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Chantal Emade Nkwelle,1,2 Smith B Babiaka,3,4 Clovis S Metuge,2,3 Kimberly Liang,5 Unique Stephens,5 Seraphine Nkie Esemu,1 David S Zuzga,5 Kristy Shuda McGuire,5 Luis J Montaner,5 Roland N Ndip,1 Ian Tietjen,5 Fidele Ntie-Kang2,3,6 1Department of Microbiology and Parasitology, Faculty of Science, University of Buea, Buea, Cameroon; 2Center for Drug Discovery, Faculty of Science, University of Buea, Buea, Cameroon; 3Department of Chemistry, Faculty of Science, University of Buea, Buea, Cameroon; 4Department of Microbial Bioactive Compound, University of Tübingen, Tübingen, Germany; 5Drug Discovery, The Wistar Institute of Anatomy and Biology, Philadelphia, PA, USA; 6Institute of Pharmacy, Martin-Luther University Halle-Wittenberg, Halle, GermanyCorrespondence: Ian Tietjen, The Wistar Institute, 3601 Spruce Street, Philadelphia, PA, 19104, USA, Tel +1-215-898-3938, Email itietjen@wistar.org Fidele Ntie-Kang, Center for Drug Discovery, Faculty of Science, University of Buea, P. O. Box 63, Buea, Cameroon, Tel +237-673872475, Email fidele.ntie-kang@ubuea.cmBackground: Croton oligandrus Pierre & Hutch is a tropical tree that grows in West and Central Africa, used in ethnomedicine to treat cancer, diabetes, headaches, convulsions, urinary diseases, and inflammatory diseases. As other Croton species have been observed to possess chemical compounds that target HIV latency-reversal, we hypothesized that this species may have similar properties.Aim of the Study: The identification of extracts and compounds of this species, which have HIV-1 latency-reversing activity in J-Lat T cell lines.Methods: The stem bark was obtained, air-dried, powdered, and extracted using dichloromethane. In vitro flow cytometry was used to monitor GFP expression, a marker of HIV latency reversal, following treatment of J-Lat T cells with extracts and compounds.Results: Four extracts were found to reverse HIV latency, the most active extract showing better activity (ie, latency reversal in 69.7 ± 7.1% [mean ± s.e.m.] of J-Lat 10.6 cells at 1 μg/mL) than control agents prostratin (46.2 ± 9.5% at 1.2 μg.mL) and the “Mukungulu” (Croton megalobotrys) extract (34.9 ± 24.2% at 1 μg/mL). Extracts reversed HIV latency through mechanisms over and above protein kinase C (PKC) activation and distinct from histone deacetylase (HDAC) inhibition. The most active extract also synergized with the control HDAC inhibitor romidepsin but did not synergize with other extracts. Isolated compounds (β-Stigmasterol and lupeol) had limited but consistent latency reversal on their own.Conclusion: The plant extracts and compounds reverse HIV latency through mechanisms additional to PKC activation and/or synergize with romidepsin in vitro. Extracts and compounds from this plant may enhance the activity of current HIV latency-reversing agents being assessed in HIV cure studies.Keywords: antiviral, compounds, extracts, HIV-1, latency reversal, screening

New therapeutic strategies are needed for malignant pleural mesothelioma (MPM). We conducted a single-center, open-label, nonrandomized, pilot and feasibility trial using two intrapleural doses of an adenoviral vector encoding human IFN-α (Ad.IFN-α2b). Nine subjects were enrolled at two dose levels. The first three subjects had very high pleural and systemic IFN-α concentrations resulting in severe "flu-like" symptoms necessitating dose de-escalation. The next six patients had reduced (but still significant) pleural and serum IFN-α levels, but with tolerable symptoms. Repeated vector administration appeared to prolong IFN-α expression levels. Anti-tumor humoral immune responses against mesothelioma cell lines were seen in seven of the eight subjects evaluated. No clinical responses were seen in the four subjects with advanced disease. However, evidence of disease stability or tumor regression was seen in the remaining five patients, including one dramatic example of partial tumor regression at sites not in contiguity with vector infusion. These data show that Ad.IFN-α2b has potential therapeutic benefit in MPM and that it generates anti-tumor immune responses that may induce anatomic and/or metabolic reductions in distant tumor. Clinical trial registered with www.clinicaltrials.gov (NCT 01212367). [ABSTRACT FROM AUTHOR]

J-Lat cells are derivatives of the Jurkat CD4 + T cell line that contain a non-infectious, inducible HIV provirus with a GFP tag. While these cells have substantially advanced our understanding of HIV latency, their use by many laboratories in low and middle-income countries is restricted by limited access to flow cytometry. To overcome this barrier, we describe a modified J-Lat assay using a standard microplate reader that detects HIV-GFP expression following treatment with latency-reversing agents (LRAs). We show that HIV reactivation by control LRAs like prostratin and romidepsin is readily detected with dose dependence and with significant correlation and sensitivity to standard flow cytometry. For example, 10 μM prostratin induced a 20.1 ± 3.3-fold increase in GFP fluorescence in the microplate reader assay, which corresponded to 64.2 ± 5.0% GFP-positive cells detected by flow cytometery. Similarly, 0.3 μM prostratin induced a 1.7 ± 1.2-fold increase compared to 8.7 ± 5.7% GFP-positive cells detected. Using this method, we screen 79 epigenetic modifiers and identify CUDC-101, molibresib, and quisinostat as novel LRAs. This microplate reader-based method offers accessibility to researchers in resource-limited regions to work with J-Lat cells and more actively participate in global HIV cure research efforts., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025 Elsevier Inc. All rights reserved.)

The only current strategy to test efficacy of novel interventions for sustained HIV control without antiretroviral therapy (ART) among people with HIV (PWH) is through an analytical treatment interruption (ATI). Inclusion of 'placebo' controls in ATIs poses ethical, logistical, and economic challenges. To understand viral dynamics and rates of post-treatment control (PTC) after ATI among PWH receiving either placebo or no intervention, we undertook an individual-participant data meta-analysis. In total, 24 eligible prospective studies with 382 individuals with ≥5 plasma HIV RNA viral loads (pVLs) within the first 84 days post-ATI were included. Early-ART was defined as ART initiation within 6 months of HIV acquisition; others were classified as late-ART or unknown. Median age was 42 years, 91% male, 75% white, 45% received early-ART. Median time to pVL >50, >400, and >10,000 copies/mL was 16 days (interquartile range [IQR]:13-25), 21 (IQR:15-28), and 32 (IQR:20-35), respectively. PTC defined as pVL <50 copies/mL at day 84 occurred in 4% (n = 14) of participants (6% early-ART and 1% late-ART). Sustained PTC of pVL <50 copies/ml after 84 days is rare in PWH, especially in those starting ART late. Our findings inform future interventional HIV cure/remission trials on study size and design., Competing Interests: Competing interests: The authors declare no conflict of interest., (© 2025. The Author(s).)

