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Introduction: Restriction factors (RFs) suppress HIV-1 in cell lines and primary cell models. Hence, RFs might be attractive targets for novel antiviral strategies, but their importance for virus control in vivo is controversial., Methods: We profiled the expression of RFs in primary blood-derived mononuclear cells (PBMC) from therapy-naïve HIV-1 patients and quantified infection., Results: Overall, there was no correlation between individual RF expression and HIV-1 status in total PBMC. However, we identified a T cell population with low levels of intracellular CD2 and reduced expression of SAMHD1, p21 and SerinC5. CD2 low T cells with reduced RF expression were markedly positive for HIV-1 p24. In contrast, CD2+ T cells were less infected and expressed higher levels of RFs. CD2 low T cell infection correlated with viral loads and was associated with HIV-1 disease progression., Conclusions: In untreated therapy naïve chronic HIV-1 patients, RF expression in T cells is associated with CD2 expression and seems to influence viral loads. Our study suggests that RFs help to control HIV-1 infection in certain T cells in vivo and supports the potential for RFs as promising targets for therapeutic intervention., Competing Interests: The authors declare that they have no competing interests.

While an essential role of HIV-1 integrase (IN) for integration of viral cDNA into human chromosome is established, studies with IN mutants and allosteric IN inhibitors (ALLINIs) have suggested that IN can also influence viral particle maturation. However, it has remained enigmatic as to how IN contributes to virion morphogenesis. Here, we demonstrate that IN directly binds the viral RNA genome in virions. These interactions have specificity, as IN exhibits distinct preference for select viral RNA structural elements. We show that IN substitutions that selectively impair its binding to viral RNA result in eccentric, non-infectious virions without affecting nucleocapsid-RNA interactions. Likewise, ALLINIs impair IN binding to viral RNA in virions of wild-type, but not escape mutant, virus. These results reveal an unexpected biological role of IN binding to the viral RNA genome during virion morphogenesis and elucidate the mode of action of ALLINIs., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Novel therapeutic options are urgently needed to improve global treatment of virus infections. Herbal products with confirmed clinical safety features are attractive starting material for the identification of new antiviral activities. Here we demonstrate that Cistus incanus (Ci) herbal products inhibit human immunodeficiency virus (HIV) infections in vitro. Ci extract inhibited clinical HIV-1 and HIV-2 isolates, and, importantly, a virus isolate with multiple drug resistances, confirming broad anti-HIV activity. Antiviral activity was highly selective for virus particles, preventing primary attachment of the virus to the cell surface and viral envelope proteins from binding to heparin. Bioassay-guided fractionation indicated that Ci extract contains numerous antiviral compounds and therefore has favorably low propensity to induce virus resistance. Indeed, no resistant viruses emerged during 24 weeks of continuous propagation of the virus in the presence of Ci extracts. Finally, Ci extracts also inhibited infection by virus particles pseudotyped with Ebola and Marburg virus envelope proteins, indicating that antiviral activity of Ci extract extends to emerging viral pathogens. These results demonstrate that Ci extracts show potent and broad in vitro antiviral activity against viruses that cause life-threatening diseases in humans and are promising sources of agents that target virus particles.

Global HIV-1 treatment would benefit greatly from safe herbal medicines with scientifically validated novel anti-HIV-1 activities. The root extract from the medicinal plant Pelargonium sidoides (PS) is licensed in Germany as the herbal medicine EPs®7630, with numerous clinical trials supporting its safety in humans. Here we provide evidence from multiple cell culture experiments that PS extract displays potent anti-HIV-1 activity. We show that PS extract protects peripheral blood mononuclear cells and macrophages from infection with various X4 and R5 tropic HIV-1 strains, including clinical isolates. Functional studies revealed that the extract from PS has a novel mode-of-action. It interferes directly with viral infectivity and blocks the attachment of HIV-1 particles to target cells, protecting them from virus entry. Analysis of the chemical footprint of anti-HIV activity indicates that HIV-1 inhibition is mediated by multiple polyphenolic compounds with low cytotoxicity and can be separated from other extract components with higher cytotoxicity. Based on our data and its excellent safety profile, we propose that PS extract represents a lead candidate for the development of a scientifically validated herbal medicine for anti-HIV-1 therapy with a mode-of-action different from and complementary to current single-molecule drugs.

There is clinical evidence showing that the Pelargonium sidoides root extract EPs 7630 is a safe and effective treatment for a range of acute infectious respiratory illnesses. Moreover, EPs 7630 has been shown to reduce the use of antibiotics, which is important in the context of rising antibiotic resistance levels. A wide range of mechanisms appears to contribute to the beneficial effects of EPs 7630, e.g. antibacterial, antiviral, immunomodulatory, and epithelial barrier effects. This broad spectrum of pharmacological activities seems to enable the clinical activity of EPs 7630 against multiple respiratory infections. In particular, the combination of antiviral and immunomodulatory effects may enable EPs 7630 to tackle acute viral respiratory infections both in early stages of the disease process, which are driven by virus replication, as well as in later stages, which are caused by an overshooting immune response. Hence, EPs 7630 is a prime example of a plant extract with evidence-based clinical efficacy, including a solid understanding of the underlying mechanisms of action. The example of EPs 7630 demonstrates that plant extracts have a potential role as evidence-based clinical treatments and that they deserve pre-clinical and clinical testing and investigation in the same way as any other drug class. [ABSTRACT FROM AUTHOR]

HIV-1 virion maturation is an essential step in the viral replication cycle to produce infectious virus particles. Gag and Gag-Pol polyproteins are assembled at the plasma membrane of the virus-producer cells and bud from it to the extracellular compartment. The newly released progeny virions are initially immature and noninfectious. However, once the Gag polyprotein is cleaved by the viral protease in progeny virions, the mature capsid proteins assemble to form the fullerene core. This core, harboring two copies of viral genomic RNA, transforms the virion morphology into infectious virus particles. This morphological transformation is referred to as maturation. Virion maturation influences the distribution of the Env glycoprotein on the virion surface and induces conformational changes necessary for the subsequent interaction with the CD4 receptor. Several host factors, including proteins like cyclophilin A, metabolites such as IP6, and lipid rafts containing sphingomyelins, have been demonstrated to have an influence on virion maturation. This review article delves into the processes of virus maturation and Env glycoprotein recruitment, with an emphasis on the role of host cell factors and environmental conditions. Additionally, we discuss microscopic technologies for assessing virion maturation and the development of current antivirals specifically targeting this critical step in viral replication, offering long-acting therapeutic options. [ABSTRACT FROM AUTHOR]

Ultrasound is an indispensable tool for preliminary diagnosis ("filter function"), during treatment ("monitoring function") and in the follow-up examinations ("follow-up function") after shock-wave lithotripsy of gallstones. It permits rapid and reliable assessment of the therapeutic outcome and early identification of complications, which present-day experience has shown to be rare.

