Evaluation of SARS-CoV-2 entry, inflammation and new therapeutics in human lung tissue cells.

The development of physiological models that reproduce SARS-CoV-2 infection in primary human cells will be instrumental to identify host-pathogen interactions and potential therapeutics. Here, using cell suspensions directly from primary human lung tissues (HLT), we have developed a rapid platform for the identification of viral targets and the expression of viral entry factors, as well as for the screening of viral entry inhibitors and anti-inflammatory compounds. The direct use of HLT cells, without long-term cell culture and in vitro differentiation approaches, preserves main immune and structural cell populations, including the most susceptible cell targets for SARS-CoV-2; alveolar type II (AT-II) cells, while maintaining the expression of proteins involved in viral infection, such as ACE2, TMPRSS2, CD147 and AXL. Further, antiviral testing of 39 drug candidates reveals a highly reproducible method, suitable for different SARS-CoV-2 variants, and provides the identification of new compounds missed by conventional systems, such as VeroE6. Using this method, we also show that interferons do not modulate ACE2 expression, and that stimulation of local inflammatory responses can be modulated by different compounds with antiviral activity. Overall, we present a relevant and rapid method for the study of SARS-CoV-2.

Entrectinib-A SARS-CoV-2 Inhibitor in Human Lung Tissue (HLT) Cells.

Since the start of the COVID-19 outbreak, pharmaceutical companies and research groups have focused on the development of vaccines and antiviral drugs against SARS-CoV-2. Here, we apply a drug repurposing strategy to identify drug candidates that are able to block the entrance of the virus into human cells. By combining virtual screening with in vitro pseudovirus assays and antiviral assays in Human Lung Tissue (HLT) cells, we identify entrectinib as a potential antiviral drug.

Treatment intensification with raltegravir in subjects with sustained HIV-1 viraemia suppression: a randomized 48-week study.

BACKGROUND: Residual viraemia is a major obstacle to HIV-1 eradication in subjects receiving HAART. The intensification with raltegravir could impact latent reservoirs and might lead to a reduction of plasma HIV-1 viraemia (viral load [VL]), complementary DNA intermediates and immune activation. METHODS: This was a prospective, open-label, randomized study comprising 69 individuals on suppressive HAART randomly assigned 2:1 to add raltegravir during 48 weeks. RESULTS: Total and integrated HIV-1 DNA, and ultrasensitive VL remained stable despite intensification. There was a significant increase in episomal HIV DNA at weeks 2-4 in the raltegravir group returning to baseline levels at week 48. Median CD4(+) T-cell counts increased 124 and 80 cells/µl in the intensified and control groups after 48 weeks (P=0.005 and P=0.027, respectively), without significant differences between groups. No major changes were observed in activation of CD4(+) T-cells. Conversely, raltegravir intensification significantly reduced activation of CD8(+) T-cells at week 48 (HLA-DR(+)CD38(+), P=0.005), especially in the memory compartment (CD38(+) of CD8(+)CD45RO(+), P<0.0001). Linear mix models also depicted a larger decrease in CD8(+) T-cell activation in the intensification group (P=0.036 and P=0.010, respectively). Raltegravir intensification was not associated to any particular adverse event. CONCLUSIONS: Intensification of HAART with raltegravir during 48 weeks was safe and associated with a significant decrease in CD8(+) T-cell activation, and a transient increase of episomal HIV-1 DNA. However, raltegravir did not significantly contribute to changes in CD4(+) T-cell counts, ultrasensitive VL, and total and integrated HIV-1 DNA. These findings suggest that raltegravir impacts residual HIV-1 replication and support new strategies to impair HIV-1 persistence. ClinicalTrials.gov identifier: NCT00554398.

HIV-1 replication and immune dynamics are affected by raltegravir intensification of HAART-suppressed subjects.

Highly active antiretroviral therapy (HAART) results in potent and durable suppression of HIV-1 viremia. However, HIV-1 replication resumes if therapy is interrupted. Although it is generally believed that active replication has been halted in individuals on HAART, immune activation and inflammation continue at abnormal levels, suggesting continued, low-level viral replication. To assess whether active replication might be driving immune activation in HAART, we examined the impact of treatment intensification with the integrase inhibitor raltegravir on viral complementary DNA and immune activation parameters. In the presence of raltegravir, linear HIV-1 cDNA is prevented from integrating into chromatin and is subsequently converted to episomal cDNAs. Raltegravir intensification of a three-drug suppressive HAART regimen resulted in a specific and transient increase in episomal DNAs in a large percentage of HAART-suppressed subjects. Furthermore, in subjects with these episomal DNAs, immune activation was higher at baseline and was subsequently normalized after raltegravir intensification. These results suggest that, despite suppressive HAART, active replication persists in some infected individuals and drives immune activation. The ability of raltegravir intensification to perturb the reservoir that supports active replication has implications for therapeutic strategies aimed at achieving viral eradication.