Transcription repressor BACH2 redirects short-lived terminally differentiated effector into long-lived memory cells. We postulate that BACH2-mediated long-lived memory programs promote HIV-1 persistence in gut CD4+ T cells. We coupled single-cell DOGMA-seq and TREK-seq to capture chromatin accessibility, transcriptome, surface proteins, T cell receptor, HIV-1 DNA and HIV-1 RNA in 100,744 gut T cells from ten aviremic HIV-1+ individuals and five HIV-1- donors. BACH2 was the leading transcription factor that shaped gut tissue resident memory T cells (TRMs) into long-lived memory with restrained interferon-induced effector function. We found that HIV-1-infected cells were enriched in TRMs (80.8%). HIV-1-infected cells had increased BACH2 transcription factor accessibility, TRM (CD49a, CD69, CD103) and survival ( IL7R ) gene expression, and Th17 polarization (RORC, CCR6 ). In vitro gut CD4+ T cell infection revealed preferential infection and persistence of HIV-1 in CCR6+ TRMs. Overall, we found BACH2-driven TRM program promotes HIV-1 persistence and BACH2 as a new therapeutic target., Competing Interests: DECLARATION OF INTERESTS The authors have no competing interests to declare.

Despite antiretroviral therapy (ART), HIV-1 persists in latently infected CD4 + T cells, preventing a cure. Antigens drive the proliferation of infected cells, precluding latent reservoir decay. However, the relationship between antigen recognition and HIV-1 gene expression is poorly understood because most studies of latency reversal use agents that induce non-specific global T cell activation. Here, we isolated rare CD4 + T cells responding to cytomegalovirus (CMV) or HIV-1 Gag antigens from people living with HIV-1 on long-term ART and assessed T cell activation and HIV-1 RNA expression upon coculture with autologous dendritic cells (DCs) presenting cognate antigens. Presentation of cognate antigens ex vivo induced broad T cell activation (median 42-fold increase in CD154 + CD69 + cells) and significantly increased HIV-1 transcription (median 4-fold), mostly through the induction of rare cells with higher viral expression. Thus, despite low proviral inducibility, antigen recognition can promote HIV-1 expression, potentially contributing to spontaneous reservoir activity and viral rebound upon ART interruption., Competing Interests: Declaration of interests R.F.S. is one of the inventors of a patent application involving aspects of the IPDA entitled “Compositions and methods Related to characterizing proviral reservoirs,” PCT/US16/28822, filed by Johns Hopkins University. F.R.S. received payments from Gilead Sciences for participating at scientific meetings. F.W. is currently an employee of Gilead Sciences. N.L.B. is currently a postdoctoral associate in medicine at Weill Cornell Medical College., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Ethical patient outreach is critical for engaging patients with HIV in HIV cure-directed research. We sought to examine HIV clinical providers' awareness of current HIV cure-directed research strategies investigated through the Martin Delaney Collaboratories (MDC) and providers' attitudes toward patient outreach for HIV cure-directed research and to identify opportunities for clinical provider education on MDC research strategies., Methods: We conducted a 1-time, cross-sectional, web-based survey with 64 HIV clinical providers (physicians, physician assistants, and nurses) in Philadelphia. A descriptive analysis was generated to determine clinical providers' awareness of MDC research strategies and attitudes toward patient outreach. Bivariate analyses were conducted to evaluate differences in awareness and attitudes by provider characteristics., Results: Most providers were aware of MDC strategies, and nearly three-fourths of providers reported familiarity with 1 of the 2 Philadelphia MDC research programs. Providers overwhelmingly endorsed the need for clinicians to assist with patient outreach for HIV cure-directed research and were willing to recommend patients to participate. Enthusiasm for patient outreach waned if a study required a patient to undergo analytic treatment interruptions (ATIs). Providers identified preferred resources for receiving HIV cure-directed research updates, including webinars with continuing education credit and conferences., Conclusions: Our study's findings advocate for added investment toward developing resources that better engage clinical providers about HIV cure-directed research updates and ongoing studies, including the importance of ATIs. As gatekeepers to patient participation on HIV cure-directed studies, added efforts to increase provider knowledge of specific HIV cure-directed research will advance patient education and ethical outreach., Competing Interests: Potential conflicts of interest. All authors: No reported conflicts of interest., (© The Author(s) 2024. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

We have synthesized a novel and highly selective severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main protease peptide mimetic inhibitor mimicking the replicase 1ab recognition sequence -Val-Leu-Gln- and utilizing a cysteine selective acyloxymethyl ketone as the electrophilic warhead to target the active site Cys145. Utilizing a constrained cyclic peptide that locks the conformation between the P3 (Val) and P2 (Leu) residues, we identified a highly selective inhibitor that fills the P2 pocket occupied by the leucine residue sidechain of PF-00835231 and the dimethyl-3-azabicyclo-hexane motif in nirmatrelvir (PF-07321332). This strategy resulted in potent and highly selective Mpro inhibitors without inhibiting essential host cathepsin cysteine or serine proteases. The lead prototype compound 1 (MPro IC 50 = 230 ± 18 nM) also inhibits the replication of multiple SARS-CoV-2 variants in vitro , including SARS-CoV-2 variants of concern, and can synergize at lower concentrations with the viral RNA polymerase inhibitor, remdesivir, to inhibit replication. It also reduces SARS-CoV-2 replication in SARS-CoV-2 Omicron-infected Syrian golden hamsters without obvious toxicities, demonstrating in vivo efficacy. This novel lead structure provides the basis for optimization of improved agents targeting evolving SARS-CoV-2 drug resistance that can selectively act on Mpro versus host proteases and are less likely to have off-target effects due to non-specific targeting. Developing inhibitors against the active site of the main protease (Mpro), which is highly conserved across coronaviruses, is expected to impart a higher genetic barrier to evolving SARS-CoV-2 drug resistance. Drugs that selectively inhibit the viral Mpro are less likely to have off-target effects warranting efforts to improve this therapy., Competing Interests: J.M.S. receives funding from the NIH P30-CA010815-53; has patents pending to Wistar; owns equity in Alliance Discovery, Inc., Barer Institute, and Context Therapeutics; and consults for Syndeavor Therapeutics, Inc. All other authors declare no competing interests.

Type I IFNs play a pivotal role in immune response modulation, yet dysregulation is implicated in various disorders. Therefore, it is crucial to develop tools that facilitate the understanding of their mechanism of action and enable the development of more effective anti-IFN therapeutic strategies. In this study, we isolated, cloned, and characterized anti-IFN-α and anti-IFN-β Abs from PBMCs of individuals treated with IFN-α or IFN-β, harboring confirmed neutralizing Abs. Clones AH07856 and AH07857 were identified as neutralizing anti-IFN-α-specific with inhibition against IFN-α2a, -α2b, and -αK subtypes. Clones AH07859 and AH07866 were identified as neutralizing anti-IFN-β1a-specific signaling and able to block lipopolysaccharide or S100 calcium-binding protein A14-induced IFN-β signaling effects. Cloned Abs bind rhesus but not murine IFNs. The specificity of inhibition between IFN-α and IFN-β suggests potential for diverse research and clinical applications., (Copyright © 2024 by The American Association of Immunologists, Inc.)