Using a subtile examination technique genuine information can be obtained from the interior of a gallstone--at least to a depth of 15 mm. This information should provide an indication of the homogeneous (crystalline) or inhomogeneous (non-cholesterol) structure of the stone. Further systematic use of differentiating gallstone ultrasonography could become a useful additional decision making criterion for selection of gallstones suitable for conservative treatment.

Cyclophilins (Cyps), characterized as peptidyl-prolyl cis-trans isomerases (PPIases), are highly conserved and ubiquitous, playing a crucial role in protein folding and cellular signaling. This review summarizes the biochemical pathways mediated by Cyps, including their involvement in pathological states such as viral replication, inflammation, and cancer progression, to underscore the therapeutic potential of Cyp inhibition. The exploration of Cyp inhibitors (CypI) in this review, particularly non-immunosuppressive cyclosporine A (CsA) derivatives, highlights their significance as therapeutic agents. The structural and functional nuances of CsA derivatives are examined, including their efficacy, mechanism of action, and the balance between therapeutic benefits and off-target effects. The landscape of CypI is evaluated to emphasize the clinical need for targeted approaches to exploit the complex biology of Cyps and to propose future directions for research that may enhance the utility of non-immunosuppressive CsA derivatives in treating diseases where Cyps play a key pathological role. [ABSTRACT FROM AUTHOR]

Retroviral integration is mediated by intasome nucleoprotein complexes wherein a pair of viral DNA ends are bridged together by a multimer of integrase (IN). Atomic-resolution structures of HIV-1 intasomes provide detailed insights into the mechanism of integration and inhibition by clinical IN inhibitors. However, previously described HIV-1 intasomes are highly heterogeneous and have the tendency to form stacks, which is a limiting factor in determining high-resolution cryo-EM maps. We have assembled HIV-1 intasomes in the presence of excess IN C-terminal domain protein, which was readily incorporated into the intasomes. The purified intasomes were largely homogeneous and exhibited minimal stacking tendencies. The cryo-EM map resolution was further improved to 2.01 Å, which will greatly facilitate structural studies of IN inhibitor action and drug resistance mechanisms. The C-terminal 18 residues of HIV-1 IN, which are critical for virus replication and integration in vitro, have not been well resolved in previous intasome structures, and its function remains unclear. We show that the C-terminal tail participates in intasome assembly, resides within the intasome core, and forms a small alpha helix (residues 271–276). Mutations that disrupt alpha helix integrity impede IN activity in vitro and disrupt HIV-1 infection at the step of viral DNA integration. [ABSTRACT FROM AUTHOR]

The current study analyzed the intersecting biophysical, biochemical, and functional properties of extracellular particles (EPs) with the human immunodeficiency virus type‐1 (HIV‐1) beyond the currently accepted size range for HIV‐1. We isolated five fractions (Frac‐A through Frac‐E) from HIV‐infected cells by sequential differential ultracentrifugation (DUC). All fractions showed a heterogeneous size distribution with median particle sizes greater than 100 nm for Frac‐A through Frac‐D but not for Frac‐E, which contained small EPs with an average size well below 50 nm. Synchronized and released cultures contained large infectious EPs in Frac‐A, with markers of amphisomes and viral components. Additionally, Frac‐E uniquely contained EPs positive for CD63, HSP70, and HIV‐1 proteins. Despite its small average size, Frac‐E contained membrane‐protected viral integrase, detectable only after SDS treatment, indicating that it is enclosed in vesicles. Single particle analysis with dSTORM further supported these findings as CD63, HIV‐1 integrase, and the viral surface envelope (Env) glycoprotein (gp) colocalized on the same Frac‐E particles. Surprisingly, Frac‐E EPs were infectious, and infectivity was significantly reduced by immunodepleting Frac‐E with anti‐CD63, indicating the presence of this protein on the surface of infectious small EPs in Frac‐E. To our knowledge, this is the first time that extracellular vesicle (EV) isolation methods have identified infectious small HIV‐1 particles (smHIV‐1) that are under 50 nm. Collectively, our data indicate that the crossroads between EPs and HIV‐1 potentially extend beyond the currently accepted biophysical properties of HIV‐1, which may have further implications for viral pathogenesis. [ABSTRACT FROM AUTHOR]

Background: Initiation of ART among people living with HIV (PLWH) having a CD4 count ≤ 350cells/µl, produces poor immunological recovery, putting them at a high risk of opportunistic infections. To mitigate this, PLWH on ART in Uganda frequently use herbal remedies like Artemisia annua and Moringa oleifera, but their clinical benefits and potential antiretroviral (ARV) interactions remain unknown. This study examined the impact of A. annua and M. oleifera on CD4 count, viral load, and potential ARV interactions among PLWH on ART at an HIV clinic in Uganda. Methods: 282 HIV-positive participants on antiretroviral therapy (ART) with a CD4 count ≤ 350cells/µl were randomized in a double-blind clinical trial to receive daily, in addition to their routine standard of care either; 1) A. annua leaf powder, 2) A. annua plus M. oleifera, and 3) routine standard of care only. Change in the CD4 count at 12 months was our primary outcome. Secondary outcomes included changes in viral load, complete blood count, and ARV plasma levels. Participants were followed up for a year and outcomes were measured at baseline, 6 and 12 months. Results: At 12 months of patient follow-up, in addition to standard of care, administration of A. annua + M. oleifera resulted in an absolute mean CD4 increment of 105.06 cells/µl, (p < 0.001), while administration of A. annua plus routine standard of care registered an absolute mean CD4 increment of 60.84 cells/µl, (p = 0.001) compared to the control group. The A. annua plus M. oleifera treatment significantly reduced viral load (p = 0.022) and increased platelet count (p = 0.025) and white blood cell counts (p = 0.003) compared to standard care alone, with no significant difference in ARV plasma levels across the groups. Conclusion: A combination of A. annua and M. oleifera leaf powders taken once a day together with the routine standard of care produced a significant increase in CD4 count, WBCs, platelets, and viral load suppression among individuals on ART. A. annua and M. oleifera have potential to offer an affordable alternative remedy for managing HIV infection, particularly in low-resource communities lacking ART access. Trial registration: ClinicalTrials.gov NCT03366922. [ABSTRACT FROM AUTHOR]

