Discovery of new acetamide derivatives of 5-indole-1,3,4-oxadiazol-2-thiol as inhibitors of HIV-1 Tat-mediated viral transcription.

Human immunodeficiency virus-1 (HIV-1) encodes a transcriptional factor called Tat, which is critical for viral transcription. Tat-mediated transcription is highly ordered apart from the cellular manner; therefore, it is considered a target for developing anti-HIV-1 drugs. However, drugs targeting Tat-mediated viral transcription are not yet available. Our high-throughput screen of a compound library employing a dual-reporter assay identified a 1,3,4-oxadiazole scaffold against Tat and HIV-1 infection. Furthermore, a serial structure-activity relation (SAR) study performed with biological assays found 1,3,4-oxadiazole derivatives (9 and 13) containing indole and acetamide that exhibited potent inhibitory effects on HIV-1 infectivity, with half-maximal effective concentrations (EC50) of 0.17 (9) and 0.24 µM (13), respectively. The prominent derivatives specifically interfered with the viral transcriptional step without targeting other infection step(s) and efficiently inhibited the HIV-1 replication cycle in the T cell lines and peripheral blood mononuclear cells infected with HIV-1. Additionally, compared to the wild type, the compounds exhibited similar potency against anti-retroviral drug-resistant HIV-1 strains. In a series of mode-of-action studies, the compounds inhibited the ejection of histone H3 for facilitating viral transcription on the long-terminal repeat (LTR) promoter. Furthermore, SAHA (a histone deacetylase inhibitor) treatment abolished the compound potency, revealing that the compounds can possibly target Tat-regulated epigenetic modulation of LTR to inhibit viral transcription. Overall, our screening identified novel 1,3,4-oxadiazole compounds that inhibited HIV-1 Tat, and subsequent SAR-based optimization led to the derivatives 9 and 13 development that could be a promising scaffold for developing a new class of therapeutic agents for HIV-1 infection.

In Silico Drug Repurposing Endorse Amprenavir, Darunavir and Saquinavir to Target Enzymes of Multidrug Resistant Uropathogenic E. Coli.

Multidrug resistance is a paramount impediment to successful treatment of most hospital acquired bacterial infections. A plethora of bacterial genera exhibit differential levels of resistance to the existing antibiotics. Prevalent Uropathogenic Escherichia coli or UPEC conduce high mortality among them. Multi-Drug Resistant bacterial strains utilize precise mechanisms to bypass effects of antibiotics. This is probably due to their familiar genomic origin. In this article drug repositioning method have been utilised to target 23 enzymes of UPEC strains viz. CFT073, 536 and UTI89. 3-D drug binding motifs have been predicted using SPRITE and ASSAM servers that compare amino acid side chain similarities. From the hit results anti-viral drugs have been considered for their uniqueness and specificity. Out of 14 anti-viral drugs 3 anti-HIV drugs viz. Amprenavir, Darunavir and Saquinavir have selected for maximum binding score or drug targetability. Finally, active sites of the enzymes were analyzed using GASS-WEB for eloquent drug interference. Further analyses with the active sites of all the enzymes showed that the three selected anti-HIV drugs were very much potent to inhibit their active sites. Combination or sole application of Amprenavir, Darunavir and Saquinavir to MDR-UPEC infections may leads to cure and inhibition of mortality. Supplementary Information: The online version contains supplementary material available at 10.1007/s12088-024-01282-x.

Efficacy and safety of artemether-lumefantrine as treatment for Plasmodium falciparum uncomplicated malaria in adult patients on efavirenz-based antiretroviral therapy in Zambia: an open label non-randomized interventional trial.