J-Lat cells are derivatives of the Jurkat CD4+ T cell line that contain a non-infectious, inducible HIV provirus with a GFP tag. While these cells have substantially advanced our understanding of HIV latency, their use by many laboratories in low and middle-income countries is restricted by limited access to flow cytometry. To overcome this barrier, we describe a modified J-Lat assay using a standard microplate reader that detects HIV-GFP expression following treatment with latency-reversing agents (LRAs). We show that HIV reactivation by control LRAs like prostratin and romidepsin is readily detected with dose dependence and with significant correlation and sensitivity to standard flow cytometry. For example, 10 µM prostratin induced a 20.1 ± 3.3-fold increase in GFP fluorescence in the microplate reader assay, which corresponded to 64.2 ± 5.0% GFP-positive cells detected by flow cytometery. Similarly, 0.3 µM prostratin induced a 1.7 ± 1.2-fold increase compared to 8.7 ± 5.7% GFP-positive cells detected. Using this method, we screen 79 epigenetic modifiers and identify molibresib, quisinostat, and CUDC-101 as novel LRAs. This microplate reader-based method offers accessibility to researchers in resource-limited regions to work with J-Lat cells and more actively participate in global HIV cure research efforts., Highlights: J-Lat T-cell lines are important to HIV cure research but require flow cytometryWe describe a method to work with J-Lat cells using a standard microplate readerThis assay can detect control LRAs similar to flow cytometry and discover new LRAsThis assay allows low-resourced laboratories to contribute to HIV cure research.

Delta (B.1.617.2) and Omicron (B.1.1.529) variants of SARS-CoV-2 represents unique clinical characteristics. However, their role in altering immunometabolic regulations during acute infection remains convoluted. Here, we evaluated the differential immunopathogenesis of Delta vs. Omicron variants in Golden Syrian hamsters (GSH). The Delta variant resulted in higher virus titers in throat swabs and the lungs and exhibited higher lung damage with immune cell infiltration than the Omicron variant. The gene expression levels of immune mediators and metabolic enzymes, Arg-1 and IDO1 in the Delta-infected lungs were significantly higher compared to Omicron. Further, Delta/Omicron infection perturbed carbohydrates, amino acids, nucleotides, and TCA cycle metabolites and was differentially regulated compared to uninfected lungs. Collectively, our data provide a novel insight into immunometabolic/pathogenic outcomes for Delta vs. Omicron infection in the GSH displaying concordance with COVID-19 patients associated with inflammation and tissue injury during acute infection that offered possible new targets to develop potential therapeutics., Competing Interests: The authors declared no competing interest., (© 2024 The Author(s).)

The coronavirus disease 2019 (COVID-19) has spread worldwide with severe health, social, and economic repercussions. Although vaccines have significantly reduced the severity of symptoms and deaths, alternative medications derived from natural products (NPs) are vital to further decrease fatalities, especially in regions with low vaccine uptake. When paired with the latest computational developments, NPs, which have been used to cure illnesses and infections for thousands of years, constitute a renewed resource for drug discovery. In the present report, a combination of computational and in vitro methods reveals the repositioning of NPs and identifies salvinorin A and deacetylgedunin (DCG) as having potential anti-SARS-CoV-2 activities. Salvinorin A was found both in silico and in vitro to inhibit both SARS-CoV-2 spike/host ACE2 protein interactions, consistent with blocking viral cell entry, and well as live virus replication. Plant extracts from Azadirachta indica and Cedrela odorata , which contain high levels of DCG, inhibited viral cell replication by targeting the main protease (Mpro) and/or inhibited viral cell entry by blocking the interaction between spike RBD-ACE2 protein at concentrations lower than salvinorin A. Our findings suggest that salvinorin A represent promising chemical starting points where further optimization may result in effective natural product-derived and potent anti-SARS-CoV-2 inhibitors to supplement vaccine efforts., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Current small-molecule-based SARS-CoV-2 treatments have limited global accessibility and pose the risk of inducing viral resistance. Therefore, a marine algae and cyanobacteria extract library was screened for natural products that could inhibit two well-defined and validated COVID-19 drug targets, disruption of the spike protein/ACE-2 interaction and the main protease (M pro ) of SARS-CoV-2. Following initial screening of 86 extracts, we performed an untargeted metabolomic analysis of 16 cyanobacterial extracts. This approach led to the isolation of an unusual saturated fatty acid, jobosic acid (2,5-dimethyltetradecanoic acid, 1 ). We confirmed that 1 demonstrated selective inhibitory activity toward both viral targets while retaining some activity against the spike-RBD/ACE-2 interaction of the SARS-CoV-2 omicron variant. To initially explore its structure-activity relationship (SAR), the methyl and benzyl ester derivatives of 1 were semisynthetically accessed and demonstrated acute loss of bioactivity in both SARS-CoV-2 biochemical assays. Our efforts have provided copious amounts of a fatty acid natural product that warrants further investigation in terms of SAR, unambiguous determination of its absolute configuration, and understanding of its specific mechanisms of action and binding site toward new therapeutic avenues for SARS-CoV-2 drug development.

Ovarian cancer remains a major health threat with limited treatment options available. It is characterized by immunosuppressive tumor microenvironment (TME) maintained by tumor- associated macrophages (TAMs) hindering anti-tumor responses and immunotherapy efficacy. Here we show that targeting retinoblastoma protein (Rb) by disruption of its LxCxE cleft pocket, causes cell death in TAMs by induction of ER stress, p53 and mitochondria-related cell death pathways. A reduction of pro-tumor Rb high M2-type macrophages from TME in vivo enhanced T cell infiltration and inhibited cancer progression. We demonstrate an increased Rb expression in TAMs in women with ovarian cancer is associated with poorer prognosis. Ex vivo, we show analogous cell death induction by therapeutic Rb targeting in TAMs in post-surgery ascites from ovarian cancer patients. Overall, our data elucidates therapeutic targeting of the Rb LxCxE cleft pocket as a novel promising approach for ovarian cancer treatment through depletion of TAMs and re-shaping TME immune landscape., Statement of Significance: Currently, targeting immunosuppressive myeloid cells in ovarian cancer microenvironment is the first priority need to enable successful immunotherapy, but no effective solutions are clinically available. We show that targeting LxCxE cleft pocket of Retinoblastoma protein unexpectedly induces preferential cell death in M2 tumor-associated macrophages. Depletion of immunosuppressive M2 tumor-associated macrophages reshapes tumor microenvironment, enhances anti-tumor T cell responses, and inhibits ovarian cancer. Thus, we identify a novel paradoxical function of Retinoblastoma protein in regulating macrophage viability as well as a promising target to enhance immunotherapy efficacy in ovarian cancer.

