quantifiable maraviroc and efavirenz colocalization with RNA-positive cells was usually greater than the in vitro concentration inhibiting 50% replication (IC50). Fibrosis markers covered more than 50% of the spleen tissue area and had negative relationships with cumulative ARV coverages. Our findings suggest that a heterogeneous ARV spatial distribution must be considered when evaluating viral persistence in lymphoid tissue reservoirs.

Intracellular islatravir pharmacology differs between species in an in vitro model: implications for preclinical study design.

BACKGROUND: Islatravir (4'-ethynyl-2-fluoro-2'-deoxyadenosine; EFdA) is a first-in-class nucleoside reverse transcriptase translocation inhibitor (NRTTI) being investigated for HIV treatment and prevention. EFdA is intracellularly phosphorylated to EFdA-triphosphate (EFdA-tp), a competitive substrate of deoxyadenosine-triphosphate (dATP). Thus, translating safety and efficacy findings from preclinical studies relies on the assumption that EFdA's intracellular pharmacology can be extrapolated across species. OBJECTIVES: We investigated how EFdA is phosphorylated across animal species commonly used for preclinical models in drug development to identify those that most closely matched humans. METHODS: PBMCs were isolated from whole blood of six species (human, rhesus macaque non-human primate (rmNHP), rat, minipig, dog, and rabbit) using Ficoll separation and counted on a haemocytometer by Trypan blue staining. One million live cells were cultured in media supplemented with 10 U/mL human IL-2, 10% FBS and 1% antibiotics and treated with 0, 17, 170, and 1700 nM EFdA (n = 3 replicates per concentration). After 24 h, representative cell counts were derived from untreated control wells (as above), cells were washed in PBS, and lysed with 70:30 methanol:water. EFdA-tp and dATP concentrations were quantified by HPLC-MS/MS and normalized to the representative live cell counts for each species. RESULTS: When compared to human values, EFdA-tp concentrations for each EFdA treatment concentration were lower in all species (rmNHP 1.5-2.1-fold, rat 4.5-15-fold, minipig 37-71-fold, dog and rabbit >100-fold). Additionally, rmNHP and dog PBMCs exhibited significantly higher (7-10-fold; P < 0.001) dATP when compared with human PBMCs. CONCLUSIONS: Given intracellular pharmacology differences, these preclinical models may be a conservative estimate of EFdA's intracellular pharmacokinetics and efficacy in humans.

Feasibility, Acceptability, and Preliminary Efficacy of a Gamified Mobile Health Contingency Management Intervention for PrEP Adherence Among Black MSM.

Oral HIV pre-exposure prophylaxis (PrEP) is effective at preventing HIV. However, low adherence is common and undermines these protective effects. This is particularly relevant for groups with disproportionately higher rates of HIV, including Black men who have sex with men (MSM). The current study tested the feasibility, acceptability, and preliminary efficacy of a gamified mobile health contingency management intervention for PrEP adherence-called mSMART (Mobile App-Based Personalized Solutions for Medication Adherence of Rx Pill Tool). Fifteen Black MSM already prescribed PrEP in the community completed baseline and follow-up assessments separated by 8 weeks of using mSMART. Regarding feasibility, there was no study attrition, no mSMART functional difficulties that significantly interfered with use, and a mean rate of 82% daily mSMART use. Acceptability ratings were in the moderately to extremely satisfied range for factors such as willingness to recommend mSMART to others and user-friendliness, and in the low range for ratings on difficulty learning how to use mSMART. Scores on a system usability measure were in the acceptable range for 73% of the sample. Qualitative analysis of follow-up interviews identified individual components of mSMART that could be modified in future iterations to make it more engaging. PrEP composite adherence scores from biomarkers indicated an improvement from baseline to follow-up with a medium effect size, as well as a decrease in the number of perceived barriers to medication adherence. Findings indicate a future efficacy trial is needed to examine the effects of this gamified mobile health contingency management intervention on PrEP adherence.

Dynamics of the Decay of Human Immunodeficiency Virus (HIV) RNA and Distribution of Bictegravir in the Genital Tract and Rectum in Antiretroviral-naive Adults Living With HIV-1 Treated With Bictegravir/Emtricitabine/Tenofovir Alafenamide (Spanish HIV/AIDS Research Network, PreEC/RIS 58).