BACKGROUND: HIV-infected individuals on antiretroviral therapy (ART) require treatment with artemisinin-based combination therapy (ACT) when infected with malaria. Artemether-lumefantrine (AL) is the most commonly used ACT for treatment of falciparum malaria in Africa but there is limited evidence on the safety and efficacy of AL in HIV-infected individuals on ART, among whom drug-drug interactions are expected. Day-42 adequate clinical and parasitological response (ACPR) and incidence of adverse events was assessed in HIV-infected individuals on efavirenz-based ART with uncomplicated falciparum malaria treated with AL. METHODS: A prospective, open label, non-randomized, interventional clinical trial was conducted at St Paul's Hospital in northern Zambia, involving 152 patients aged 15-65 years with uncomplicated falciparum malaria, who were on efavirenz-based ART. They received a 3-day directly observed standard treatment of AL and were followed up until day 63. Day-42 polymerase chain reaction (PCR)-corrected ACPRs (95% confidence interval [CI]) were calculated for the intention-to-treat population. RESULTS: Enrolled patients had a baseline geometric mean (95% CI) parasite density of 1108 (841-1463) parasites/µL; 16.4% (25/152) of the participants had a recurrent malaria episode by day 42. However, PCR data was available for 17 out of the 25 patients who had malaria recurrence. Among all the 17 patients, PCR findings demonstrated malaria re-infection, making the PCR-adjusted day-42 ACPR 100% in the 144 patients who could be evaluated. Even when eight patients with missing PCR data were considered very conservatively as failures, the day-42 ACPR was over 94%. None of the participants, disease or treatment characteristics, including day-7 lumefantrine concentrations, predicted the risk of malaria recurrence by day 42. AL was well tolerated following administration. There were only two cases of grade 3 neutropaenia and one serious adverse event of lobar pneumonia, none of which was judged as probably related to intake of AL. CONCLUSIONS: AL was well tolerated and efficacious in treating uncomplicated falciparum malaria in HIV co-infected adults on efavirenz-based ART. However, a higher than anticipated proportion of participants experienced malaria re-infection, which highlights the need for additional malaria prevention measures in this sub-population after treatment with AL. Trial registration Pan African Clinical Trials Registry (PACTR): PACTR201311000659400. Registered on 4 October 2013. https://pactr.samrc.ac.za/Search.aspx.

The HIV-1 late domain-2 S40A polymorphism in antiretroviral (or ART)-exposed individuals influences protease inhibitor susceptibility.

BACKGROUND: The p6 region of the HIV-1 structural precursor polyprotein, Gag, contains two motifs, P7TAP11 and L35YPLXSL41, designated as late (L) domain-1 and -2, respectively. These motifs bind the ESCRT-I factor Tsg101 and the ESCRT adaptor Alix, respectively, and are critical for efficient budding of virus particles from the plasma membrane. L domain-2 is thought to be functionally redundant to PTAP. To identify possible other functions of L domain-2, we examined this motif in dominant viruses that emerged in a group of 14 women who had detectable levels of HIV-1 in both plasma and genital tract despite a history of current or previous antiretroviral therapy. RESULTS: Remarkably, variants possessing mutations or rare polymorphisms in the highly conserved L domain-2 were identified in seven of these women. A mutation in a conserved residue (S40A) that does not reduce Gag interaction with Alix and therefore did not reduce budding efficiency was further investigated. This mutation causes a simultaneous change in the Pol reading frame but exhibits little deficiency in Gag processing and virion maturation. Whether introduced into the HIV-1 NL4-3 strain genome or a model protease (PR) precursor, S40A reduced production of mature PR. This same mutation also led to high level detection of two extended forms of PR that were fairly stable compared to the WT in the presence of IDV at various concentrations; one of the extended forms was effective in trans processing even at micromolar IDV. CONCLUSIONS: Our results indicate that L domain-2, considered redundant in vitro, can undergo mutations in vivo that significantly alter PR function. These may contribute fitness benefits in both the absence and presence of PR inhibitor.

A lipid index for risk of hyperlipidemia caused by anti-retroviral drugs.