Type I interferons (IFNs) play a pivotal role in immune response modulation, yet dysregulation is implicated in various disorders. Therefore, it is crucial to develop tools that facilitate the understanding of their mechanism of action and enable the development of more effective anti-IFN therapeutic strategies. In this study, we isolated, cloned, and characterized anti-IFN-α and anti-IFN-β antibodies (Abs) from peripheral blood mononuclear cells of individuals treated with IFN-α or IFN-β, harboring confirmed neutralizing Abs. Clones AH07856 and AH07857 were identified as neutralizing anti-IFN-α-specific with inhibition against IFN-α2a, -α2b, and -αK subtypes. Clones AH07859 and AH07866 were identified as neutralizing anti-IFN-β1a-specific signaling, and able to block Lipopolysaccharide or S100 calcium binding protein A14-induced IFN-β signaling effects. Cloned Abs bind rhesus but not murine IFNs. The specificity of inhibition between IFN-α and IFN-β suggests potential for diverse research and clinical applications.

Seven furanochromene-quinoline derivatives containing a hydrazone linker were synthesized by condensing a furanochromene hydrazide with quinoline 2-, 3-, 4-, 5-, 6-, and 8-carbaldehydes, including 8-hydroxyquinoline-2-carbaldehye. Structure-activity correlations were investigated to determine the influence of the location of the hydrazone linker on the quinoline unit on SARS-CoV-2 M pro enzyme inhibition. The 3-, 5-, 6- and 8-substituted derivatives showed moderate inhibition of SARS-CoV-2 M pro with IC 50 values ranging from 16 to 44 μM. Additionally, all of the derivatives showed strong interaction with the SARS-CoV-2 M pro substrate binding pocket, with docking energy scores ranging from -8.0 to -8.5 kcal/mol. These values are comparable to that of N3 peptide (-8.1 kcal/mol) and more favorable than GC-373 (-7.6 kcal/mol) and ML-188 (-7.5 kcal/mol), all of which are known SARS-CoV-2 M pro inhibitors. Furthermore, in silico absorption, distribution, metabolism, and excretion (ADME) profiles indicate that the derivatives have good drug-likeness properties. Overall, this study highlights the potential of the furanochromene-quinoline hydrazone scaffold as a SARS-CoV-2 M pro inhibitor., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The persistence of CD4 + T cells carrying latent human immunodeficiency virus-1 (HIV-1) proviruses is the main barrier to a cure. New therapeutics to enhance HIV-1-specific immune responses and clear infected cells will probably be necessary to achieve reduction of the latent reservoir. In the present study, we report two single-chain diabodies (scDbs) that target the HIV-1 envelope protein (Env) and the human type III Fcγ receptor (CD16). We show that the scDbs promoted robust and HIV-1-specific natural killer (NK) cell activation and NK cell-mediated lysis of infected cells. Cocultures of CD4 + T cells from people with HIV-1 on antiretroviral therapy (ART) with autologous NK cells and the scDbs resulted in marked elimination of reservoir cells that was dependent on latency reversal. Treatment of human interleukin-15 transgenic NSG mice with one of the scDbs after ART initiation enhanced NK cell activity and reduced reservoir size. Thus, HIV-1-specific scDbs merit further evaluation as potential therapeutics for clearance of the latent reservoir., (© 2024. The Author(s).)

The latent reservoir of HIV persists for decades in people living with HIV (PWH) on antiretroviral therapy (ART). To determine if persistence arises from the natural dynamics of memory CD4+ T cells harboring HIV, we compared the clonal dynamics of HIV proviruses to that of memory CD4+ T cell receptors (TCRβ) from the same PWH and from HIV-seronegative people. We show that clonal dominance of HIV proviruses and antigen-specific CD4+ T cells are similar but that the field's understanding of the persistence of the less clonally dominant reservoir is significantly limited by undersampling. We demonstrate that increasing reservoir clonality over time and differential decay of intact and defective proviruses cannot be explained by mCD4+ T cell kinetics alone. Finally, we develop a stochastic model of TCRβ and proviruses that recapitulates experimental observations and suggests that HIV-specific negative selection mediates approximately 6% of intact and 2% of defective proviral clearance. Thus, HIV persistence is mostly, but not entirely, driven by natural mCD4+ T cell kinetics.

Chemical prototypes with broad-spectrum antiviral activity are important toward developing new therapies that can act on both existing and emerging viruses. Binding of the SARS-CoV-2 spike protein to the host angiotensin-converting enzyme 2 (ACE2) receptor is required for cellular entry of SARS-CoV-2. Toward identifying new chemical leads that can disrupt this interaction, including in the presence of SARS-CoV-2 adaptive mutations found in variants like omicron that can circumvent vaccine, immune, and therapeutic antibody responses, we synthesized 5-chloro-3-(2-(2,4-dinitrophenyl)hydrazono)indolin-2-one (H 2 L) from the condensation reaction of 5-chloroisatin and 2,4-dinitrophenylhydrazine in good yield. H 2 L was characterised by elemental and spectral (IR, electronic, Mass) analyses. The NMR spectrum of H 2 L indicated a keto-enol tautomerism, with the keto form being more abundant in solution. H 2 L was found to selectively interfere with binding of the SARS-CoV-2 spike receptor-binding domain (RBD) to the host angiotensin-converting enzyme 2 receptor with a 50% inhibitory concentration (IC 50 ) of 0.26 μM, compared to an unrelated PD-1/PD-L1 ligand-receptor-binding pair with an IC 50 of 2.06 μM in vitro (Selectivity index = 7.9). Molecular docking studies revealed that the synthesized ligand preferentially binds within the ACE2 receptor-binding site in a region distinct from where spike mutations in SARS-CoV-2 variants occur. Consistent with these models, H 2 L was able to disrupt ACE2 interactions with the RBDs from beta, delta, lambda, and omicron variants with similar activities. These studies indicate that H 2 L-derived compounds are potential inhibitors of multiple SARS-CoV-2 variants, including those capable of circumventing vaccine and immune responses., Supplementary Information: The online version contains supplementary material available at 10.1007/s11696-023-03274-5., Competing Interests: Conflict of interestWe declare none., (© The Author(s) 2024.)

Background: A feature of HIV cure trials is the need to interrupt treatment to test the efficacy of experimental interventions-a process known as analytical treatment interruptions (ATIs)., Objectives: We report the experiences of participants after they completed an extended ATI., Methods: From April to November 2022, we conducted post-ATI in-depth interviews with BEAT2 clinical trial (NCT03588715) participants who stopped ART while receiving an immunotherapy regimen. We used conventional content analysis to code the data., Results: We conducted interviews with 11 Black/African American and three White/Caucasian participants (11 males, two females, and one transgender woman). The mean ATI was 38 weeks. Participants noted several significant experiences surrounding the interventions' side effects, ATI, and returning to medication. Some participants had positive experiences with their ATI. Other participants were nervous during the ATI. Rising viral loads led some to feel a sense of failure. Although trial experiences were heterogeneous, participants unanimously had positive interactions with the clinical trial staff which facilitated their retention in the trial. Participants shared their experiences with the trial, including changes in expectations, experiences with experimental interventions and procedures, compensation as a measure of respect, effort, transportation, and effects of COVID-19 during the trial. Based on these results, we provide considerations for the conduct of future HIV cure-directed clinical trials involving ATIs., Conclusions: Managing expectations, focusing on participants' contributions, and providing support to reduce feelings of having failed the research team and/or the HIV community following viral rebound should be part of HIV cure trial design. Discussing the mental health impact of rebound during consent, distinct from risk, is needed. Continued efforts to understand how people with HIV experience ATIs will improve future designs of HIV cure clinical trials.