BACKGROUND: The pharmacokinetics of bictegravir (BIC) and its association with the decay of human immunodeficiency virus (HIV)-1 RNA in genital fluids and the rectum have not yet been addressed. METHODS: We conducted a prospective, multicenter study of antiretroviral-naive people living with HIV-1 and initiating BIC/emtricitabine (FTC)/tenofovir alafenamide (TAF). HIV-1 RNA was measured (limit of quantification, 40 copies/mL) in blood plasma (BP), seminal plasma (SP), rectal fluid (RF), and cervicovaginal fluid (CVF) at baseline; Days 3, 7, 14, and 28; and Weeks 12 and 24. Total and protein-unbound BIC concentrations at 24 hours postdose (C24h) were quantified in BP, SP, CVF and rectal tissue (RT) on Day 28 and Week 12 using a validated liquid chromatography-tandem mass spectrometry assay. RESULTS: The study population comprised 15 males and 8 females. In SP, RF, and CVF, the baseline HIV-1 RNA was >40 copies/mL in 12/15, 13/15, and 4/8 individuals, respectively, with medians of 3.54 (2.41-3.79), 4.19 (2.98-4.70), and 2.56 (1.61-3.56) log10 copies/mL, respectively. The initial decay slope was significantly lower in SP than in RF and BP. The time to undetectable HIV-1 RNA was significantly shorter in SP and RF than in BP. All women achieved undetectable HIV-1 RNA in CVF at Day 14. The median total BIC concentrations in SP, RT, and CVF were 65.5 (20.1-923) ng/mL, 74.1 (6.0-478.5) ng/g, and 61.6 (14.4-1760.2) ng/mL, respectively, representing 2.7%, 2.6%, and 2.8% of the BP concentration, respectively, while the protein-unbound fractions were 51.1%, 44.6%, and 42.6%, respectively. CONCLUSIONS: BIC/FTC/TAF led to rapid decay of HIV-1 RNA in genital and rectal fluids. Protein-unbound BIC concentrations in SP, RT, and CVF highly exceeded the half-maximal effective concentration (EC50) value (1.1 ng/mL). CLINICAL TRIALS REGISTRATION: EudraCT 2018-002310-12.

Quantitative Imaging Analysis of the Spatial Relationship between Antiretrovirals, Reverse Transcriptase Simian-Human Immunodeficiency Virus RNA, and Collagen in the Mesenteric Lymph Nodes of Nonhuman Primates.

Human immunodeficiency virus (HIV) persistence in tissue reservoirs is a major barrier to HIV cure. While antiretrovirals (ARVs) suppress viral replication, antiretroviral therapy (ART) interruption results in rapid rebound viremia that may originate from lymphoid tissues. To understand the relationship between anatomic distribution of ARV exposure and viral expression in lymph nodes, we performed mass spectrometry imaging (MSI) of 6 ARVs, RNAscope in situ hybridization for viral RNA (vRNA), and immunohistochemistry of collagen in mesenteric lymph nodes from 8 uninfected and 10 reverse transcriptase simian/human immunodeficiency virus (RT-SHIV)-infected rhesus macaques dosed to steady state with combination ART. MATLAB-based quantitative imaging analysis was used to evaluate spatial and pharmacological relationships between these ARVs, viral RNA (both vRNA+ cells and follicular dendritic cell [FDC]-bound virions), and collagen deposition. Using MSI, 31% of mesenteric lymph node tissue area was found to be not covered by any ARV. Additionally, 28% of FDC-trapped virions and 21% of infected cells were not exposed to any detected ARV. Of the 69% of tissue area that was covered by cumulative ART exposure, nearly 100% of concentrations were greater than in vitro 50% inhibitory concentration (IC50) values; however, 52% of total tissue coverage was from only one ARV, primarily maraviroc. Collagen covered ∼35% of tissue area but did not influence ARV distribution heterogeneity. Our findings are consistent with our hypothesis that ARV distribution, in addition to total-tissue drug concentration, must be considered when evaluating viral persistence in lymph nodes and other reservoir tissues.

A cross-species comparison of antiretroviral penetration into lymph nodes using novel physiologically based pharmacokinetic models.