HIV-associated lipodystrophy has been reported in people taking anti-retroviral therapy (ART). Lipodystrophy can cause cardiovascular diseases, affecting the quality of life of HIV-infected individuals. In this study, we propose a pharmacological lipid index to estimate the risk of hyperlipidemia caused by anti-retroviral drugs. Lipid droplets were stained in cells treated with anti-retroviral drugs and cyclosporin A. Signal intensities of lipid droplets were plotted against the drug concentrations to obtain an isodose of 10 µM of cyclosporin A, which we call the Pharmacological Lipid Index (PLI). The PLI was then normalized by EC50. PLI/EC50 values were low in early proteinase inhibitors and the nucleoside reverse transcriptase inhibitor, d4T, indicating high risk of hyperlipidemia, which is consistent with previous findings of hyperlipidemia. In contrast, there are few reports of hyperlipidemia for drugs with high PLI/EC50 scores. Data suggests that PLI/EC50 is a useful index for estimating the risk of hyperlipidemia.

Development and In-House Validation of an Enzyme-Linked Immunosorbent Assay and a Lateral Flow Immunoassay for the Dosage of Tenofovir in Human Saliva.

Highly active antiretroviral therapy (HAART) includes very potent drugs that are often characterized by high toxicity. Tenofovir (TFV) is a widely used drug prescribed mainly for pre-exposure prophylaxis (PreP) and the treatment of human immunodeficiency virus (HIV). The therapeutic range of TFV is narrow, and adverse effects occur with both underdose and overdose. The main factor contributing to therapeutic failure is the improper management of TFV, which may be caused by low compliance or patient variability. An important tool to prevent inappropriate administration is therapeutic drug monitoring (TDM) of compliance-relevant concentrations (ARCs) of TFV. TDM is performed routinely using time-consuming and expensive chromatographic methods coupled with mass spectrometry. Immunoassays, such as enzyme-linked immunosorbent assays (ELISAs) and lateral flow immunoassays (LFIAs), are based on antibody-antigen specific recognition and represent key tools for real-time quantitative and qualitative screening for point-of-care testing (POCT). Since saliva is a non-invasive and non-infectious biological sample, it is well-suited for TDM. However, saliva is expected to have a very low ARC for TFV, so tests with high sensitivity are required. Here, we have developed and validated a highly sensitive ELISA (IC50 1.2 ng/mL, dynamic range 0.4-10 ng/mL) that allows the quantification of TFV in saliva at ARCs and an extremely sensitive LFIA (visual LOD 0.5 ng/mL) that is able to distinguish between optimal and suboptimal ARCs of TFV in untreated saliva.

HIV and SARS-CoV-2 Co-Infection: From Population Study Evidence to In Vitro Studies.

Human immunodeficiency virus type 1 (HIV-1) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have caused two major viral outbreaks during the last century. Two major aspects of HIV-1 and SARS-CoV-2 co-infection have been extensively investigated and deserve attention. First, the impact of the co-infection on the progression of disease caused by HIV-1 or SARS-CoV-2. Second, the impact of the HIV-1 anti-retroviral treatment on SARS-CoV-2 infection. In this review, we aim to summarize and discuss the works produced since the beginning of the SARS-CoV-2 pandemic ranging from clinical studies to in vitro experiments in the context of co-infection and drug development.

Scaffolding viral protein NC nucleates phase separation of the HIV-1 biomolecular condensate.

Membraneless biomolecular condensates (BMCs) contribute to the replication of a growing number of viruses but remain to be functionally characterized. Previously, we demonstrated that pan-retroviral nucleocapsid (NC) proteins phase separated into condensates regulating virus assembly. Here we discover that intrinsically disordered human immunodeficiency virus-type 1 (HIV-1) core proteins condense with the viral genomic RNA (vRNA) to assemble as BMCs attaining a geometry characteristic of viral reverse transcription complexes. We explore the predisposition, mechanisms, and pharmacologic sensitivity of HIV-1 core BMCs in living cells. HIV-1 vRNA-interacting NC condensates were found to be scaffolds onto which client capsid, reverse transcriptase, and integrase condensates assemble. HIV-1 core BMCs exhibit fundamental characteristics of BMCs and are drug-sensitive. Lastly, protease-mediated maturation of Gag and Gag-Pol precursor proteins yield abundant and visible BMCs in cells. This study redefines HIV-1 core components as fluid BMCs and advances our understanding of the nature of viral cores during ingress.

The Indian Network of Drug-Induced Liver Injury: Etiology, Clinical Features, Outcome and Prognostic Markers in 1288 Patients.

