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While vaccination campaigns are ongoing worldwide, there is still a tremendous medical need for efficient antivirals against SARS-CoV-2 infection. Among several drug candidates, chloroquine (CQN) and hydroxychloroquine (H-CQN) were tested intensively, and any contentious therapeutic effect of both has been discussed controversially in the light of severe side effects and missing efficacy. Originally, H-CQN descended from the natural substance quinine, a medicinal product used since the Middle Ages, which actually is regulatory approved for various indications. We hypothesized that quinine also exerts anti-SARS-CoV-2 activity. In Vero cells, quinine inhibited SARS-CoV-2 infection more effectively than CQN, and H-CQN and was less toxic. In human Caco-2 colon epithelial cells as well as the lung cell line A549 stably expressing ACE2 and TMPRSS2, quinine also showed antiviral activity. In consistence with Vero cells, quinine was less toxic in A549 as compared to CQN and H-CQN. Finally, we confirmed our findings in Calu-3 lung cells, expressing ACE2 and TMPRSS2 endogenously. In Calu-3, infections with high titers of SARS-CoV-2 were completely blocked by quinine, CQN, and H-CQN in concentrations above 50 µM. The estimated IC50s were ~25 µM in Calu-3, while overall, the inhibitors exhibit IC50 values between ~3.7 to ~50 µM, dependent on the cell line and multiplicity of infection (MOI). Conclusively, our data indicate that quinine could have the potential of a treatment option for SARS-CoV-2, as the toxicological and pharmacological profile seems more favorable when compared to its progeny drugs H-CQN or CQN.

Since there is no vaccine or regulatory approved therapy available for treatment of SARS-CoV-2 infection, the medical need to prevent the transition of a mild into the severe COVID-19 stage of infection is of outmost importance. Among several drug candidates, Chloroquine (CQN) and Hydroxy-Chloroquine (H-CQN) have been tested most intensively. However, the therapeutic effect of H-CQN and CQN has been discussed controversially in the light of severe side effects. Originally, H-CQN descended from the natural substance Quinine, a medicinal product used since the Middle Ages and is now regulatory approved for various indications. We hypothesized that Quinine also exerts anti-SARS-CoV-2 activity. First, virus production in Vero B4 cells was analyzed by Western blot, showing that Quinine exerts antiviral activity against SARS-CoV-2 that at 10 µM was even stronger than that of H-CQN or CQN. Second, fluorescence end-point and time lapse analysis of SARS-CoV-2-mNeonGreen-infected Caco-2 cells could confirm a similar antiviral effect of Quinine in a human-derived cell line. Thereby, our in vitro studies revealed, that the antiviral effect appears to be specific, since in Vero cells Quinine impacted cell viability at approximately 50-fold higher concentration, while the therapeutic window of H-CQN and CQN was approximately 10-fold lower. In Caco-2 cells no toxic effect was observed while complete block of infection occurred between 50 and 100 µM at high MOIs. In conclusion, our data indicate that Quinine would have the potential of a well tolerable and widely used treatment option for SARS-CoV-2 infections, with a predictable and significantly better toxicological profile when compared to H-CQN or CQN.

Psoriasis is a frequent systemic inflammatory autoimmune disease characterized primarily by skin lesions with massive infiltration of leukocytes, but frequently also presents with cardiovascular comorbidities. Especially polymorphonuclear neutrophils (PMNs) abundantly infiltrate psoriatic skin but the cues that prompt PMNs to home to the skin are not well-defined. To identify PMN surface receptors that may explain PMN skin homing in psoriasis patients, we screened 332 surface antigens on primary human blood PMNs from healthy donors and psoriasis patients. We identified platelet surface antigens as a defining feature of psoriasis PMNs, due to a significantly increased aggregation of neutrophils and platelets in the blood of psoriasis patients. Similarly, in the imiquimod-induced experimental in vivo mouse model of psoriasis, disease induction promoted PMN-platelet aggregate formation. In psoriasis patients, disease incidence directly correlated with blood platelet counts and platelets were detected in direct contact with PMNs in psoriatic but not healthy skin. Importantly, depletion of circulating platelets in mice in vivo ameliorated disease severity significantly, indicating that both PMNs and platelets may be relevant for psoriasis pathology and disease severity.

