Background The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in millions of infections worldwide. While the search for an effective antiviral is still ongoing, experimental therapies based on repurposing of available antivirals is being attempted, of which HIV protease inhibitors (PIs) have gained considerable interest. Inhibition profiling of the PIs directly against the viral protease has never been attempted in vitro, and while few studies reported an efficacy of lopinavir and ritonavir in SARS-CoV-2 context, the mechanism of action of the drugs remains to be validated. Methods We carried out an in-depth analysis of the efficacy of HIV PIs against the main protease of SARS-CoV-2 (Mpro) in cell culture and in vitro enzymatic assays, using a methodology that enabled us to focus solely on any potential inhibitory effects of the inhibitors against the viral protease. For cell culture experiments a dark-to-bright GFP reporter substrate system was designed. Results Lopinavir, ritonavir, darunavir, saquinavir, and atazanavir were able to inhibit the viral protease in cell culture, albeit in concentrations much higher than their achievable plasma levels, given their current drug formulations. While inhibition by lopinavir was attributed to its cytotoxicity, ritonavir was the most effective of the panel, with IC50 of 13.7 µM. None of the inhibitors showed significant inhibition of SARS-CoV-2 Mpro in our in vitro enzymatic assays up to 100 µM concentration. Conclusion Targeting of SARS-CoV-2 Mpro by some of the HIV PIs might be of limited clinical potential, given the high concentration of the drugs required to achieve significant inhibition. Therefore, given their weak inhibition of the viral protease, any potential beneficial effect of the PIs in COVID-19 context might perhaps be attributed to acting on other molecular target(s), rather than SARS-CoV-2 Mpro.

Dolutegravir-based antiretroviral therapy (ART) has been scaled up in many developing countries, including Ethiopia. However, subtype-dependent polymorphic differences might influence the occurrence of HIV-drug-resistance mutations (HIVDRMs). We analyzed the prevalence of pre-treatment integrase strand transfer inhibitor (INSTI) HIVDRMs and naturally occurring polymorphisms (NOPs) of the integrase gene, using plasma samples collected as part of the national HIVDR survey in Ethiopia in 2017. We included a total of 460 HIV-1 integrase gene sequences from INSTI-naïve (n = 373 ART-naïve and n = 87 ART-experienced) patients. No dolutegravir-associated HIVDRMs were detected, regardless of previous exposure to ART. However, we found E92G in one ART-naïve patient specimen and accessory mutations in 20/460 (4.3%) of the specimens. Moreover, among the 288 integrase amino acid positions of the subtype C, 187/288 (64.9%) were conserved (<1.0% variability). Analysis of the genetic barrier showed that the Q148H/K/R dolutegravir resistance pathway was less selected in subtype C. Docking analysis of the dolutegravir showed that protease- and reverse-transcriptase-associated HIVDRMs did not affect the native structure of the HIV-1 integrase. Our results support the implementation of a wide scale-up of dolutegravir-based regimes. However, the detection of polymorphisms contributing to INSTI warrants the continuous surveillance of INSTI resistance.

Retroviral protease inhibitors (PIs) are fundamental pillars in the treatment of HIV infection and acquired immunodeficiency syndrome (AIDS). Currently used PIs are designed against HIV-1, and their effect on HIV-2 is understudied. Using a modular HIV-2 protease cassette system, inhibition profiling assays were carried out for protease inhibitors both in enzymatic and cell culture assays. Moreover, the treatment-associated resistance mutations (I54M, L90M) were introduced into the modular system, and comparative inhibition assays were performed to determine their effect on the susceptibility of the protease. Our results indicate that darunavir, saquinavir, indinavir and lopinavir were very effective HIV-2 protease inhibitors, while tipranavir, nelfinavir and amprenavir showed a decreased efficacy. I54M, L90M double mutation resulted in a significant reduction in the susceptibility to most of the inhibitors with the exception of tipranavir. To our knowledge, this modular system constitutes a novel approach in the field of HIV-2 protease characterization and susceptibility testing.

Abstract Background The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in millions of infections worldwide. While the search for an effective antiviral is still ongoing, experimental therapies based on repurposing of available antivirals is being attempted, of which HIV protease inhibitors (PIs) have gained considerable interest. Inhibition profiling of the PIs directly against the viral protease has never been attempted in vitro, and while few studies reported an efficacy of lopinavir and ritonavir in SARS-CoV-2 context, the mechanism of action of the drugs remains to be validated. Methods We carried out an in-depth analysis of the efficacy of HIV PIs against the main protease of SARS-CoV-2 (Mpro) in cell culture and in vitro enzymatic assays, using a methodology that enabled us to focus solely on any potential inhibitory effects of the inhibitors against the viral protease. Results Lopinavir, ritonavir, darunavir, saquinavir, and atazanavir were able to inhibit the viral protease in cell culture, albeit in concentrations much higher than their achievable plasma levels, given their current drug formulations. While inhibition by lopinavir was attributed to its cytotoxicity, ritonavir was the most effective of the panel, with IC50 of 13.7 µM. Atazanavir on the other hand was the only PI to inhibit the viral protease both in cell culture and in our in vitro enzymatic assay. Conclusion Targeting of SARS-CoV-2 Mpro by some of the HIV PIs might be of limited clinical potential, given the high concentration of the drugs required to achieve significant inhibition. Therefore, given their weak inhibition of the viral protease, any potential beneficial effect of the PIs in COVID-19 context might perhaps be attributed to acting on other molecular target(s), rather than SARS-CoV-2 Mpro.