BACKGROUND: Etiology of and outcomes following idiosyncratic drug-induced liver injury (DILI) vary geographically. We conducted a prospective study of DILI in India, from 2013 to 2018 and summarize the causes, clinical features, outcomes and predictors of mortality. METHODS: We enrolled patients with DILI using international DILI expert working group criteria and Roussel Uclaf causality assessment method. Follow-up was up to 3 months from onset of DILI or until death. Multivariate logistics regression was carried out to determine predictors of non-survival. RESULTS: Among 1288 patients with idiosyncratic DILI, 51.4% were male, 68% developed jaundice, 68% required hospitalization and 8.2% had co-existing HIV infection. Concomitant features of skin reaction, ascites, and encephalopathy (HE) were seen in 19.5%, 16.4%, and 10% respectively. 32.4% had severe disease. Mean MELD score at presentation was 18.8 ± 8.8. Overall mortality was 12.3%; 65% in those with HE, 17.6% in patients who fulfilled Hy's law, and 16.6% in those that developed jaundice. Combination anti-TB drugs (ATD) 46.4%, complementary and alternative medicines (CAM) 13.9%, anti-epileptic drugs (AED) 8.1%, non-ATD antimicrobials 6.5%, anti-metabolites 3.8%, anti-retroviral drugs (ART)3.5%, NSAID2.6%, hormones 2.5%, and statins 1.4% were the top 9 causes. Univariate analysis identified, ascites, HE, serum albumin, bilirubin, creatinine, INR, MELD score (p < 0.001), transaminases (p < 0.04), and anti-TB drugs (p = 0.02) as predictors of non-survival. Only serum creatinine (p = 0.017), INR (p < 0.001), HE (p < 0.001), and ascites (p = 0.008), were significantly associated with mortality on multivariate analysis. ROC yielded a C-statistic of 0.811 for MELD and 0.892 for combination of serum creatinine, INR, ascites and HE. More than 50 different agents were associated with DILI. Mortality varied by drug class: 15% with ATD, 13.6% with CAM, 15.5% with AED, 5.8% with antibiotics. CONCLUSION: In India, ATD, CAM, AED, anti-metabolites and ART account for the majority of cases of DILI. The 3-month mortality was approximately 12%. Hy's law, presence of jaundice or MELD were predictors of mortality.

Dissecting the Therapeutic Relevance of Gene Therapy in NeuroAIDS: An Evolving Epidemic.

NeuroAIDS, a disease incorporating both infectious and neurodegenerative pathways, is still a formidable challenge for the researchers to deal with. The primary concern for the treatment of neuroAIDS still remains the inaccessibility of the viral reservoir, making it indispensable for novel techniques to be continuously innovated. Since the brain serves as a reservoir for viral replication, it is pragmatic and a prerequisite to overcome the related barriers in order to improve the drug delivery to the brain. The current treatment ideology is based on the combinatorial approach of a mocktail of antiretroviral drugs. However, complete eradication of the disease could not be achieved. Thereby the arena of gene-based cellular delivery is trending and has created a niche for itself in the present scenario. To establish the supremacy of gene delivery, it is advisable to have a better understanding of the molecular mechanism involved in the due process. The mechanism associated with the activity of the anti-HIV gene lies in their intrinsic property to impart resistance to the HIV infection by targeting the viral entry channels. This review principally emphasizes on different types of gene therapies explored so far for the management of AIDS and its associated neurological conditions. Therefore it could rightly be said that we are at the crossroad where the need of the hour is to develop novel strategies for curbing AIDS and its associated neurological conditions.

Croton megalobotrys Müll Arg. and Vitex doniana (Sweet): Traditional medicinal plants in a three-step treatment regimen that inhibit in vitro replication of HIV-1.