BACKGROUND: Investigating antiretroviral (ARV) penetration and pharmacology in lymph nodes is crucial to understanding mechanisms of HIV persistence in tissue, but sampling these tissues in humans is invasive and costly. Physiologically based pharmacokinetic (PBPK) modelling is a non-invasive solution for understanding lymph node penetration of ARVs across multiple species. OBJECTIVES: To develop customized PBPK models with a novel lymph node compartment, and use these models to describe the distribution of three ARVs-tenofovir, emtricitabine and efavirenz-into the plasma and lymph nodes of non-human primates (NHPs) and humans. MATERIALS AND METHODS: In this analysis, we utilized standard monkey and human PBPK models in PK-Sim, and added a novel lymph node compartment using MoBi. We used these models to describe the distribution of tenofovir, emtricitabine and efavirenz into NHP and human plasma and lymph nodes, and compared model-predicted versus observed AUC and Cmax. RESULTS: For all three ARVs, population simulations using the base and final models reasonably characterized observed plasma and tissue data in NHPs and humans, with predicted/observed AUC and Cmax ratios within 0.7-2.0. CONCLUSIONS: Overall, our novel PBPK model provides a framework for understanding lymph node penetration of ARVs or future HIV cure therapies.

Long-Acting Rilpivirine (RPV) Preexposure Prophylaxis Does Not Inhibit Vaginal Transmission of RPV-Resistant HIV-1 or Select for High-Frequency Drug Resistance in Humanized Mice.

As a long-acting formulation of the nonnucleoside reverse transcriptase inhibitor rilpivirine (RPV LA) has been proposed for use as preexposure prophylaxis (PrEP) and the prevalence of transmitted RPV-resistant viruses can be relatively high, we evaluated the efficacy of RPV LA to inhibit vaginal transmission of RPV-resistant HIV-1 in humanized mice. Vaginal challenges of wild-type (WT), Y181C, and Y181V HIV-1 were performed in mice left untreated or after RPV PrEP. Plasma viremia was measured for 7 to 10 weeks, and single-genome sequencing was performed on plasma HIV-1 RNA in mice infected during PrEP. RPV LA significantly prevented vaginal transmission of WT HIV-1 and Y181C HIV-1, which is 3-fold resistant to RPV. However, it did not prevent transmission of Y181V HIV-1, which has 30-fold RPV resistance in the viruses used for this study. RPV LA did delay WT HIV-1 dissemination in infected animals until genital and plasma RPV concentrations waned. Animals that became infected despite RPV LA PrEP did not acquire new RPV-resistant mutations above frequencies in untreated mice or untreated people living with HIV-1, and the mutations detected conferred low-level resistance. These data suggest that high, sustained concentrations of RPV were required to inhibit vaginal transmission of HIV-1 with little or no resistance to RPV but could not inhibit virus with high resistance. HIV-1 did not develop high-level or high-frequency RPV resistance in the majority of mice infected after RPV LA treatment. However, the impact of low-frequency RPV resistance on virologic outcome during subsequent antiretroviral therapy still is unclear.IMPORTANCE The antiretroviral drug rilpivirine was developed into a long-acting formulation (RPV LA) to improve adherence for preexposure prophylaxis (PrEP) to prevent HIV-1 transmission. A concern is that RPV LA will not inhibit transmission of drug-resistant HIV-1 and may select for drug-resistant virus. In female humanized mice, we found that RPV LA inhibited vaginal transmission of WT or 3-fold RPV-resistant HIV-1 but not virus with 30-fold RPV resistance. In animals that became infected despite RPV LA PrEP, WT HIV-1 dissemination was delayed until genital and plasma RPV concentrations waned. RPV resistance was detected at similar low frequencies in untreated and PrEP-treated mice that became infected. These results indicate the importance of maintaining RPV at a sustained threshold after virus exposure to prevent dissemination of HIV-1 after vaginal infection and low-frequency resistance mutations conferred low-level resistance, suggesting that RPV resistance is difficult to develop after HIV-1 infection during RPV LA PrEP.

S-warfarin limited sampling strategy with a population pharmacokinetic approach to estimate exposure and cytochrome P450 (CYP) 2C9 activity in healthy adults.