HIV-1 persists in a latent reservoir in resting CD4+ T cells despite antiretroviral therapy (ART). The reservoir decays slowly over the first 7 years of ART (t1/2 = 44 months). However, whether decay continues with long-term ART is unclear. Recent integration site studies indicate gradual selection against inducible, intact proviruses, raising speculation that decades of ART might allow treatment interruption without viral rebound. Therefore, we measured the reservoir in 42 people on long-term ART (mean 22 years) using a quantitative viral outgrowth assay. After 7 years of ART, there was no long-term decrease in the frequency of inducible, replication-competent proviruses but rather an increase with an estimated doubling time of 23 years. Another reservoir assay, the intact proviral DNA assay, confirmed that reservoir decay with t1/2 of 44 months did not continue with long-term ART. The lack of decay reflected proliferation of infected cells. Most inducible, replication-competent viruses (79.8%) had env sequences identical to those of other isolates from the same sample. Thus, although integration site analysis indicates changes in reservoir composition, the proliferation of CD4+ T cells counteracts decay, maintaining the frequency of inducible, replication-competent proviruses at roughly constant levels over the long term. These results reinforce the need for lifelong ART.

Analytical treatment interruption (ATI), defined as a closely monitored clinical pause in antiretroviral therapy (ART), is a core component of many HIV cure-directed clinical studies. ATIs may cause significant physical and psychosocial risks for people living with HIV and, as a result, integrating participant and community perspectives into clinical trial designs that include an ATI is crucial to ensuring a successful and person-centered trial. We conducted semi-structured interviews with participants enrolling in the BEAT-2 cure-directed trial (NCT03588715). Interviews elicited participant motivations and decision-making processes for trial participation along with participants' perceptions of the ATI. Interviews were recorded, transcribed, and analyzed using a directed content analysis. Fourteen of 15 trial participants completed interviews. The majority were Black (79%) cisgender male (79%). Participants noted several significant motivating factors contributing to their desire to enroll in the HIV cure-directed clinical trial, the most prominent being a desire to find a cure for HIV and help others in the HIV community. HIV care teams were the most commonly identified resource for patients when making the decision to enroll in the trial, and family, friends, and romantic partners also played a significant role. Altruism was a primary motivation for participation, although participants also shared interest in learning about HIV science and research. Participants had a strong understanding of trial procedures and displayed significant trust in the study team to keep them informed and healthy during their participation. The ATI was a significant source of anxiety for participants. Their primary worry was that their prior antiretroviral therapy (ART) regimen would no longer be effective once they resumed ART. Despite these concerns, participants shared considerable excitement for continued participation in the trial and being a part of the search toward an HIV cure.

Objective: The aim of this study was to assess the susceptibility of HIV to two HIV monoclonal antibodies (bnAbs), 3BNC117 and 10-1074, in individuals with chronically antiretroviral therapy (ART) suppressed HIV infection., Design: The susceptibility of bnAbs was determined using the PhenoSense mAb Assay, which is a cell-based infectivity assay designed to assess the susceptibility of luciferase-reporter pseudovirions. This assay is the only Clinical Laboratory Improvement Ammendment (CLIA)/College of American Pathologist (CAP) compliant screening test specifically developed for evaluating bnAb susceptibility in people with HIV infection., Method: The susceptibility of luciferase-reporter pseudovirions, derived from HIV-1 envelope proteins obtained from peripheral bloodmononuclear cells of 61 ART-suppressed individuals, to 3BNC117 and 10-1074 bnAbs was assessed using the PhenoSense mAb assay. Susceptibility was defined as an IC 90 of <2.0 μg/ml and 1.5 μg/ml for 3BNC117 and 10-1074, respectively., Results: About half of the individuals who were chronically infected and virologically suppressed were found to harbor virus with reduced susceptibility to one or both of the tested bnAbs., Conclusions: The reduced combined susceptibility of 3BNC117 and 10-1074 highlights a potential limitation of using only two bnAbs for pre-exposure prophylaxis or treatment. Further studies are needed to define and validate the clinical correlates of bnAb susceptibility., (Copyright © 2023 Wolters Kluwer Health, Inc. All rights reserved.)

Despite effective combination antiretroviral therapy (cART), people living with HIV (PLWH) continue to harbor replication-competent and transcriptionally active virus in infected cells, which in turn can lead to ongoing viral antigen production, chronic inflammation, and increased risk of age-related comorbidities. To identify new agents that may inhibit postintegration HIV beyond cART, we screened a library of 512 pure compounds derived from natural products and identified (-)-hopeaphenol as an inhibitor of HIV postintegration transcription at low to submicromolar concentrations without cytotoxicity. Using a combination of global RNA sequencing, plasmid-based reporter assays, and enzyme activity studies, we document that hopeaphenol inhibits protein kinase C (PKC)- and downstream NF-κB-dependent HIV transcription as well as a subset of PKC-dependent T-cell activation markers, including interleukin-2 (IL-2) cytokine and CD25 and HLA-DRB1 RNA production. In contrast, it does not substantially inhibit the early PKC-mediated T-cell activation marker CD69 production of IL-6 or NF-κB signaling induced by tumor necrosis factor alpha (TNF-α). We further show that hopeaphenol can inhibit cyclin-dependent kinase 9 (CDK9) enzymatic activity required for HIV transcription. Finally, it inhibits HIV replication in peripheral blood mononuclear cells (PBMCs) infected in vitro and dampens viral reactivation in CD4 + cells from PLWH. Our study identifies hopeaphenol as a novel inhibitor that targets a subset of PKC-mediated T-cell activation pathways in addition to CDK9 to block HIV expression. Hopeaphenol-based therapies could complement current antiretroviral therapy otherwise not targeting cell-associated HIV RNA and residual antigen production in PLWH.

