Non-nucleoside reverse transcriptase inhibitors (NNRTIs) induce pyroptosis of HIV-1-infected CD4
T cells through induction of intracellular HIV-1 protease activity, which activates the CARD8 ...inflammasome. Because high concentrations of NNRTIs are required for efficient elimination of HIV-1-infected cells, it is important to elucidate ways to sensitize the CARD8 inflammasome to NNRTI-induced activation. We show that this sensitization can be achieved through chemical inhibition of the CARD8 negative regulator DPP9. The DPP9 inhibitor Val-boroPro (VbP) can kill HIV-1-infected cells without the presence of NNRTIs and act synergistically with NNRTIs to promote clearance of HIV-1-infected cells in vitro and in humanized mice. More importantly, VbP is able to enhance clearance of residual HIV-1 in CD4
T cells isolated from people living with HIV (PLWH). We also show that VbP can partially overcome NNRTI resistance. This offers a promising strategy for enhancing NNRTI efficacy in the elimination of HIV-1 reservoirs in PLWH.
Since January 2017, the recommended first-line antiretroviral regimen in Brazil is the fixed-dose combination of tenofovir plus lamivudine with dolutegravir (TL + D). According to the literature, ...integrase resistance-associated mutations (INRAMs) are rarely found upon virologic failure to first-line dolutegravir plus two nucleoside reverse transcriptase inhibitors. We evaluated the HIV antiretroviral genotypic resistance profile of patients referred for genotyping in the public health system who failed first-line TL + D after at least six months of therapy on or before December 31, 2018.
HIV Sanger sequences of the pol gene were generated from plasma of patients with confirmed virologic failure to first-line TL + D in the Brazilian public health system before December 31, 2018.
One hundred thirteen individuals were included in the analysis. Major INRAMs were detected in seven patients (6.19%), four with R263K, one with G118R, one with E138A, and one with G140R. Four patients with major INRAMs also had the K70E and M184V mutations in the RT gene. Sixteen (14.2%) additional individuals presented minor INRAMs, and five (4,42%) patients had both major and minor INRAMS. Thirteen (11.5%) patients also presented mutations in the RT gene selected by tenofovir and lamivudine, including four with both the K70E and M184V mutations and four with only M184V. The integrase mutations L101I and T124A, which are in the in vitro pathway for integrase inhibitor resistance, were found in 48 and 19 patients, respectively. Mutations not related to TL + D, thus probable transmitted resistance mutations (TDR), were present in 28 patients (24.8%): 25 (22.1%) to nucleoside reverse transcriptase inhibitors, 19 (16.8%) to non-nucleoside reverse transcriptase inhibitors, and 6 (5.31%) to protease inhibitors.
In marked contrast to previous reports, we report a relatively high frequency of INRAMs among selected patients failing first-line TL + D in the public health system in Brazil. Possible reasons for this discrepancy include delays in detecting virologic failure, patients inadvertently on dolutegravir monotherapy, TDR, and/or infecting subtype.
Abstract
Background
Increased access to antiretroviral therapy (ART) has resulted in rising levels of pretreatment human immunodeficiency virus drug resistance (PDR). This is the first systematic ...review and meta-analysis to assess the impact of PDR on treatment outcomes among people initiating nonnucleoside reverse transcriptase inhibitor (NNRTI)–based ART, including the combination of efavirenz (EFV), tenofovir (TDF), and lamivudine or emtricitabine (XTC).
Methods
We systematically reviewed studies and conference proceedings comparing treatment outcomes in populations initiating NNRTI-based ART with and without PDR. We conducted subgroup analyses by regimen: (1) NNRTIs + 2 nucleoside reverse transcriptase inhibitors (NRTIs), (2) EFV + 2 NRTIs, or (3) EFV/TDF/XTC; by population (children vs adults); and by definition of resistance (PDR vs NNRTI PDR).
Results
Among 6197 studies screened, 32 were analyzed (31 441 patients). We found that individuals with PDR initiating NNRTIs across all the subgroups had increased risk of virological failure compared to those without PDR. Risk of acquisition of new resistance mutations and ART switch was also higher in people with PDR.