PURPOSE: S-warfarin is used to phenotype cytochrome P450 (CYP) 2C9 activity. This study evaluated S-warfarin limited sampling strategy with a population pharmacokinetic (PK) approach to estimate CYP2C9 activity in healthy adults. METHODS: In 6 previously published studies, a single oral dose of warfarin 10 mg was administered alone or with a CYP2C9 inducer to 100 healthy adults. S-warfarin concentrations were obtained from adults during conditions when subjects were not on any prescribed medications. A population PK model was developed using non-linear mixed effects modeling. Limited sampling models (LSMs) using single- or 2-timepoint concentrations were compared with full PK profiles from intense sampling using empiric Bayesian post hoc estimations of S-warfarin AUC derived from the population PK model. Preset criterion for LSM selection and validation were a correlation coefficient (R2) >0.9, relative percent mean prediction error (%MPE) >-5 to <5%, relative percent mean absolute error (%MAE) ≤ 10%, and relative percent root mean squared error (%RMSE) ≤ 15%. RESULTS: S-warfarin concentrations (n=2540) were well described with a two-compartment model. Mean apparent oral clearance was 0.56 L/hr and volume of distribution was 35.5 L. Clearance decreased 33% with the CYP2C9 *3 allele and increased 42% with lopinavir/ritonavir co-administration. During CYP2C9 constitutive conditions, LSMs at 48 hr and at 72 hr as well as 2-timepoint LSMs were within acceptable limits for R2, %MPE, %MAE, and %RMSE. During CYP2C9 induction, S-warfarin LSMs had unacceptable %MPE, %MAE, and %RMSE. CONCLUSIONS: Phenotyping studies with S-warfarin in healthy subjects can utilize a single- and/or a 2-timepoint LSM with a population PK approach to estimate constitutive CYP2C9 activity.

Food Insecurity Is Associated With Lower Levels of Antiretroviral Drug Concentrations in Hair Among a Cohort of Women Living With Human Immunodeficiency Virus in the United States.

BACKGROUND: Food insecurity is a well-established determinant of suboptimal, self-reported antiretroviral therapy (ART) adherence, but few studies have investigated this association using objective adherence measures. We examined the association of food insecurity with levels of ART concentrations in hair among women living with human immunodeficiency virus (WLHIV) in the United States. METHODS: We analyzed longitudinal data collected semiannually from 2013 through 2015 from the Women's Interagency HIV Study, a multisite, prospective, cohort study of WLHIV and controls not living with HIV. Our sample comprised 1944 person-visits from 677 WLHIV. Food insecurity was measured using the US Household Food Security Survey Module. ART concentrations in hair, an objective and validated measure of drug adherence and exposure, were measured using high-performance liquid chromatography with mass spectrometry detection for regimens that included darunavir, atazanavir, raltegravir, or dolutegravir. We conducted multiple 3-level linear regressions that accounted for repeated measures and the ART medication(s) taken at each visit, adjusting for sociodemographic and clinical characteristics. RESULTS: At baseline, 67% of participants were virally suppressed and 35% reported food insecurity. In the base multivariable model, each 3-point increase in food insecurity was associated with 0.94-fold lower ART concentration in hair (95% confidence interval, 0.89 to 0.99). This effect remained unchanged after adjusting for self-reported adherence. CONCLUSIONS: Food insecurity was associated with lower ART concentrations in hair, suggesting that food insecurity may be associated with suboptimal ART adherence and/or drug absorption. Interventions seeking to improve ART adherence among WLHIV should consider and address the role of food insecurity.

Translational Approach to Predicting the Efficacy of Maraviroc-Based Regimens as HIV Preexposure Prophylaxis.

Maraviroc-based regimens have been explored as preexposure prophylaxis (PrEP) against human immunodeficiency virus (HIV). In this study, we utilized mucosal tissue drug exposure data, combined with target concentrations generated in vitro, in a pharmacokinetic-pharmacodynamic analysis to predict the effects of drug combinations and adherence on PrEP efficacy. Mucosal tissue concentrations of maraviroc were measured in 24 healthy women. The 90% effective concentrations (EC90) of maraviroc (alone and combined with tenofovir and emtricitabine) for protection against HIV were identified in CD4+ T cells. Monte Carlo simulations were performed to identify dosing strategies to protect colorectal and female genital tract (FGT) tissues from HIV infection. Colorectal maraviroc concentrations were 350-fold higher than in the FGT. Under steady-state conditions, our model predicted that one 300-mg dose/week was sufficient to protect colorectal tissue from HIV in 99% of the population, while 300 mg daily would protect the FGT in only 63% of the population. FGT protection increased to >90% when maraviroc was used in combination with tenofovir (5 doses/week) or emtricitabine (3 doses/week). Poor adherence resulted in a drastic decrease in efficacy in the FGT but not colorectal tissue. However, greater forgiveness was seen when maraviroc was combined with tenofovir or emtricitabine, suggesting that maraviroc should not be used alone as PrEP.