ARID1A, encoding a subunit of the SWI/SNF complex, is mutated in ∼50% of clear cell ovarian carcinoma (OCCC) cases. Here we show that inhibition of the mevalonate pathway synergizes with immune checkpoint blockade (ICB) by driving inflammasome-regulated immunomodulating pyroptosis in ARID1A-inactivated OCCCs. SWI/SNF inactivation downregulates the rate-limiting enzymes in the mevalonate pathway such as HMGCR and HMGCS1, which creates a dependence on the residual activity of the pathway in ARID1A-inactivated cells. Inhibitors of the mevalonate pathway such as simvastatin suppresses the growth of ARID1A mutant, but not wild-type, OCCCs. In addition, simvastatin synergizes with anti-PD-L1 antibody in a genetic OCCC mouse model driven by conditional Arid1a inactivation and in a humanized immunocompetent ARID1A mutant patient-derived OCCC mouse model. Our data indicate that inhibition of the mevalonate pathway simultaneously suppresses tumor cell growth and boosts antitumor immunity by promoting pyroptosis, which synergizes with ICB in suppressing ARID1A-mutated cancers., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Thorectidiols isolated from the marine sponge Dactylospongia elegans (family Thorectidae, order Dictyoceratida) collected in Papua New Guinea are a family of symmetrical and unsymmetrical dimeric biphenyl meroterpenoid stereoisomers presumed to be products of oxidative phenol coupling of a co-occurring racemic monomer, thorectidol ( 3 ). One member of the family, thorectidiol A ( 1 ), has been isolated in its natural form, and its structure has been elucidated by analysis of NMR, MS, and ECD data. Acetylation of the sponge extract facilitated isolation of additional thorectidiol diacetate stereoisomers and the isolation of the racemic monomer thorectidol acetate ( 6 ). Racemic thorectidiol A ( 1 ) showed selective inhibition of the SARS-CoV-2 spike receptor binding domain (RBD) interaction with the host ACE2 receptor with an IC 50 = 1.0 ± 0.7 μM.

The known Eremophila microtheca -derived diterpenoid 3,7,8-trihydroxyserrulat-14-en-19-oic acid ( 1 ) was targeted for large-scale purification, as this bioactive plant compound has proven to be an attractive scaffold for semisynthetic studies and subsequent library generation. Compound 1 was converted to a selectively protected trimethyl derivative, 3-hydroxy-7,8-dimethoxyserrulat-14-en-19-oic acid methyl ester ( 2 ), using simple and rapid methylation conditions. The resulting scaffold 2 was reacted with a diverse series of commercially available isocyanates to generate an 11-membered carbamate-based library. The chemical structures of the 11 new semisynthetic analogues were fully characterized by spectroscopic and spectrometric analysis. All natural products and semisynthetic compounds were evaluated for their anthelmintic, antimalarial, and anti-HIV activities. Compound 3 was shown to elicit the greatest antiplasmodial activity of all compounds tested, with IC 50 values of 4.6 and 11.6 μM against Plasmodium falciparum 3D7 and Dd2, respectively. Compound 11 showed the greatest inhibition of development to fourth-stage Haemonchus contortus larvae (L4) and induction of a skinny ( Ski ) phenotype (67.5% of nematodes) at 50 μM. Compound 7 , which inhibited 59.0% of HIV production at 100 μg/mL, was the carbamate analogue that displayed the best antiviral activity.

Objective: The human endogenous protein galectin-9 (Gal-9) reactivates latently HIV-infected cells in vitro and ex vivo , which may allow for immune-mediated clearance of these cells. However, Gal-9 also activates several immune cells, which could negatively affect HIV persistence by promoting chronic activation/exhaustion. This potential 'double-edged sword' effect of Gal-9 raises the question of the overall impact of Gal-9 on HIV persistence in vivo ., Design: We used the BLT (bone marrow, liver, thymus) humanized mouse model to evaluate the impact of Gal-9 on HIV persistence in vivo during antiretroviral therapy (ART)., Methods: Two independent cohorts of ART-suppressed HIV-infected BLT mice were treated with either recombinant Gal-9 or phosphate-buffered saline control. Plasma viral loads and levels of tissue-associated HIV DNA and RNA were measured by qPCR. Immunohistochemistry and HIV RNAscope were used to quantify CD4 + T, myeloid, and HIV RNA+ cells in tissues. T cell activation and exhaustion were measured by flow cytometry, and plasma markers of inflammation were measured by multiplex cytokine arrays., Results: Gal-9 did not induce plasma markers of inflammation or T cell markers of activation/exhaustion in vivo . However, the treatment significantly increased levels of tissue-associated HIV DNA and RNA compared to controls ( P  = 0.0007 and P  = 0.011, respectively, for cohort I and P  = 0.002 and P  = 0.005, respectively, for cohort II). RNAscope validated the Gal-9 mediated induction of HIV RNA in tissue-associated myeloid cells, but not T cells., Conclusions: Our study highlights the overall adverse effects of Gal-9 on HIV persistence and the potential need to block Gal-9 interactions during ART-suppressed HIV infection., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

A significant number of persons with coronavirus disease 2019 (COVID-19) experience persistent, recurrent, or new symptoms several months after the acute stage of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. This phenomenon, termed post-acute sequelae of SARS-CoV-2 (PASC) or long COVID, is associated with high viral titers during acute infection, a persistently hyperactivated immune system, tissue injury by NETosis-induced micro-thrombofibrosis (NETinjury), microbial translocation, complement deposition, fibrotic macrophages, the presence of autoantibodies, and lymphopenic immune environments. Here, we review the current literature on the immunological imbalances that occur during PASC. Specifically, we focus on data supporting common immunopathogenesis and tissue injury mechanisms shared across this highly heterogenous disorder, including NETosis, coagulopathy, and fibrosis. Mechanisms include changes in leukocyte subsets/functions, fibroblast activation, cytokine imbalances, lower cortisol, autoantibodies, co-pathogen reactivation, and residual immune activation driven by persistent viral antigens and/or microbial translocation. Taken together, we develop the premise that SARS-CoV-2 infection results in PASC as a consequence of acute and/or persistent single or multiple organ injury mediated by PASC determinants to include the degree of host responses (inflammation, NETinjury), residual viral antigen (persistent antigen), and exogenous factors (microbial translocation). Determinants of PASC may be amplified by comorbidities, age, and sex., Competing Interests: Conflict of interest: The authors declare no conflicts of interest., (© The Author(s) 2023. Published by Oxford University Press on behalf of Society for Leukocyte Biology.)

Understanding the complexity of the long-lived HIV reservoir during antiretroviral therapy (ART) remains a considerable impediment in research towards a cure for HIV. To address this, we developed a single-cell strategy to precisely define the unperturbed peripheral blood HIV-infected memory CD4 + T cell reservoir from ART-treated people living with HIV (ART-PLWH) via the presence of integrated accessible proviral DNA in concert with epigenetic and cell surface protein profiling. We identified profound reservoir heterogeneity within and between ART-PLWH, characterized by new and known surface markers within total and individual memory CD4 + T cell subsets. We further uncovered new epigenetic profiles and transcription factor motifs enriched in HIV-infected cells that suggest infected cells with accessible provirus, irrespective of reservoir distribution, are poised for reactivation during ART treatment. Together, our findings reveal the extensive inter- and intrapersonal cellular heterogeneity of the HIV reservoir, and establish an initial multiomic atlas to develop targeted reservoir elimination strategies., (© 2022. The Author(s).)