Conclusions
This review shows poorer treatment outcomes in the presence of PDR, supporting the World Health Organization’s recommendation to avoid using NNRTIs in countries where levels of PDR are high.
Higher risks of virological failure, acquisition of new resistance mutations, and treatment switch were observed among adults and children with pretreatment HIV drug resistance initiating nonnucleoside reverse transcriptase inhibitor–containing regimens, including efavirenz/tenofovir/lamivudine or emtricitabine.
Like normal cellular nucleosides, the nucleoside reverse transcriptase (RT) inhibitor (NRTI) 4'-ethynyl-2-fluoro-2'-deoxyadenosine (EFdA) has a 3'-hydroxyl moiety, and yet EFdA is a highly potent ...inhibitor of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) replication with activity against a broad range of clinically important drug-resistant HIV isolates. We evaluated the anti-HIV activity of EFdA in primary human cells and in HIV-infected humanized mice. EFdA exhibited excellent potency against HIVJR-CSF in phytohemagglutinin-stimulated peripheral blood mononuclear cells (PBMCs), with a 50% inhibitory concentration of 0.25 nM and a selectivity index of 184,000; similar antiviral potency was found against 12 different HIV clinical isolates from multiple clades (A, B, C, D, and CRF01_AE). EFdA was readily absorbed after oral dosing (5 mg/kg of body weight) in both mice and the rhesus macaque, with micromolar levels of the maximum concentration of drug in serum (Cmax) attained at 30 min and 90 min, respectively. Trough levels were at or above 90% inhibitory concentration (IC90) levels in the macaque at 24 h, suggesting once-daily dosing. EFdA showed reasonable penetration of the blood-brain barrier in the rhesus macaque, with cerebrospinal fluid levels at approximately 25% of plasma levels 8 h after single oral dosing. Rhesus PBMCs isolated 24 h following a single oral dose of 5 mg/kg EFdA were refractory to SIV infection due to sufficiently high intracellular EFdA-triphosphate levels. The intracellular half-life of EFdA-triphosphate in PBMCs was determined to be >72 h following a single exposure to EFdA. Daily oral administration of EFdA at low dosage levels (1 to 10 mg/kg/day) was highly effective in protecting humanized mice from HIV infection, and 10 mg/kg/day oral EFdA completely suppressed HIV RNA to undetectable levels within 2 weeks of treatment.
The NNRTI efavirenz has long been one of the most frequently employed antiretroviral drugs in the multidrug regimens used to treat HIV infection, in accordance with its well-demonstrated ...antiretroviral efficacy and favourable pharmacokinetics. However, growing concern about its adverse effects has sometimes led to efavirenz being replaced by other drugs in the initial treatment selection or to switching of therapy to efavirenz-free regimens in experienced patients. Neurological and neuropsychiatric reactions are the manifestations most frequently experienced by efavirenz-treated patients and range from transitory effects, such as nightmares, dizziness, insomnia, nervousness and lack of concentration, to more severe symptoms including depression, suicidal ideation or even psychosis. In addition, efavirenz has recently been associated with mild/moderate neurocognitive impairment, which is of specific relevance given that half of the patients receiving ART eventually suffer some form of HIV-associated neurocognitive disorder. The mechanisms responsible for efavirenz-induced neurotoxicity are unclear, although growing evidence points to disturbances in brain mitochondrial function and bioenergetics. This review offers a comprehensive overview of the current evidence on the interaction that efavirenz displays with the CNS, including the penetration and concentration of the drug in the brain. We discuss the prevalence, types and specificities of its side effects and recently uncovered cellular mechanisms that may be involved in their development.
Antiretroviral medications that are used as prophylaxis can prevent acquisition of human immunodeficiency virus type 1 (HIV-1) infection. However, in clinical trials among African women, the ...incidence of HIV-1 infection was not reduced, probably because of low adherence. Longer-acting methods of drug delivery, such as vaginal rings, may simplify use of antiretroviral medications and provide HIV-1 protection.