Viruses interact with multiple host cell factors. Some of these are required to promote viral propagation, others have roles in inhibiting infection. Here, we delineate the function of the cellular factor PHF13 (or SPOC1), a putative HIV-1 restriction factor. Early in the HIV-1 replication cycle PHF13 increased the number of integrated proviral copies and the number of infected cells. However, after HIV-1 integration, high levels of PHF13 suppressed viral gene expression. The antiviral activity of PHF13 is counteracted by the viral accessory protein Vpr, which mediates PHF13 degradation. Altogether, the transcriptional master regulator and chromatin binding protein PHF13 does not have purely repressive effects on HIV-1 replication, but also promotes viral integration. By the functional characterization of the dual role of PHF13 during the HIV-1 replication cycle, we reveal a surprising and intricate mechanism through which HIV-1 might regulate the switch from integration to viral gene expression. Furthermore, we identify PHF13 as a cellular target specifically degraded by HIV-1 Vpr. ispartof: Open Biology vol:7 issue:10 ispartof: location:England status: published

U2OS cells were treated with 1 μg/ml doxycycline or left untreated for 24 h before cells were infected with 100 ng p24 VSVG pseudotyped HIV-1 NL4-3 IRESeGFP. 24 hpi cells were analyzed by flow cytometry. The mean percentage of GFP+ cells from three independent experiments was calculated and the resulting data was normalized to untreated cells (100 %).

Jurkat-TAg or SupT1 cells were infected with 200 ng p24 VSVG pseudotyped HIV-1 NL4-3 or mock infected. Cells were harvested 48 hpi, total cell extracts were prepared and analyzed for expression of PHF13, HIV-1 capsid p24 and tubulin by immunoblot.

U2OS cells were infected with 100 ng p24 VSVG pseudotyped HIV-1 NL4-3 IRES-eGFP for 24 h. Thereafter, cells were treated with 1 μg/ml doxycycline.Additional 24 h later cells were analyzed by flow cytometry for the % and MFI of GFP+ (HIV-1-infected) cells. The graph shows mean values and standard deviations of three independent experiments.

293T cells were treated with the indicated amounts of recombinant Interferon-α and cultured for 6 h or 24 h. At the time points indicated cells were lysed to detect PHF13, tetherin and actin by immunoblot.

U2OS-C5 cells were mock transfected or with a control siRNA (csiRNA) or directed against PHF13 (siPHF13). 24 h later cells were infected with 100 ng p24 VSVG pseudotyped HIV-1 NL4-3 IRES-eGFP or the ΔVpr variant for 6 h, washed and incubated for additional 18 h. At that time point, cells were induced with 1 μg/ml doxycycline. 24 h later cells were lysed to detect PHF13, tubulin, HIV-1 p24 and Vpr by immunoblot.

The majority of T cells encountered by HIV-1 are non-activated and do not readily allow productive infection. HIV-1 Vpr is highly abundant in progeny virions, and induces signalling and HIV-1 LTR transcription. We hence hypothesized that Vpr might be a determinant of non-activated T-cell infection. Virion-delivered Vpr activated nuclear factor of activated T cells (NFAT) through Ca(2+) influx and interference with the NFAT export kinase GSK3β. This leads to NFAT translocation and accumulation within the nucleus and was required for productive infection of unstimulated primary CD4(+) T cells. A mutagenesis approach revealed correlation of Vpr-mediated NFAT activation with its ability to enhance LTR transcription and mediate cell cycle arrest. Upon NFAT inhibition, Vpr did not augment resting T-cell infection, and showed reduced G2/M arrest and LTR transactivation. Altogether, Vpr renders unstimulated T cells more permissive for productive HIV-1 infection and stimulates activation of productively infected as well as virus-exposed T cells. Therefore, it could be involved in the establishment and reactivation of HIV-1 from viral reservoirs and might have an impact on the levels of immune activation, which are determinants of HIV-1 pathogenesis.