The field of retroviral integration research has a long history that started with the provirus hypothesis and subsequent discoveries of the retroviral reverse transcriptase and integrase enzymes. Because both enzymes are essential for retroviral replication, they became valued targets in the effort to discover effective compounds to inhibit HIV-1 replication. In 2007, the first integrase strand transfer inhibitor was licensed for clinical use, and subsequently approved second-generation integrase inhibitors are now commonly co-formulated with reverse transcriptase inhibitors to treat people living with HIV. International meetings specifically focused on integrase and retroviral integration research first convened in 1995, and this paper is part of the Viruses Special Issue on the 7th International Conference on Retroviral Integration, which was held in Boulder Colorado in the summer of 2023. Herein, we overview key historical developments in the field, especially as they pertain to the development of the strand transfer inhibitor drug class. Starting from the mid-1990s, research advancements are presented through the lens of the international conferences. Our overview highlights the impact that regularly scheduled, subject-specific international meetings can have on community-building and, as a result, on field-specific collaborations and scientific advancements. [ABSTRACT FROM AUTHOR]

Most plants have developed unique mechanisms to cope with harsh environmental conditions to compensate for their lack of mobility. A key part of their coping mechanisms is the synthesis of secondary metabolites. In addition to their role in plants' defense against pathogens, they also possess therapeutic properties against diseases, and their use by humans predates written history. Viruses are a unique class of submicroscopic agents, incapable of independent existence outside a living host. Pathogenic viruses continue to pose a significant threat to global health, leading to innumerable fatalities on a yearly basis. The use of medicinal plants as a natural source of antiviral agents has been widely reported in literature in the past decades. Metabolomics is a powerful research tool for the identification of plant metabolites with antiviral potentials. It can be used to isolate compounds with antiviral capacities in plants and study the biosynthetic pathways involved in viral disease progression. This review discusses the use of medicinal plants as antiviral agents, with a special focus on the metabolomics evidence supporting their efficacy. Suggestions are made for the optimization of various metabolomics methods of characterizing the bioactive compounds in plants and subsequently understanding the mechanisms of their operation. [ABSTRACT FROM AUTHOR]

Allosteric HIV-1 Integrase (IN) Inhibitors or ALLINIs bind at the dimer interface of the IN, away from the enzymatic catalytic site, and disable viral replication by inducing over-multimerization of IN. Interestingly, these inhibitors are capable of impacting both the early and late stages of viral replication. To better understand the important binding features of multi-substituted quinoline-based ALLINIs, we have surveyed published studies on IN multimerization and antiviral properties of various substituted quinolines at the 4, 6, 7, and 8 positions. Here we show how the efficacy of these inhibitors can be modulated by the nature of the substitutions at those positions. These features not only improve the overall antiviral potencies of these compounds but also significantly shift the selectivity toward the viral maturation stage. Thus, to fully maximize the potency of ALLINIs, the interactions between the inhibitor and multiple IN subunits need to be simultaneously optimized. [ABSTRACT FROM AUTHOR]

Coronavirus disease 2019 (COVID-19) is an infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Available data suggests that there is no specific treatment in modern medicine. The present study aims to investigate the anti-SARSCoV-2 activity of Annona muricata and Artemisia annua leaf extracts as well as their acute toxicity study. The phytochemical screening of the plant extracts was done using standard methods. The acute toxicity evaluation was done using Lorke method. The antiviral activity screening against SARS-CoV-2 was done by plaque assay with Vero-E6 cell line. Phytochemical screening revealed the presence of alkaloids, terpenoids, flavonoids, saponins and steroids in both plants, while tannins and anthraquinone glycosides were absent. The quantification of SARS-CoV-2 by plaque assay with Vero (E6) cell line shows a reduction in the zones of cellular death for the extracts treated cells compared to the cells with the virus only. Acute toxicity test showed no death or abnormal behavioural changes in mice during and after 24 hours of extract administration. On the basis of the results obtained from the present study, both Annona muricata and Artemisia annua leaf extracts are considered to have some level of inhibition against SARS-CoV-2 virus growth. [ABSTRACT FROM AUTHOR]

Integration of HIV-1 genomic cDNA results in the formation of single-strand breaks in cellular DNA, which must be repaired for efficient viral replication. Post-integration DNA repair mainly depends on the formation of the HIV-1 integrase complex with the Ku70 protein, which promotes DNA-PK assembly at sites of integration and its activation. Here, we have developed a first-class inhibitor of the integrase-Ku70 complex formation that inhibits HIV-1 replication in cell culture by acting at the stage of post-integration DNA repair. This inhibitor, named s17, does not affect the main cellular function of Ku70, namely its participation in the repair of double-strand DNA breaks through the non-homologous end-joining pathway. Using a molecular dynamics approach, we have constructed a model for the interaction of s17 with Ku70. According to this model, the interaction of two phenyl radicals of s17 with the L76 residue of Ku70 is important for this interaction. The requirement of two phenyl radicals in the structure of s17 for its inhibitory properties was confirmed using a set of s17 derivatives. We propose to stimulate compounds that inhibit post-integration repair by disrupting the integrase binding to Ku70 KuINins. [ABSTRACT FROM AUTHOR]

The authors of the article respond to a comment made by Dr. Žarko Kulić regarding their research on the antiviral activity of Pelargonium sidoides extracts. They clarify that their aim was to identify antiviral bioactive compounds from the extracts, not to characterize the plant material used. They obtained the extracts from a reputable company in South Africa and performed extraction methods twice to ensure reproducibility. They identified the dominant compound in the extract as Neoilludin B and are currently conducting further studies to validate their findings. They acknowledge an error in the use of an incorrect image in the original manuscript and express their commitment to addressing all comments and concerns. [Extracted from the article]