We conducted a phase 3, randomized, double-blind, placebo-controlled trial of a monthly vaginal ring containing dapivirine, a non-nucleoside HIV-1 reverse-transcriptase inhibitor, involving women between the ages of 18 and 45 years in Malawi, South Africa, Uganda, and Zimbabwe.
Among the 2629 women who were enrolled, 168 HIV-1 infections occurred: 71 in the dapivirine group and 97 in the placebo group (incidence, 3.3 and 4.5 per 100 person-years, respectively). The incidence of HIV-1 infection in the dapivirine group was lower by 27% (95% confidence interval CI, 1 to 46; P=0.046) than that in the placebo group. In an analysis that excluded data from two sites that had reduced rates of retention and adherence, the incidence of HIV-1 infection in the dapivirine group was lower by 37% (95% CI, 12 to 56; P=0.007) than that in the placebo group. In a post hoc analysis, higher rates of HIV-1 protection were observed among women over the age of 21 years (56%; 95% CI, 31 to 71; P<0.001) but not among those 21 years of age or younger (-27%; 95% CI, -133 to 31; P=0.45), a difference that was correlated with reduced adherence. The rates of adverse medical events and antiretroviral resistance among women who acquired HIV-1 infection were similar in the two groups.
A monthly vaginal ring containing dapivirine reduced the risk of HIV-1 infection among African women, with increased efficacy in subgroups with evidence of increased adherence. (Funded by the National Institutes of Health; ClinicalTrials.gov number, NCT01617096 .).
Islatravir (MK-8591) is a highly potent type 1 human immunodeficiency virus (HIV-1) nucleoside reverse transcriptase translocation inhibitor with a long intracellular half-life that is in development ...for the prevention and treatment of HIV-1. We conducted a randomized, double-blind, placebo-controlled, phase 1 trial in adults without HIV-1 infection. Participants received islatravir or placebo subdermal implants for 12 weeks and were monitored throughout this period and after implant removal. The co-primary end points were safety and tolerability of the islatravir implant and pharmacokinetics, including concentration at day 85, of islatravir triphosphate in peripheral blood mononuclear cells (PBMCs). Secondary end points included additional pharmacokinetic parameters for islatravir triphosphate in PBMCs and the plasma pharmacokinetic profile of islatravir. Based on preclinical data, two doses were assessed: 54 mg (n = 8, two placebo) and 62 mg (n = 8, two placebo). The most frequently reported adverse events were mild-to-moderate implant-site reactions (induration, hematoma, pain). Throughout the 12-week trial, geometric mean islatravir triphosphate concentrations were above a pharmacokinetic threshold of 0.05 pmol per 10
PBMCs, which was estimated to provide therapeutic reverse transcriptase inhibition (concentration at day 85 (percentage of geometric coefficient of variation): 54 mg, 0.135 pmol per 10
cells (27.3); 62 mg, 0.272 pmol per 10
cells (45.2)). Islatravir implants at both doses were safe and resulted in mean concentrations above the pharmacokinetic threshold through 12 weeks, warranting further investigation of islatravir implants as a potential HIV prevention strategy.
The human DNA polymerase gamma (Pol γ) is responsible for DNA replication in mitochondria. Pol γ is particularly susceptible to inhibition by dideoxynucleoside‐based inhibitors designed to fight ...viral infection. Here, we report crystal structures of the replicating Pol γ–DNA complex bound to either substrate or zalcitabine, an inhibitor used for HIV reverse transcriptase. The structures reveal that zalcitabine binds to the Pol γ active site almost identically to the substrate dCTP, providing a structural basis for Pol γ‐mediated drug toxicity. When compared to the apo form, Pol γ undergoes intra‐ and inter‐subunit conformational changes upon formation of the ternary complex with primer/template DNA and substrate. We also find that the accessory subunit Pol γB, which lacks intrinsic enzymatic activity and does not contact the primer/template DNA directly, serves as an allosteric regulator of holoenzyme activities. The structures presented here suggest a mechanism for processivity of the holoenzyme and provide a model for understanding the deleterious effects of Pol γ mutations in human disease. Crystal structures of the mitochondrial DNA polymerase, Pol γ, in complex with substrate or antiviral inhibitor zalcitabine provide a basis for understanding Pol γ‐mediated drug toxicity.