ETHNOPHARMACOLOGICAL RELEVANCE: Human Immunodeficiency Virus (HIV) strains resistant to licensed anti-retroviral drugs (ARVs) continue to emerge. On the African continent, uneven access to ARVs combined with occurrence of side-effects after prolonged ARV therapy have led to searches for traditional medicines as alternative or complementary remedies to conventional HIV/AIDS management. AIM OF THE STUDY: Here we characterize a specific three-step traditional HIV/AIDS treatment regimen consisting of Cassia sieberiana root, Vitex doniana root, and Croton megalobotrys bark by combining qualitative interviews of traditional medical knowledge users in Botswana with in vitro HIV replication studies. MATERIALS AND METHODS: Crude extracts from a total of seven medicinal plants were tested for in vitro cytotoxicity and inhibition of wild-type (NL4.3) and ARV-resistant HIV-1 replication in an immortalized GFP-reporter CD4+ T-cell line. RESULTS: C. sieberiana root, V. doniana root, and C. megalobotrys bark extracts inhibited HIV-1NL4.3 replication with dose-dependence and without concomitant cytotoxicity. C. sieberiana and V. doniana extracts inhibited HIV-1 replication by 50% at 84.8µg/mL and at 25µg/mL, respectively, while C. megalobotrys extracts inhibited HIV-1 replication by a maximum of 45% at concentrations as low as 0.05µg/mL. Extracts did not interfere with antiviral activities of licensed ARVs when applied in combination and exhibited comparable efficacies against viruses harboring major resistance mutations to licensed protease, reverse-transcriptase, or integrase inhibitors. CONCLUSIONS: We report for the first time a three-step traditional HIV/AIDS regimen, used alone or in combination with standard ARV regimens, where each step exhibited more potent ability to inhibit HIV replication in vitro. Our observations support the "reverse pharmacology" model where documented clinical experiences are used to identify natural products of therapeutic value.

Quantification of darunavir and etravirine in human peripheral blood mononuclear cells using high performance liquid chromatography tandem mass spectrometry (LC-MS/MS), clinical application in a cohort of 110 HIV-1 infected patients and evidence of a potential drug-drug interaction.

OBJECTIVES: To describe the validation of a sensitive high performance liquid chromatography tandem mass spectrometry (LC-MS/MS) method allowing the simultaneous quantification of darunavir (DRV) and etravirine (ETR) in peripheral blood mononuclear cells (PBMCs) and its application in a cohort of HIV-1 infected patients. METHODS: Blood samples were obtained from 110 patients. PMBCs were isolated using density gradient centrifugation. Drug extraction from PBMCs was performed with a 60:40 methanol-water (MeOH-H2O) solution containing deuterated IS (DRV-d9 and ETR-d8). The chromatographic separation was performed on a RP18 XBridge™ column. RESULTS: The geometric mean (GM) of cell associated concentration ([DRV]CC) and plasmatic concentration ([DRV]plasma) were 360.5ng/mL (CI95%:294.5-441.2) and 1733ng/mL (CI95%:1486-2021), respectively. A geometric mean intracellular (IC)/plasma ratio (GMR) of 0.21 (CI95%:0.18-0.24) was calculated. Adjusted for dose/body surface area and post-intake time, a statistically significant correlation was observed between [DRV]Plasma and the eGFR (p=0.002) and between [DRV]Plasma and the concomitant use of ETR (p=0.038). For the 10 patients receiving ETR in addition to DRV, the GM of [ETR]Plasma (available for 8 out of 10 patients) and [ETR]CC were 492.3ng/mL and 2951ng/mL respectively. The GMR of ETR was 7.6 (CI95%: 3.61-13.83). CONCLUSIONS: A handy and sensitive high performance LC-MS/MS method allowing the simultaneous quantification of DRV and ETR in PBMCs has been described and successfully applied in the largest cohort of DRV-treated patients reported to date. ETR accumulates more efficiently in PBMCs compared to DRV. We have also highlighted a possible impact of ETR on DRV plasma concentrations requiring further investigations.

Preparation of molecularly imprinted solid-phase microextraction fiber for the selective removal and extraction of the antiviral drug abacavir in environmental and biological matrices.