Background: Several anti-retroviral drugs are available against Human immunodeficiency virus type-1, but have multiple adverse side effects. Hence, there is an incessant compulsion for effectual anti-retroviral agents with minimal or no intricacy. Traditionally, natural products have been the most successful source for the development of new medications. Withania somnifera, also known as Ashwagandha, is the utmost treasured medicinal plant used in Ayurveda, which holds the potential to give adaptogenic, immunomodulatory, and antiviral effects. However, its effect on HIV-1 replication at the cellular level has never been explored. Herein, we focused on the anti-HIV-1 activity and the probable mechanism of action of hydroalcoholic and aqueous extracts of Withania somnifera roots and its phytomolecules. Methods: The cytotoxicity of the extracts was determined through MTT assay, while the in vitro anti-HIV-1 activity was assessed in TZM-bl cells against the HIV-1 strains of X4 and R5 subtypes. Results were confirmed in peripheral blood mononuclear cells, using the HIV-1 p24 antigen assay. Additionally, the mechanism of action was determined through the Time of Addition assay, which was further validated through the series of enzymatic assays, i.e. HIV-1 Integrase, Reverse transcriptase, and Protease assays. To explore the role of the identified active metabolites of Withania somnifera in antiretroviral activity, molecular docking analyses were performed against these key HIV-1 replication enzymes. Results: The hydroalcoholic and aqueous extracts of Withania somnifera roots were found to be safer at the sub-cytotoxic concentrations and exhibited their ability to inhibit replication of two primary isolates of HIV-1 through cell-associated and cell-free assays, in dose-dependent kinetics. Several active phytomolecules found in Withania somnifera successfully established hydrogens bonds in the active binding pocket site residues responsible for the catalytic activity of HIV replication and therefore, signifying their role in the attenuation of HIV-1 infection as implied through the in silico molecular docking studies. Conclusions: Our research identified both the hydroalcoholic and aqueous extracts of Withania somnifera roots as potent inhibitors of HIV-1 infection. The in silico analyses also indicated the key components of Withania somnifera with the highest binding affinity against the HIV-1 Integrase by 12-Deoxywithastramonolide and 27-Hydroxywithanone, HIV-1 Protease by Ashwagandhanolide and Withacoagin, and HIV-1 Reverse transcriptase by Ashwagandhanolide and Withanolide B, thereby showing possible mechanisms of HIV-1 extenuation. Overall, this study classified the role of Withania somnifera extracts and their active compounds as potential agents against HIV-1 infection. [ABSTRACT FROM AUTHOR]

In the effort to identify and develop new HIV-1 inhibitors endowed with innovative mechanisms, we focused our attention on the possibility to target more than one viral encoded enzymatic function with a single molecule. In this respect, we have previously identified by virtual screening a new indolinonebased scaffold for dual allosteric inhibitors targeting both reverse transcriptaseassociated functions: polymerase and RNase H. Pursuing with the structural optimization of these dual inhibitors, we synthesized a series of 35 new 3-[2-(4- aryl-1,3-thiazol-2-ylidene)hydrazin-1-ylidene]1-indol-2-one and 3-[3-methyl4-arylthiazol-2-ylidene)hydrazine-1-ylidene)indolin-2-one derivatives, which maintain their dual inhibitory activity in the low micromolar range. Interestingly, compounds 1a, 3a, 10a, and 9b are able to block HIV-1 replication with EC50 < 20 µM. Mechanism of action studies showed that such compounds could block HIV-1 integrase. In particular, compound 10a is the most promising for further multitarget compound development. [ABSTRACT FROM AUTHOR]

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has so far infected 762 million people with over 6.9 million deaths worldwide. Broad-spectrum viral inhibitors that block the initial stages of infection by reducing virus binding and proliferation, thereby reducing disease severities, are still an unmet global medical need. We studied Bi121, which is a standardized polyphenolic-rich compound isolated from Pelargonium sidoides, against recombinant vesicular stomatitis virus (rVSV)-pseudotyped SARS-CoV-2S (mutations in the spike protein) of six different variants of SARS-CoV-2. Bi121 was effective at neutralizing all six rVSV-ΔG-SARS-CoV-2S variants. The antiviral activity of Bi121 was also assessed against SARS-CoV-2 variants (USA WA1/2020, Hongkong/VM20001061/2020, B.1.167.2 (Delta), and Omicron) in Vero cells and HEK-ACE2 cell lines using RT-qPCR and plaque assays. Bi121 showed significant antiviral activity against all the four SARS-CoV-2 variants tested, suggesting a broad-spectrum activity. Bi121 fractions generated using HPLC showed antiviral activity in three fractions out of eight against SARS-CoV-2. The dominant compound identified in all three fractions using LC/MS/MS analysis was Neoilludin B. In silico structural modeling studies with Neoilludin B showed that it has a novel RNA-intercalating activity toward RNA viruses. In silico findings and the antiviral activity of this compound against several SARS-CoV-2 variants support further evaluation as a potential treatment of COVID-19. [ABSTRACT FROM AUTHOR]