Synopsis
Crystal structures of the mitochondrial DNA polymerase, Pol γ, in complex with substrate or antiviral inhibitor zalcitabine provide a basis for understanding Pol γ‐mediated drug toxicity.
Zalcitabine binds to the Pol γ active site in a similar manner as the substrate dCTP.
The human mitochondrial DNA polymerase ternary complex halted by zalcitabine provides a structural mechanism for antiviral drug toxicity.
The crystal structure suggests the accessory subunit Pol γB to allosterically regulate processivity and proofreading of Pol γ.
This study sets the stage for understanding human diseases associated with the mitochondrial DNA polymerase.
Crystal structures of the mitochondrial DNA polymerase, Pol γ, in complex with substrate or antiviral inhibitor zalcitabine provide a basis for understanding Pol γ‐mediated drug toxicity.
Efavirenz (EFV) is principally metabolized by CYP2B6 to 8-hydroxy-efavirenz (8OH-EFV) and to a lesser extent by CYP2A6 to 7-hydroxy-efavirenz (7OH-EFV). So far, most metabolite profile analyses have ...been restricted to 8OH-EFV, 7OH-EFV, and EFV-N-glucuronide, even though these metabolites represent a minor percentage of EFV metabolites present in vivo. We have performed a quantitative phase I and II metabolite profile analysis by tandem mass spectrometry of plasma, cerebrospinal fluid (CSF), and urine samples in 71 human immunodeficiency virus patients taking efavirenz, prior to and after enzymatic (glucuronidase and sulfatase) hydrolysis. We have shown that phase II metabolites constitute the major part of the known circulating efavirenz species in humans. The 8OH-EFV-glucuronide (gln) and 8OH-EFV-sulfate (identified for the first time) in humans were found to be 64- and 7-fold higher than the parent 8OH-EFV, respectively. In individuals (n = 67) genotyped for CYP2B6, 2A6, and CYP3A metabolic pathways, 8OH-EFV/EFV ratios in plasma were an index of CYP2B6 phenotypic activity (P < 0.0001), which was also reflected by phase II metabolites 8OH-EFV-glucuronide/EFV and 8OH-EFV-sulfate/EFV ratios. Neither EFV nor 8OH-EFV, nor any other considered metabolites in plasma were associated with an increased risk of central nervous system (CNS) toxicity. In CSF, 8OH-EFV levels were not influenced by CYP2B6 genotypes and did not predict CNS toxicity. The phase II metabolites 8OH-EFV-gln, 8OH-EFV-sulfate, and 7OH-EFV-gln were present in CSF at 2- to 9-fold higher concentrations than 8OH-EFV. The potential contribution of known and previously unreported EFV metabolites in CSF to the neuropsychological effects of efavirenz needs to be further examined in larger cohort studies.
There is an urgent unmet medical need for novel human immunodeficiency virus type 1 (HIV-1) inhibitors that are effective against a variety of NNRTI-resistance mutations. We report our research ...efforts aimed at discovering a novel chemotype of anti-HIV-1 agents with improved potency against a variety of NNRTI-resistance mutations in this paper. Structural modifications of the lead K-5a2 led to the identification of a potent inhibitor 16c. 16c yielded highly potent anti-HIV-1 activities and improved resistance profiles compared with the approved drug etravirine. The co-crystal structure revealed the key role of the water networks surrounding the NNIBP for binding and for resilience against resistance mutations, while suggesting further extension of 16c toward the NNRTI-adjacent site as a lead development strategy. Furthermore, 16c demonstrated favorable pharmacokinetic and safety properties, suggesting the potential of 16c as a promising anti-HIV-1 drug candidate.
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