BackgroundThe pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in millions of infections worldwide. While the search for an effective antiviral is still ongoing, experimental therapies based on repurposing of available antivirals is being attempted, of which HIV protease inhibitors (PIs) have gained considerable interest. Inhibition profiling of the PIs directly against the viral protease has never been attempted in vitro, and while few studies reported an efficacy of lopinavir and ritonavir in SARS-CoV-2 context, the mechanism of action of the drugs remains to be validated. MethodsWe carried out an in-depth analysis of the efficacy of HIV PIs against the main protease of SARS-CoV-2 (Mpro) in cell culture and in vitro enzymatic assays, using a methodology that enabled us to focus solely on any potential inhibitory effects of the inhibitors against the viral protease. For cell culture experiments a dark-to-bright GFP reporter substrate system was designed.ResultsLopinavir, ritonavir, darunavir, saquinavir, and atazanavir were able to inhibit the viral protease in cell culture, albeit in concentrations much higher than their achievable plasma levels, given their current drug formulations. While inhibition by lopinavir was attributed to its cytotoxicity, ritonavir was the most effective of the panel, with IC50 of 13.7 µM. None of the inhibitors showed significant inhibition of SARS-CoV-2 Mpro in our in vitro enzymatic assays up to 100 µM concentration. ConclusionTargeting of SARS-CoV-2 Mpro by some of the HIV PIs might be of limited clinical potential, given the high concentration of the drugs required to achieve significant inhibition. Therefore, given their weak inhibition of the viral protease, any potential beneficial effect of the PIs in COVID-19 context might perhaps be attributed to acting on other molecular target(s), rather than SARS-CoV-2 Mpro.

The relationship between HIV evolution and disease progression is fundamental to our understanding of HIV immune control and vaccine design. There are no clear definitions for faster and slower HIV-2 disease progression and for the relationship of the rate of progression with HIV-2 evolution. To address the hypothesis that viral evolution is correlated with disease progression in HIV-2 infection, we determined faster and slower disease progression based on follow-up data from a prospective cohort of police officers in Guinea-Bissau. The analysis showed that although the CD4+ T-cell level and the decline in the level were independently associated with progression to AIDS, only the CD4+ T-cell level or a combined CD4+ T-cell level/decline stratification was associated with the rate of HIV-2 evolution. The HIV-2 evolutionary rate was almost twice as high among the faster progressors as among the slower progressors. Importantly, this report defines previously unknown characteristics linking HIV-2 disease progression with virus evolution.

Disease progression of human immunodeficiency virus type 1 (HIV-1) is delayed by HIV type 2 (HIV-2) in individuals with dual HIV-1/HIV-2 infection. The protective mechanisms, however, are still to be revealed. In the current study we examined type-specific and cross-reactive antibody-dependent cellular cytotoxicity (ADCC) in HIV-1 and HIV-2 monoinfection or dual infection. Of note, intertype cross-reactive antibodies that mediated HIV-1 envelope glycoprotein (Env)–targeted ADCC were frequently identified in HIV-2–infected individuals. Furthermore, the magnitude of HIV-1 cross-reactive ADCC activity during HIV-2 infections depended on the HIV-1 Env origin and was associated with the duration of infection. These results suggest that preexisting antibodies against HIV-2, which mediate intertype ADCC, might contribute to control of HIV-1 during dual infection.

Human immunodeficiency virus type 1 (HIV-1) and HIV-2 share a striking genomic resemblance; however, variability in the genetic sequence accounts for the presence of unique accessory genes, such as the viral protein X () gene in HIV-2. Dual infection with both viruses has long been described in the literature, yet the molecular mechanism of how dually infected patients tend to do better than those who are monoinfected with HIV-1 has not yet been explored. We hypothesized that in addition to extracellular mechanisms, an HIV-2 accessory gene is the culprit, and interference at the viral accessory/regulatory protein level is perhaps responsible for the attenuated pathogenicity of HIV-1 observed in dually infected patients. Following simulation of dual infection in cell culture experiments, we found that pretransduction of cells with HIV-2 significantly protects against HIV-1 transduction. Importantly, we have found that this dampening of the infectivity of HIV-1 was a result of interviral interference carried out by viral protein X of HIV-2, resulting in a severe hindrance to the replication dynamics of HIV-1, influencing both its early and late phases of the viral life cycle. Our findings shed light on potential intracellular interactions between the two viruses and broaden our understanding of the observed clinical spectrum in dually infected patients, highlighting HIV-2 Vpx as a potential candidate worth exploring in the fight against HIV-1. Dual infection with human immunodeficiency virus types 1 and 2 is relatively common in areas of endemicity. For as-yet-unclarified reasons, patients who are dually infected were shown to have lower viral loads and generally a lower rate of progression to AIDS than those who are monoinfected. We aimed to explore dual infection in cell culture, to elucidate possible mechanisms by which HIV-2 may be able to exert such an effect. Our results indicate that on the cellular level, pretransduction of cells with HIV-2 significantly protects against HIV-1 transduction, which was found to be a result of interviral interference carried out by viral protein X of HIV-2. These findings broaden our knowledge of interviral interactions on the cellular level and may provide an explanation for the decreased pathogenicity of HIV-1 in dually infected patients, highlighting HIV-2 Vpx as a potential candidate worth exploring in the fight against HIV.