The mature HIV-1 capsid is stabilized by host and viral determinants. The capsid protein CA binds to the cellular metabolites inositol hexakisphosphate (IP6) and its precursor inositol (1, 3, 4, 5, 6) pentakisphosphate (IP5) to stabilize the mature capsid. In target cells, capsid destabilization by the antiviral compounds lenacapavir and PF74 reveals a HIV-1 infectivity defect due to IP5/IP6 (IP5/6) depletion. To test whether intrinsic HIV-1 capsid stability and/or host factor binding determines HIV-1 insensitivity to IP5/6 depletion, a panel of CA mutants was assayed for infection of IP5/6-depleted T cells and wildtype cells. Four CA mutants with unstable capsids exhibited dependence on host IP5/6 for infection and reverse transcription (RTN). Adaptation of one such mutant, Q219A, by spread in culture resulted in Vpu truncation and a capsid three-fold interface mutation, T200I. T200I increased intrinsic capsid stability as determined by in vitro uncoating of purified cores and partially reversed the IP5/6-dependence in target cells for each of the four CA mutants. T200I further rescued the changes to lenacapavir sensitivity associated with the parental mutation. The premature dissolution of the capsid caused by the IP5/6-dependent mutations imparted a unique defect in integration targeting that was rescued by T200I. Collectively, these results demonstrate that T200I restored other capsid functions after RTN for the panel of mutants. Thus, the hyperstable T200I mutation stabilized the instability defects imparted by the parental IP5/6-dependent CA mutation. The contribution of Vpu truncation to mutant adaptation was linked to BST-2 antagonization, suggesting that cell-to-cell transfer promoted replication of the mutants. We conclude that interactions at the three-fold interface are adaptable, key mediators of capsid stability in target cells and are able to antagonize even severe capsid instability to promote infection. Author summary: The US FDA- and EU-approved compound lenacapavir targets the HIV-1 capsid protein (CA) to inhibit HIV-1 infection. Lenacapavir blocks infection at multiple stages of infection offering benefits over current antiretroviral therapies. Identification of other capsid surfaces that can be targeted by next-generation inhibitors is of importance due to the high mutability of the virus. We sought to identify surfaces on CA that modulate capsid stability in the cell targeted for infection by HIV-1. We identified CA substitutions that render the virus dependent on host metabolites that promote capsid stability. Replication of the host factor-dependent virus resulted in alteration of the Vpu protein and a second site on CA that stabilized the capsid. The compensatory mutation suppressed defects associated with the parental mutation, including decreased infection, changes to lenacapavir antiviral efficiency, and redistribution of integration targeting, a characteristic that affects latency. Our study reveals that an interface between the hexameric CA units in the capsid adapts to conditions that destabilize the capsid and that this interface is a primary modulator of capsid stability in target cells. We propose targeting of our identified capsid surfaces with inhibitors to increase lenacapavir potency and efficacy in patients. [ABSTRACT FROM AUTHOR]

Viral infections represent one of the main causes of disease worldwide, especially today due to the increase of migration, global travel, and urbanization. The several side effects of the conventional drugs and the growing phenomenon of resistance have led researchers to turn to the plant kingdom as a source of potential new antiviral drugs. The aim of this work is to summarize the updated evidence for antiviral activity of different plants and their isolated bioactive compounds, evaluating also the potential interactions, which can occur in cotreatment with conventional antiviral drugs. The plant complexes have proved to be usually more active than their most abundant isolated compounds by hypothesizing synergistic mechanisms. In addition to cellular and molecular investigations, molecular docking studies have proved essential in highlighting the interaction mechanisms of bioactive compounds with target molecules. However, the use of nonstandardized extracts, or too high concentrations in vitro, which do not reproduce their bioavailability in vivo, are often limiting factors. Moreover, the lack of studies concerning the safety profile of plant extracts and their isolated compounds, alone or in combination with conventional antiviral drugs, is the most worrying aspect. In light of this, further studies are needed to validate their possible therapeutic use., (© 2019 John Wiley & Sons, Ltd.)

The emergence of drug-resistant Human Immunodeficiency Virus-1 strains against anti-HIV therapies in the clinical pipeline, and the persistence of HIV in cellular reservoirs remains a significant concern. Therefore, there is a continuous need to discover and develop new, safer, and effective drugs targeting novel sites to combat HIV-1. The fungal species are gaining increasing attention as alternative sources of anti-HIV compounds or immunomodulators that can escape the current barriers to cure. Despite the potential of the fungal kingdom as a source for diverse chemistries that can yield novel HIV therapies, there are few comprehensive reports on the progress made thus far in the search for fungal species with the capacity to produce anti-HIV compounds. This review provides insights into the recent research developments on natural products produced by fungal species, particularly fungal endophytes exhibiting immunomodulatory or anti-HIV activities. In this study, we first explore currently existing therapies for various HIV-1 target sites. Then we assess the various activity assays developed for gauging antiviral activity production from microbial sources since they are crucial in the early screening phases for discovering novel anti-HIV compounds. Finally, we explore fungal secondary metabolites compounds that have been characterized at the structural level and demonstrate their potential as inhibitors of various HIV-1 target sites. [ABSTRACT FROM AUTHOR]

Transmissions of simian viruses to humans has originated the different groups of HIV-1. We recently identified a functional motif (CLA), in the C-terminal domain of the integrase, essential for integration in HIV-1 group M. Here, we found that the motif is instead dispensable in group O isolates, because of the presence, in the N-terminal domain of HIV-1 O of a specific sequence, Q7G27P41H44, that we define as the NOG motif. Alterations of reverse transcription and of 3' processing observed by mutating the CLA motif of IN M are fully rescued to wt levels by inserting the sequence of the NOG motif in the N-ter of the protein. These results indicate that the two motifs (CLA and NOG) functionally complement each other and a working model accounting for these observations is proposed. The establishment of these two alternative motifs seems to be due to the different phylogenetic origin and history of these two groups. Indeed, the NOG motif is already present in the ancestor of group O (SIVgor) while it is absent from SIVcpzPtt, the ancestor of group M. The CLA motif, instead, seems to have emerged after SIVcpzPtt has been transferred to humans, since no conservation is found at the same positions in these simian viruses. These results show the existence of two-group specific motifs in HIV-1 M and O integrases. In each group, only one of the motifs is functional, potentially leading the other motif to diverge from its original function and, in an evolutionary perspective, assist other functions of the protein, further increasing HIV genetic diversity. Author summary: HIV-1 establishes a permanent infection by integrating its genome in the one of the host cell. This key step is achieved by the viral integrase (IN), which is composed of three domains: N-terminal (NTD), catalytic core (CCD), and C-terminal (CTD). We recently reported, in the CTD, the existence of a functional motif (CLA) that is essential for integration in HIV-1 group M. Here we show that in HIV-1 group O, the function exerted by the integrase M CLA motif is instead ensured by another motif (NOG), located in the NTD of the protein. We show that these two motifs can functionally complement each other by acting on the same steps of the viral cycle, highlighting for the first time such an important functional duality between HIV-1 integrases. We also show that, while the NOG motif in HIV-1 O was likely inherited from its simian ancestor (SIVgor), the CLA motif seems to have emerged in the human host, probably in the process of adaptation to the new host of the simian virus on its way to become HIV-1 M. Such a requirement did not seem to have existed for HIV-1 O, possibly because of the presence of the NOG motif. [ABSTRACT FROM AUTHOR]