Antiretroviral Penetration and Drug Transporter Concentrations in the Spleens of Three Preclinical Animal Models and Humans.

Adequate antiretroviral (ARV) concentrations in lymphoid tissues are critical for optimal antiretroviral therapy (ART). While the spleen contains 25% of the body's lymphocytes, there are minimal data on ARV penetration in this organ. This study quantified total and protein-unbound splenic ARV concentrations and determined whether drug transporters, sex, or infection status were modifiers of these concentrations in animal models and humans. Two humanized mice models (hu-HSC-Rag [n = 36; 18 HIV-positive (HIV+) and 18 HIV-negative (HIV-)] and bone marrow-liver-thymus [n = 13; 7 HIV+ and 6 HIV-]) and one nonhuman primate (NHP) model (rhesus macaque [n = 18; 10 SHIV+ and 8 SHIV-]) were dosed to steady state with ARV combinations. HIV+ human spleens (n = 14) from the National NeuroAIDS Tissue Consortium were analyzed postmortem (up to 24 h postdose). ARV concentrations were measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS), drug transporter concentrations were measured with LC-MS proteomics, and protein binding in NHP spleens was determined by rapid equilibrium dialysis. Mice generally had the lowest splenic concentrations of the three species. Protein binding in splenic tissue was 6 to 96%, compared to 76 to 99% in blood plasma. NHPs had quantifiable Mrp4, Bcrp, and Ent1 concentrations, and humans had quantifiable ENT1 concentrations. None significantly correlated with tissue ARV concentrations. There was also no observable influence of infection status or sex. With these dosing strategies, NHP splenic penetration most closely resembled that of humans. These data can inform tissue pharmacokinetic scaling to humans to target HIV reservoirs by identifying important species-related differences.

Application of a Scavenger Receptor A1-Targeted Polymeric Prodrug Platform for Lymphatic Drug Delivery in HIV.

We have developed a macromolecular prodrug platform based on poly(l-lysine succinylated) (PLS) that targets scavenger receptor A1 (SR-A1), a receptor expressed by myeloid and endothelial cells. We demonstrate the selective uptake of PLS by murine macrophage, RAW 264.7 cells, which was eliminated upon cotreatment with the SR-A inhibitor polyinosinic acid (poly I). Further, we observed no uptake of PLS in an SR-A1-deficient RAW 264.7 cell line, even after 24 h incubation. In mice, PLS distributed to lymphatic organs following i.v. injection, as observed by ex vivo fluorescent imaging, and accumulated in lymph nodes following both i.v. and i.d. administrations, based on immunohistochemical analysis with high-resolution microscopy. As a proof-of-concept, the HIV antiviral emtricitabine (FTC) was conjugated to the polymer's succinyl groups via ester bonds, with a drug loading of 14.2% (wt/wt). The prodrug (PLS-FTC) demonstrated controlled release properties in vitro with a release half-life of 15 h in human plasma and 29 h in esterase-inhibited plasma, indicating that drug release occurs through both enzymatic and nonenzymatic mechanisms. Upon incubation of PLS-FTC with human peripheral blood mononuclear cells (PBMCs), the released drug was converted to the active metabolite FTC triphosphate. In a pharmacokinetic study in rats, the prodrug achieved ∼7-19-fold higher concentrations in lymphatic tissues compared to those in FTC control, supporting lymphatic-targeted drug delivery. We believe that the SR-A1-targeted macromolecular PLS prodrug platform has extraordinary potential for the treatment of infectious diseases.

Influence of hair treatments on detection of antiretrovirals by mass spectrometry imaging.