Background: People with HIV (PWH) and community members have advocated for the development of a home-based viral load test device that could make analytical treatment interruptions (ATIs) less burdensome., Objective: We assessed community acceptability of a novel home-based viral load test device., Methods: In 2021, we conducted 15 interviews and 3 virtual focus groups with PWH involved in HIV cure research. We used conventional thematic analysis to analyze the data., Results: PWH viewed the home-based viral load test device as a critical adjunct in ongoing HIV cure trials with ATIs. The ability to test for viral load at home on demand would alleviate anxiety around being off ART. Participants drew parallels with glucometers used for diabetes. A preference was expressed for the home-based test to clearly indicate whether one was detectable or undetectable for HIV to mitigate risk of HIV transmission to partners. Perceived advantages of the device included convenience, sense of control, and no puncturing of veins. Perceived concerns were possible physical marks, user errors and navigating the logistics of mailing samples to a laboratory and receiving test results. Participants expressed mixed effects on stigma, such as helping normalize HIV, but increased potential for inadvertent disclosure of HIV status or ATI participation. Increasing pluri-potency of the device beyond viral load testing (e.g., CD4+ count test) would increase its utility. Participants suggested pairing the device with telemedicine and mobile health technologies., Conclusions: If proven effective, the home-based viral load test device will become a critical adjunct in HIV cure research and HIV care.

Cancer immunotherapy often depends on recognition of peptide epitopes by cytotoxic T lymphocytes (CTLs). The tumor microenvironment (TME) is enriched for peroxynitrite (PNT), a potent oxidant produced by infiltrating myeloid cells and some tumor cells. We demonstrate that PNT alters the profile of MHC class I bound peptides presented on tumor cells. Only CTLs specific for PNT-resistant peptides have a strong antitumor effect in vivo, whereas CTLs specific for PNT-sensitive peptides are not effective. Therapeutic targeting of PNT in mice reduces resistance of tumor cells to CTLs. Melanoma patients with low PNT activity in their tumors demonstrate a better clinical response to immunotherapy than patients with high PNT activity. Our data suggest that intratumoral PNT activity should be considered for the design of neoantigen-based therapy and also may be an important immunotherapeutic target., Competing Interests: Declaration of interests E.C. is a paid consultant for Oncovir, Inc. E.M., Q.Z., A.M., and D.I.G. are employees and shareholders of AstraZeneca., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Opioid use has negative effects on immune responses and may impair immune reconstitution in persons living with HIV (PLWH) infection undergoing antiretroviral treatment (ART). The effects of treatment with μ opioid receptor (MOR) agonists (e.g., methadone, MET) and antagonists (e.g., naltrexone, NTX) on immune reconstitution and immune activation in ART-suppressed PLWH have not been assessed in-depth. We studied the effects of methadone or naltrexone on measures of immune reconstitution and immune activation in a cross-sectional community cohort of 30 HIV-infected individuals receiving suppressive ART and medications for opioid use disorder (MOUD) (12 MET, 8 NTX and 10 controls). Plasma markers of inflammation and immune activation were measured using ELISA, Luminex, or Simoa. Plasma IgG glycosylation was assessed using capillary electrophoresis. Cell subsets and activation were studied using whole blood flow cytometry. Individuals in the MET group, but no in the NTX group, had higher plasma levels of inflammation and immune activation markers than controls. These markers include soluble CD14 (an independent predictor of morbidity and mortality during HIV infection), proinflammatory cytokines, and proinflammatory IgG glycans. This effect was independent of time on treatment. Our results indicate that methadone-based MOUD regimens may sustain immune activation and inflammation in ART-treated HIV-infected individuals. Our pilot study provides the foundation and rationale for future longitudinal functional studies of the impact of MOUD regimens on immune reconstitution and residual activation after ART-mediated suppression., (©2022 Society for Leukocyte Biology.)

Background : HIV cure-directed clinical trials using analytical treatment interruptions (ATIs) require participants to adhere to frequent monitoring visits for viral load tests. Novel viral load monitoring strategies are needed to decrease participant burden during ATIs. Objective : To examine acceptability of a novel home-based blood collection device for viral load testing in the context of two ongoing ATI trials in Philadelphia, PA, United States. Methods : From January 2021 to February 2022, participants completed three in-depth interviews via teleconference during their participation in an ATI: (1) within two weeks of enrollment in the device study, (2) approximately four weeks after beginning to use the device, and (3) within two weeks of the end of the ATI when ART was re-initiated. We used conventional content analysis to analyze the data. Results : We recruited 17 participants: 15 were cisgender males, 1 cisgender female, and 1 transgender woman. We observed an overall 87% success rate in drawing blood with the device from home collection and found overall high acceptance of the device. A mean of 91.5 devices per participant were used for home-based blood collection. Most PWH viewed the device as relatively convenient, painless, easy to use, and a simple solution to frequent blood draws. The main challenge encountered was the inability to completely fill up devices with blood in some cases. Most participants reported positive experiences with mailing blood samples and could see themselves using the device on a regular basis outside of ATIs. Conclusions : Our study showed participant valued the novel home-based peripheral blood collection for viral load testing in the context of ATI trials. More research will be necessary to optimize implementation of the device and to assess whether blood collected can reliably measure viral loads in the context of ATI trials.

Despite effective antiretroviral therapy, HIV-1 persistence in latent reservoirs remains a major obstacle to a cure. We postulate that HIV-1 silencing factors suppress HIV-1 reactivation and that inhibition of these factors will increase HIV-1 reactivation. To identify HIV-1 silencing factors, we conducted a genome-wide CRISPR inhibition (CRISPRi) screen using four CRISPRi-ready, HIV-1-d6-GFP-infected Jurkat T cell clones with distinct integration sites. We sorted cells with increased green fluorescent protein (GFP) expression and captured single guide RNAs (sgRNAs) via targeted deep sequencing. We identified 18 HIV-1 silencing factors that were significantly enriched in HIV-1-d6-GFP high cells. Among them, SLTM (scaffold attachment factor B-like transcription modulator) is an epigenetic and transcriptional modulator having both DNA and RNA binding capacities not previously known to affect HIV-1 transcription. Knocking down SLTM by CRISPRi significantly increased HIV-1-d6-GFP expression (by 1.9- to 4.2-fold) in three HIV-1-d6-GFP-Jurkat T cell clones. Furthermore, SLTM knockdown increased the chromatin accessibility of HIV-1 and the gene in which HIV-1 is integrated but not the housekeeping gene POLR2A . To test whether SLTM inhibition can reactivate HIV-1 and further induce cell death of HIV-1-infected cells ex vivo , we established a small interfering RNA (siRNA) knockdown method that reduced SLTM expression in CD4 + T cells from 10 antiretroviral therapy (ART)-treated, virally suppressed, HIV-1-infected individuals ex vivo . Using limiting dilution culture, we found that SLTM knockdown significantly reduced the frequency of HIV-1-infected cells harboring inducible HIV-1 by 62.2% (0.56/10 6 versus 1.48/10 6 CD4 + T cells [ P  = 0.029]). Overall, our study indicates that SLTM inhibition reactivates HIV-1 in vitro and induces cell death of HIV-1-infected cells ex vivo . Our study identified SLTM as a novel therapeutic target. IMPORTANCE HIV-1-infected cells, which can survive drug treatment and immune cell killing, prevent an HIV-1 cure. Immune recognition of infected cells requires HIV-1 protein expression; however, HIV-1 protein expression is limited in infected cells after long-term therapy. The ways in which the HIV-1 provirus is blocked from producing protein are unknown. We identified a new host protein that regulates HIV-1 gene expression. We also provided a new method of studying HIV-1-host factor interactions in cells from infected individuals. These improvements may enable future strategies to reactivate HIV-1 in infected individuals so that infected cells can be killed by immune cells, drug treatment, or the virus itself.