Antiretroviral therapy is the only treatment option for HIV-infected patients; however, it has certain drawbacks in terms of developing multiple toxic side effects. Thus, there is a continuous need to explore safe and efficacious anti-retroviral agents. Carica papaya Linn and Psidium guajava are known for their various biological activities. In this study, we characterized the bioactive fractions of methanolic leaves extract from both plants using the High-resolution electrospray ionization mass spectrometry (HR-ESI–MS) technique, followed by the investigation of their potential as anti-HIV-1 and antioxidant agents through in vitro mechanistic assays. The anti-HIV-1 activity was examined in TZM-bl cells through luciferase gene assay against two different clades of HIV-1 strains, whereas the intracellular ROS generation was analyzed by Fluorescence-Activated Cell Sorting. Additionally, the mechanisms of action of these phyto-extracts were determined through the Time-of-addition assay. The characterization of Carica papaya Linn and Psidium guajava leaves extract through HR-ESI–MS fragmentation showed high enrichment of various alkaloids, glycosides, lipids, phenolic compounds, terpenes, and fatty acids like bioactive constituents. Both the phyto-extracts were found to be less toxic and exhibited potent antiviral activity against HIV-1 strains. Furthermore, the phyto-extracts also showed a decreased intracellular ROS in HIV-1 infected cells due to their high antioxidant potential. Overall, our study suggests the anti-HIV-1 potential of Carica papaya Linn and Psidium guajava leaves extract due to the synergistic action of multiple bioactive constituents. [ABSTRACT FROM AUTHOR]

HIV integrase (IN) inserts viral DNA into the host genome and is the target of the strand transfer inhibitors (STIs), a class of small molecules currently in clinical use. Another potent class of antivirals is the allosteric inhibitors of integrase, or ALLINIs. ALLINIs promote IN aggregation by stabilizing an interaction between the catalytic core domain (CCD) and carboxy-terminal domain (CTD) that undermines viral particle formation in late replication. Ongoing challenges with inhibitor potency, toxicity, and viral resistance motivate research to understand their mechanism. Here, we report a 2.93 Å X-ray crystal structure of the minimal ternary complex between CCD, CTD, and the ALLINI BI-224436. This structure reveals an asymmetric ternary complex with a prominent network of π-mediated interactions that suggest specific avenues for future ALLINI development and optimization. Author summary: The global burden of the HIV/AIDS pandemic and continued emergence of drug resistance drives the need for novel antivirals. The allosteric integrase inhibitors, or ALLINIs, are a potent class of antivirals in development that target the enzyme integrase in a surprising fashion: the small molecules act to stabilize an inappropriate protein-protein interaction to attain biological effect. Here, we report the first atomic resolution (2.93 Å) X-ray crystal structure of the minimal ternary complex between domains of the integrase and the first ALLINI preclinical lead BI-224436. Our structure provides a more precise view of the molecular interactions that underlie drug potency, and several aspects of our data suggest routes to improving ALLINI design and minimizing acquisition of resistance. [ABSTRACT FROM AUTHOR]

The article "Significant Broad-Spectrum Antiviral Activity of Bi121 against Different Variants of SARS-CoV-2" by Subhadra et al. discusses the antiviral activity of a standardized extract called Bi121 from Pelargonium sidoides against various SARS-CoV-2 variants. The authors identify a specific compound called Neoilludin B as the active component and demonstrate its RNA-intercalating activity in RNA viruses through structural modeling. However, the article raises concerns about the lack of comprehensive characterization of the study material, including the authentication of the plant material used and the absence of a clear standardization parameter for Bi121. The article also questions the similarity between the HPLC profile presented in this study and a previously published profile, suggesting possible plagiarism. Additionally, the identification of Neoilludin B as an active substance is deemed putative and requires further evidence. These concerns limit the validity of the study and call for a more thorough phytochemical reassessment of the material. [Extracted from the article]

Psychological stress exerts its effects mainly through the release of corticotropin releasing hormone (CRH), which activates inflammatory pathways in skin (inter alia), resulting in redness, extracellular matrix degradation, loss of skin elasticity and firmness, and the appearance of wrinkles—namely, accelerated skin aging. In order to propose a solution to this neurogenic aging phenomenon, we report here on studies using a myricitrin-rich extract of Cistus incanus, a Mediterranean shrub used in traditional medicine for the treatment of inflammatory and other diseases. These studies include a CRH receptor (CRH-R1) blocking assay; in vitro inflammatory cytokine reduction under CRH stimulation, and ex vivo NF-kB inhibition; and a double-blind clinical trial performed on highly stressed panelists, evaluating skin inflammation and wrinkling (active formulation vs. placebo control, applied split-face following a computer-generated randomization scheme; 36 subjects recruited and randomized, 30 analyzed; no adverse effects recorded; EMA/INFARMED registration #118505, internally funded). The results show that this extract can effectively block the CRH-R1 receptor, preventing NF-κB activation and the production of related pro-inflammatory cytokines. In a clinical setting, this same extract delivered significant anti-inflammatory and anti-aging effects. Taken together, these results demonstrate the value of this extract as a cosmetic active to counter neurogenic inflammation and skin aging. [ABSTRACT FROM AUTHOR]

Reverse transcriptase (RT) and integrase (IN) are encoded tandemly in the pol genes of retroviruses. We reported recently that HIV-1 RT and IN need to be supplied as the pol precursor intermediates, in which RT and IN are in fusion form (RTIN) to exert efficient reverse transcription in the context of HIV-1 replication. The mechanism underlying RTIN's effect, however, remains to be elucidated. In this study, we examined the effect of IN fusion on RT during reverse transcription by an in vitro cell-free assay, using recombinant HIV-1 RTIN (rRTIN). We found that, compared to recombinant RT (rRT), rRTIN generated significantly higher cDNAs under physiological concentrations of dNTPs (less than 10 μM), suggesting increased affinity of RTIN to dNTPs. Importantly, the cleavage of RTIN with HIV-1 protease reduced cDNA levels at a low dose of dNTPs. Similarly, sensitivities against RT inhibitors were significantly altered in RTIN form. Finally, analysis of molecular dynamics simulations of RT and RTIN suggested that IN can influence the structural dynamics of the RT active center and the inhibitor binding pockets in cis. Thus, we demonstrated, for the first time, the cis-allosteric regulatory roles of IN in RT structure and enzymatic activity. [ABSTRACT FROM AUTHOR]