Analysis of drugs in hair by mass spectrometry imaging (MSI) has great potential as an objective, long-term measure of medication adherence. However, the fidelity of the chemical record in hair may be compromised by any cosmetic hair treatments. Here, we investigate infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) MSI response to multiple antiretrovirals (ARVs) in cosmetically treated hair. Hair strands from patients on different ARV regimens were mechanically treated with dye, bleach, and relaxer. The treatments had little or no effect relative to untreated controls for cobicistat, abacavir, dolutegravir, maraviroc, efavirenz, and darunavir, but all three treatments removed emtricitabine (FTC) to undetectable levels from patient hair strands. We also evaluated hair strands by IR-MALDESI MSI from 8 patients on FTC-based regimens who reported a range of hair treatments at varying recency prior to hair collection. While FTC was undetectable in the treated portion of these hair strands, ARVs coadministered with FTC remained detectable in hair strands after treatment. We conclude that IR-MALDESI MSI can be used when measuring adherence to ARV therapy, provided that ARVs other than FTC are targeted in people using hair treatments.

Seminal Tenofovir Concentrations, Viral Suppression, and Semen Quality With Tenofovir Alafenamide, Compared With Tenofovir Disoproxil Fumarate (Spanish HIV/AIDS Research Network, PreEC/RIS 40).

BACKGROUND: This study assessed the penetration and efficacy of tenofovir alafenamide (TAF) in the male genital tract (MGT) and the semen quality of individuals infected with human immunodeficiency virus (HIV)-1 who were treated with a TAF-containing regimen. METHODS: This was a prospective, open-label, single-arm study of 14 virologically-suppressed, HIV-1-infected men on stable antiretroviral therapy with elvitegravir, cobicistat, emtricitabine (E/C/F) and tenofovir disoproxil fumarate (TDF) who switched to E/C/F and TAF. At baseline (pre-switch) and at 12 weeks post-switch, we measured HIV-1 RNA in seminal plasma (SP) and blood plasma (BP), tenofovir (TFV) in SP and BP, and TFV-diphosphate (dp) in peripheral blood mononuclear cells (PBMCs) and seminal mononuclear cells (SMCs) at the end of the dosing interval (C24h). Semen quality was assessed before switching and after 12 weeks on TAF. RESULTS: With TAF, TFV C24 was 11.9-fold higher in SP than in BP. This concentration was significantly lower than TFV C24 in SP with TDF, but 9.6-fold higher than the 50% inhibitory concentration (IC50) (11.5 ng/mL). By contrast, the median TFV-dp concentration achieved with TAF in SMCs was 6% that of TFV-dp in PBMCs. The TFV-dp SMC:PBMC ratio was also significantly lower with TAF. Nonetheless, TFV-dp C24 in SMC was comparable with TAF and TDF. All the patients had HIV-1 RNA <40 copies/mL in BP and SP at baseline and at 12 weeks post-switch. No significant differences were observed in semen quality between TAF and TDF. CONCLUSIONS: Extracellular and intracellular seminal TFV distribution differs between TAF and TDF. Nevertheless, both formulations, combined with elvitegravir/cobicistat/emtricitabine, maintained HIV-1 RNA suppression in semen. Differences in MGT distribution were not associated with differences in semen quality. CLINICAL TRIALS REGISTRATION: EudraCT: 2016-001371-69.

Decreased Tenofovir Diphosphate Concentrations in a Transgender Female Cohort: Implications for Human Immunodeficiency Virus Preexposure Prophylaxis.

Feminizing hormone therapy (FHT) may interact with human immunodeficiency virus preexposure prophylaxis (PrEP). We found that transgender women who took FHT exhibited a 7-fold lower rectal tissue ratio of PrEP's active metabolites vs competing deoxynucleotides compared to cisgender women and men (P = .03) that inversely correlated with estradiol (ρ = -0.79; P < .05). Thus, FHT may negatively impact PrEP efficacy. Clinical Trials Registration . NCT02983110.

Antiretroviral Penetration across Three Preclinical Animal Models and Humans in Eight Putative HIV Viral Reservoirs.

For HIV cure strategies like "kick and kill" to succeed, antiretroviral (ARV) drugs must reach effective concentrations in putative viral reservoirs. We characterize penetration of six ARVs in three preclinical animal models and humans. We found that standard dosing strategies in preclinical species closely mimicked tissue concentrations in humans for some, but not all, ARVs. These results have implications for interpreting HIV treatment, prevention, or cure interventions between preclinical and clinical models.