Despite the success of combination antiretroviral therapy (cART), HIV persists in low- and middle-income countries (LMIC) due to emerging drug resistance and insufficient drug accessibility. Furthermore, cART does not target latently-infected CD4+ T cells, which represent a major barrier to HIV eradication. The “shock and kill” therapeutic approach aims to reactivate provirus expression in latently-infected cells in the presence of cART and target virus-expressing cells for elimination. An attractive therapeutic prototype in LMICs would therefore be capable of simultaneously inhibiting viral replication and inducing latency reversal. Here we report that Gnidia sericocephala, which is used by traditional health practitioners in South Africa for HIV/AIDS management to supplement cART, contains at least four daphnane-type compounds (yuanhuacine A (1), yuanhuacine as part of a mixture (2), yuanhuajine (3), and gniditrin (4)) that inhibit viral replication and/or reverse HIV latency. For example, 1 and 2 inhibit HIV replication in peripheral blood mononuclear cells (PBMC) by >80% at 0.08 µg/mL, while 1 further inhibits a subtype C virus in PBMC with a half-maximal effective concentration (EC50) of 0.03 µM without cytotoxicity. Both 1 and 2 also reverse HIV latency in vitro consistent with protein kinase C activation but at 16.7-fold lower concentrations than the control prostratin. Both 1 and 2 also reverse latency in primary CD4+ T cells from cART-suppressed donors with HIV similar to prostratin but at 6.7-fold lower concentrations. These results highlight G. sericocephala and components 1 and 2 as anti-HIV agents for improving cART efficacy and supporting HIV cure efforts in resource-limited regions.

People with HIV (PWH) are at increased risk of developing active tuberculosis (TB), and anemia is a common complication in both conditions. Anemia in TB patients has been linked to immune activation, levels of inflammatory biomarkers in blood, and risk for HIV disease progression and death. In this study we show that anemia was associated with a more pronounced inflammatory profile in HIV-TB coinfected persons in a cohort of 159 individuals with advanced HIV disease (CD4 count < 100 cells/µL) recruited as part of a randomized clinical trial (NCT00988780). A panel of plasma biomarkers was assessed on plasma obtained prior to combination antiretroviral therapy (cART) initiation. We performed a series of multidimensional analyses including clinical variables and concentrations of inflammatory biomarkers to profile systemic inflammation of PWH with and without anemia. We observed that TB participants presented with moderately lower levels of hemoglobin than non-TB participants. These participants also presented a higher Degree of Inflammatory Perturbation (DIP) score, related to increased levels of IFN-γ and TNF. The DIP was associated with TB coinfection and anemia before cART initiation. Future mechanistic studies are warranted to assess the determinants of such associations and the implications on treatment outcomes., Competing Interests: JS-M reports grants from BMS, Pfizer, MSD, and Stendhal outside the submitted work. LJM reports grants from Merck and Gene-One. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Araújo-Pereira, Barreto-Duarte, Arriaga, Musselwhite, Vinhaes, Belaunzaran-Zamudio, Rupert, Montaner, Lederman, Sereti, Madero and Andrade.)

Understanding the drivers and markers of clonally expanding HIV-1-infected CD4 + T cells is essential for HIV-1 eradication. We used single-cell ECCITE-seq, which captures surface protein expression, cellular transcriptome, HIV-1 RNA, and TCR sequences within the same single cell to track clonal expansion dynamics in longitudinally archived samples from six HIV-1-infected individuals (during viremia and after suppressive antiretroviral therapy) and two uninfected individuals, in unstimulated conditions and after CMV and HIV-1 antigen stimulation. Despite antiretroviral therapy, persistent antigen and TNF responses shaped T cell clonal expansion. HIV-1 resided in Th1-polarized, antigen-responding T cells expressing BCL2 and SERPINB9 that may resist cell death. HIV-1 RNA + T cell clones were larger in clone size, established during viremia, persistent after viral suppression, and enriched in GZMB + cytotoxic effector memory Th1 cells. Targeting HIV-1-infected cytotoxic CD4 + T cells and drivers of clonal expansion provides another direction for HIV-1 eradication., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Five new minor sesterterpenoids, ansellones H ( 4 ), I ( 5 ), J ( 6 ), and K ( 7 ) and phorone C ( 8 ), have been isolated from a Phorbas sp. marine sponge collected in British Columbia. Their structures have been elucidated by detailed analysis of NMR and MS data. Ansellone J ( 6 ) and phorone C ( 8 ) are potent in vitro HIV-1 latency reversal agents that are more potent than the reference compound and control protein kinase C activator prostratin ( 3 ). The most potent Phorbas sesterterpenoid, ansellone J ( 6 ), was evaluated for HIV latency reversal in a primary cell context using CD4+ T cells obtained directly from four combination antiretroviral therapy-suppressed donors with HIV. To a first approximation, ansellone J ( 6 ) induced HIV latency reversal at levels similar to prostratin ( 3 ) ex vivo , but at a 10-fold lower concentration.

SARS-CoV-2, the causative agent of COVID-19, continues to cause global morbidity and mortality despite the increasing availability of vaccines. Alongside vaccines, antivirals are urgently needed to combat SARS-CoV-2 infection and spread, particularly in resource-limited regions which lack access to existing therapeutics. Small molecules isolated from medicinal plants may be able to block cellular entry by SARS-CoV-2 by antagonising the interaction of the viral spike glycoprotein receptor-binding domain (RBD) with the host angiotensin-converting enzyme II (ACE2) receptor. As the medicinal plant Gunnera perpensa L. is being used by some South African traditional healers for SARS-CoV-2/COVID-19 management, we hypothesised that it may contain chemical constituents that inhibit the RBD-ACE2 interaction. Using a previously described AlphaScreen-based protein interaction assay, we show here that the DCM:MeOH extract of G. perpensa readily disrupts RBD (USA-WA1/2020)-ACE2 interactions with a half-maximal inhibition concentration (IC 50 ) of < 0.001 µg/mL, compared to an IC 50 of 0.025 µg/mL for the control neutralising antibody REGN10987. Employing hyphenated analytical techniques like UPLC-IMS-HRMS (method developed and validated as per the International Conference on Harmonization guidelines), we identified two ellagitannins, punicalin (2.12% w/w) and punicalagin (1.51% w/w), as plant constituents in the DCM:MeOH extract of G. perpensa which antagonised RBD-ACE2 binding with respective IC 50 s of 9 and 29 nM. This good potency makes both compounds promising leads for development of future entry-based SARS-CoV-2 antivirals. The results also highlight the advantages of combining reverse pharmacology (based on medicinal plant use) with hyphenated analytical techniques to expedite identification of urgently needed antivirals., (© 2022. Springer-Verlag GmbH Germany, part of Springer Nature.)