African swine fever virus (ASFV) causes a viral disease in swine with a mortality rate of approximately 100%, threatening the global pig industry's economic development. However, vaccines are not yet commercially available, and other antiviral therapeutics, such as antiviral drugs, are urgently needed. In this study, berbamine hydrochloride, a natural bis-benzylisoquinoline alkaloid isolated from the traditional Chinese herb Berberis amurensis, showed significant antiviral activity against ASFV. The 50% cytotoxic concentration (CC50) of berbamine hydrochloride in porcine alveolar macrophages (PAMs) was 27.89 μM. The antiviral activity assay demonstrated that berbamine hydrochloride inhibits ASFV in a dose-dependent manner. In addition, a 4.14 log TCID50 decrease in the viral titre resulting from non-cytotoxic berbamine hydrochloride was found. Moreover, the antiviral activity of berbamine hydrochloride was maintained for 48h and took effect at multiplicities of infection (MOI) of 0.01, 0.1, and 1. The time-of-addition analysis revealed an inhibitory effect throughout the entire virus life-cycle. A subsequent viral entry assay verified that berbamine hydrochloride blocks the early stage of ASFV infection. Moreover, similar anti-ASFV activity of berbamine hydrochloride was also found in PK-15 and 3D4/21 cells. In summary, these results indicate that berbamine hydrochloride is an effective anti-ASFV natural product and may be considered a novel antiviral drug. [ABSTRACT FROM AUTHOR]

Previously, we reported that an HIV-1 variant containing Met-to-Ile change at codon 50 and Val-to-Ile mutation at codon 151 of integrase (IN), HIV(IN:M50I/V151I), was an impaired virus. Despite the mutations being in IN, the virus release was significantly suppressed (p < 0.0001) and the initiation of autoprocessing was inhibited; the mechanism of the defect remains unknown. In the current study, we attempted to identify the critical domains or amino acid (aa) residue(s) that promote defects in HIV(IN:M50I/V151I), using a series of variants, including truncated or aa-substituted RNase H (RH) or IN. The results demonstrated that virus release and the initiation of autoprocessing were regulated by the C-terminal domains (CTDs) of RH and IN. Further studies illustrated that Asp at codon 109 of RH CTD and Asp at the C terminus of IN induces the defect. This result indicated that the CTDs of RH and IN in GagPol and particular aa positions in RH and IN regulated the virus release and the initiation of autoprocessing, and these sites could be potential targets for the development of new therapies. [ABSTRACT FROM AUTHOR]

Genus Cistus L. (Cistaceae) occurs mainly in the Mediterranean region. Many plants of this genus have been used in different folk medicines since ancient times, mainly against peptic ailments and skin disorders, such as burns, wounds, and infections. Over the last years, Cistus species have attracted great interest due to the variety of their bioactive compounds and pharmacological activities. Comprehensive research of previously published literature was performed for the collection of the available data on the traditional uses, phytochemistry, and pharmacological properties of the genus Cistus since 2014, using electronic databases with several key search words. A total of 66 published scientific studies was examined. Over the last years, 111 secondary metabolites have been described in Cistus taxa. In vitro and in vivo studies, as well as a clinical study are included, highlighting the interesting pharmacological aspects of these plants. This review summarizes and discusses the current knowledge of the traditional uses, phytochemistry, and biological activities of Cistus plants, revealing the uncharted scientific territory and may provide critical information for future perspectives on these plants. [ABSTRACT FROM AUTHOR]

Recent evidence indicates that the HIV-1 Integrase (IN) binds the viral genomic RNA (gRNA), playing a critical role in the morphogenesis of the viral particle and in the stability of the gRNA once in the host cell. By combining biophysical, molecular biology, and biochemical approaches, we found that the 18-residues flexible C-terminal tail of IN acts as a sensor of the peculiar apical structure of the trans-activation response element RNA (TAR), interacting with its hexaloop. We show that the binding of the whole IN C-terminal domain modifies TAR structure, exposing critical nucleotides. These modifications favour the subsequent binding of the HIV transcriptional trans-activator Tat to TAR, finally displacing IN from TAR. Based on these results, we propose that IN assists the binding of Tat to TAR RNA. This working model provides a mechanistic sketch accounting for the emerging role of IN in the early stages of proviral transcription and could help in the design of anti-HIV-1 therapeutics against this new target of the viral infectious cycle. [ABSTRACT FROM AUTHOR]

There is an increasingly urgent call to shift industrial processes from fossil fuel feedstock to sustainable bio-based resources. This change becomes of high importance considering new budget requirements for a carbon-neutral economy. Such a transformation can be driven by traditionally used plants that are able to produce large amounts of valuable biologically relevant secondary metabolites. Tobacco plants can play a leading role in providing value-added products in remote areas of the world. In this study, we propose a non-exhaustive list of compounds with potential economic interest that can be sourced from the tobacco plant. In order to optimize extraction methodologies, we first analyzed their physico-chemical properties using rapid solubility tests and high-resolution microfractionation techniques. Next, to identify an optimal extraction for a selected list of compounds, we compared 13 different extraction method–solvent combinations. We proceeded with profiling some of these compounds in a total of six varieties from Nicotiana tabacum and Nicotiana rustica species, identifying the optimal variety for each. The estimated expected yields for each of these compounds demonstrate that tobacco plants can be a superior source of valuable compounds with diverse applications beyond nicotine. Among the most interesting results, we found high variability of anatabine content between species and varieties, ranging from 287 to 1699 µg/g. In addition, we found that CGA (1305 µg/g) and rutin (7910 µg/g) content are orders of magnitude lower in the Burley variety as compared to all others. [ABSTRACT FROM AUTHOR]

The Covid-19 pandemic has elicited much laboratory and clinical research attention on vaccines, mAbs, and certain small-molecule antivirals against SARS-CoV-2 infection. By contrast, there has been comparatively little attention on plant-derived compounds, especially those that are understood to be safely ingested at common doses and are frequently consumed in the diet in herbs, spices, fruits and vegetables. Examining plant secondary metabolites, we review recent elucidations into the pharmacological activity of flavonoids and other polyphenolic compounds and also survey their putative frequent-hitter behavior. Polyphenols, like many drugs, are glucuronidated post-ingestion. In an inflammatory milieu such as infection, a reversion back to the active aglycone by the release of β-glucuronidase from neutrophils and macrophages allows cellular entry of the aglycone. In the context of viral infection, virions and intracellular virus particles may be exposed to promiscuous binding by the polyphenol aglycones resulting in viral inhibition. As the mechanism's scope would apply to the diverse range of virus species that elicit inflammation in infected hosts, we highlight pre-clinical studies of polyphenol aglycones, such as luteolin, isoginkgetin, quercetin, quercetagetin, baicalein, curcumin, fisetin and hesperetin that reduce virion replication spanning multiple distinct virus genera. It is hoped that greater awareness of the potential spatial selectivity of polyphenolic activation to sites of pathogenic infection will spur renewed research and clinical attention for natural products antiviral assaying and trialing over a wide array of infectious viral diseases. [ABSTRACT FROM AUTHOR]