In the present study, a molecularly imprinted solid-phase microextraction fiber (MIP-SPMEf) was synthesized and applied for the selective removal and extraction of the antiviral drug, abacavir (ABA). Morphology and structure characterization of fibers were performed by scanning electron microscopy and Fourier transform infrared spectra, respectively. The effects on the adsorption behavior of the process parameters were studied and the equilibrium data were fitted by the Langmuir, Freundlich and Langmuir-Freundlich models. The maximum adsorption capability (Qmax) was determined by Langmuir- Freundlich model and was 149 mg/g for MIP-SPMEf. In the next step, SPME methodology followed by liquid desorption and liquid chromatography with mass spectrometry (LC/MS) has been developed and evaluated for the determination of the target compound in environmental and biological matrices (surface waters, wastewaters and urine). Parameters that could influence SPME efficiency were investigated. Then, optimization of stirring speed, extraction time and salt content was carried out by using a central composite design (CCD) and response surface methodology (RSM). A quadratic model between dependent and independent variables was built. Under the optimum conditions (extraction time 40 min, stirring rate 650 rpm and salt content 0.3% NaCl w/v) the validated method presented a high sensitivity and selectivity with LODs and LOQs in the range of 10.1-13.6 and 33.3-43.9 ng/L, respectively. The developed method was successfully applied to the analysis of ABA in real samples. The percentage extraction efficiency ranged from 88 to 99% revealing good accuracy and absence of matrix effects.

Recent advances of resveratrol in nanostructured based delivery systems and in the management of HIV/AIDS.

Resveratrol, a natural polyphenolic compound present in trees, in peanuts, in grapevines and exhibited multiple pharmacological activities. Extensive research in last two decades suggested that resveratrol possesses anti-inflammatory, anti-cancer, anti-viral, anti-amyloid, anti-arthritic and antioxidant properties. Some clinical reports have proposed that resveratrol might be a potential candidate for the prevention and/or treatment of HIV/AIDS and synergistically enhances the anti-HIV-1 activity. Resveratrol is not toxic to cells, and by itself reduces viral replication by 20% to 30%. With almost 12% of the world population suffering from HIV/AIDS including its resurgence in the developed world, better management of this global threat is highly desired. Further, various studies demonstrated several issues associated with resveratrol which account for its poor systemic bioavailability (almost zero) due to rapid and extensive first pass metabolism and existence of enterohepatic recirculation. In order to improve bioavailability and cellular uptake of resveratrol, various strategies have been adopted to date which includes resveratrol prodrug and the development of nanostructured delivery systems. Besides, nanostructured delivery systems are also known to inhibit the P-glycoprotein (P-gp) efflux, reduced metabolism by gut cytochrome P-450 enzymes, and circumnavigate the hepatic first-pass effect, facilitating absorption of drugs via intestinal lymphatic pathways. This review paper provides an updated bird's-eye view account on the publications and patents study on the recent novel approaches to deliver resveratrol in order to enhance oral bioavailability, overcome first pass metabolism and trounce enterohepatic recirculation to make resveratrol a therapeutically potent drug. Providing a relatively pithy overview, this paper thus presents recent advances of resveratrol for the treatment and prevention of HIV/AIDS.

Heart Disease in Patients with HIV/AIDS-An Emerging Clinical Problem.

HIV/AIDS (Human immunodeficiency virus/ Acquired immuno deficiency syndrome) is a growing global problem, in terms of its incidence and mortality. Patients with HIV/AIDS are living much longer with HAART (Highly active antiretroviral therapy) therapy so much so that HIV/AIDS has now become a part of the chronic disease burden, like hypertension and diabetes. Patients with HIV/AIDS and symptoms suggestive of cardiac disease represent a diagnostic and therapeutic challenge in clinical practice; Cardiologists are more frequently encountering this problem. An algorithmic, anatomic approach to diagnosis, localizing disease to the endocardium, myocardium and pericardium can be useful. An intimate knowledge of opportunistic infections affecting the heart, effects of HAART therapy and therapy for opportunistic infections on the heart is needed to be able to formulate a differential diagnosis. Effects of HAART therapy, especially protease inhibitors on lipid and glucose metabolism, and their influence on progression to premature vascular disease require consideration. Treatment of cardiac disease, in HIV/AIDS patients can vary from non-HIV patients, based on drug interactions, differences in responsiveness, and other factors; and this area requires further research.