Allosteric integrase (IN) inhibitors (ALLINIs), which are promising preclinical compounds that engage the lens epithelium-derived growth factor (LEDGF)/p75 binding site on IN, can inhibit different aspects of human immunodeficiency virus 1 (HIV-1) replication. During the late phase of replication, ALLINIs induce aberrant IN hyper-multimerization, the consequences of which disrupt IN binding to genomic RNA and virus particle morphogenesis. During the early phase of infection, ALLINIs can suppress HIV-1 integration into host genes, which is also observed in LEDGF/p75-depelted cells. Despite this similarity, the roles of LEDGF/p75 and its paralog hepatoma-derived growth factor like 2 (HDGFL2) in ALLINI-mediated integration retargeting are untested. Herein, we mapped integration sites in cells knocked out for LEDGF/p75, HDGFL2, or both factors, which revealed that these two proteins in large part account for ALLINI-mediated integration retargeting during the early phase of infection. We also determined that ALLINI-treated viruses are defective during the subsequent round of infection for integration into genes associated with speckle-associated domains, which are naturally highly targeted for HIV-1 integration. Class II IN mutant viruses with alterations distal from the LEDGF/p75 binding site moreover shared this integration retargeting phenotype. Altogether, our findings help to inform the molecular bases and consequences of ALLINI action. [ABSTRACT FROM AUTHOR]

Human immunodeficiency virus-1 (HIV-1) is the major cause of acquired immunodeficiency syndrome (AIDs) worldwide. In HIV-1 infection, innate immunity is the first defensive line for immune recognition and viral clearance to ensure the normal biological function of the host cell and body health. Under the strong selected pressure generated by the human body over thousands of years, HIV has evolved strategies to counteract and deceive the innate immune system into completing its lifecycle. Recently, several studies have demonstrated that HIV capsid core which is thought to be a protector of the cone structure of genomic RNA, also plays an essential role in escaping innate immunity surveillance. This mini-review summarizes the function of capsid in viral immune evasion, and the comprehensive elucidation of capsid-host cell innate immunity interaction could promote our understanding of HIV-1's pathogenic mechanism and provide insights for HIV-1 treatment in clinical therapy. [ABSTRACT FROM AUTHOR]

Hepatitis C virus is the major cause of chronic liver diseases and the only cytoplasmic RNA virus known to be oncogenic in humans. The viral genome gives rise to ten mature proteins and to additional proteins, which are the products of alternative translation initiation mechanisms. A protein—known as ARFP (alternative reading frame protein) or Core+1 protein—is synthesized by an open reading frame overlapping the HCV Core coding region in the (+1) frame of genotype 1a. Almost 20 years after its discovery, we still know little of the biological role of the ARFP/Core+1 protein. Here, our differential proteomic analysis of stable hepatoma cell lines expressing the Core+1/Long isoform of HCV-1a relates the expression of the Core+1/Long isoform with the progression of the pathology of HCV liver disease to cancer. [ABSTRACT FROM AUTHOR]

Exploration of the traditional medicinal plants is essential for drug discovery and development for various pharmacological targets. Various phytochemicals derived from medicinal plants were extensively studied for antiviral activity. This review aims to highlight the role of medicinal plants against viral infections that remains to be the leading cause of human death globally. Antiviral properties of phytoconstituents isolated from 45 plants were discussed for five different types of viral infections. The ability of the plants' active compounds with antiviral effects was highlighted as well as their mechanism of action, pharmacological studies, and toxicological data on a variety of cell lines. The experimental values, such as IC50, EC50, CC50, ED50, TD50, MIC100, and SI of the active compounds, were compiled and discussed to determine their potential. Among the plants mentioned, 11 plants showed the most promising medicinal plants against viral infections. Sambucus nigra and Clinacanthus nutans manifested antiviral activity against three different types of viral infections. Echinacea purpurea, Echinacea augustofolia, Echinacea pallida, Plantago major, Glycyrrhiza uralensis, Phyllanthus emblica, Camellia sinensis, and Cistus incanus exhibited antiviral activity against two different types of viral infections. Interestingly, Nicotiana benthamiana showed antiviral effects against mosquito-borne infections. The importance of phenolic acids, alkamides, alkylamides, glycyrrhizin, epicatechin gallate (ECG), epigallocatechin gallate (EGCG), epigallocatechin (EGC), protein-based plant-produced ZIKV Envelope (PzE), and anti-CHIKV monoclonal antibody was also reviewed. An exploratory approach to the published literature was conducted using a variety of books and online databases, including Scopus, Google Scholar, ScienceDirect, Web of Science, and PubMed Central, with the goal of obtaining, compiling, and reconstructing information on a variety of fundamental aspects, especially regarding medicinal plants. This evaluation gathered important information from all available library databases and Internet searches from 1992 to 2022. [ABSTRACT FROM AUTHOR]

Allosteric HIV-1 integrase (IN) inhibitors, or ALLINIs, are a new class of antiviral agents that bind at the dimer interface of the IN, away from the enzymatic catalytic site and block viral replication by triggering an aberrant multimerization of the viral enzyme. To further our understanding of the important binding features of multi-substituted quinoline-based ALLINIs, we have examined the IN multimerization and antiviral properties of substitution patterns at the 6 or 8 position. We found that the binding properties of these ALLINIs are negatively impacted by the presence of bulky substitutions at these positions. In addition, we have observed that the addition of bromine at either the 6 (6-bromo) or 8 (8-bromo) position conferred better antiviral properties. Finally, we found a significant loss of potency with the 6-bromo when tested with the ALLINI-resistant IN A128T mutant virus, while the 8-bromo analog retained full effectiveness. [ABSTRACT FROM AUTHOR]