| 1. |
- Haid, Sibylle, et al.
(författare)
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Repurposing screen identifies novel candidates for broad-spectrum coronavirus antivirals and druggable host targets
- 2024
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Ingår i: Antimicrobial Agents and Chemotherapy. - : American Society for Microbiology. - 0066-4804 .- 1098-6596. ; 68:3
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Tidskriftsartikel (refereegranskat)abstract
- Libraries composed of licensed drugs represent a vast repertoire of molecules modulating physiological processes in humans, providing unique opportunities for the discovery of host-targeting antivirals. We screened the Repurposing, Focused Rescue, and Accelerated Medchem (ReFRAME) repurposing library with approximately 12,000 molecules for broad-spectrum coronavirus antivirals and discovered 134 compounds inhibiting an alphacoronavirus and mapping to 58 molecular target categories. Dominant targets included the 5-hydroxytryptamine receptor, the dopamine receptor, and cyclin-dependent kinases. Gene knock-out of the drugs’ host targets including cathepsin B and L (CTSB/L; VBY-825), the aryl hydrocarbon receptor (AHR; Phortress), the farnesyl-diphosphate farnesyltransferase 1 (FDFT1; P-3622), and the kelch-like ECH-associated protein 1 (KEAP1; Omaveloxolone), significantly modulated HCoV-229E infection, providing evidence that these compounds inhibited the virus through acting on their respective host targets. Counter-screening of all 134 primary compound candidates with SARS-CoV-2 and validation in primary cells identified Phortress, an AHR activating ligand, P-3622-targeting FDFT1, and Omaveloxolone, which activates the NFE2-like bZIP transcription factor 2 (NFE2L2) by liberating it from its endogenous inhibitor KEAP1, as antiviral candidates for both an Alpha- and a Betacoronavirus. This study provides an overview of HCoV-229E repurposing candidates and reveals novel potentially druggable viral host dependency factors hijacked by diverse coronaviruses.
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| 2. |
- Ricke-Hoch, Melanie, et al.
(författare)
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Impaired immune response mediated by prostaglandin E2 promotes severe COVID-19 disease
- 2021
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 16:8
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Tidskriftsartikel (refereegranskat)abstract
- The SARS-CoV-2 coronavirus has led to a pandemic with millions of people affected. The present study finds that risk-factors for severe COVID-19 disease courses, i.e. male sex, older age and sedentary life style are associated with higher prostaglandin E2 (PGE2) serum levels in blood samples from unaffected subjects. In COVID-19 patients, PGE2 blood levels are markedly elevated and correlate positively with disease severity. SARS-CoV-2 induces PGE2 generation and secretion in infected lung epithelial cells by upregulating cyclo-oxygenase (COX)-2 and reducing the PG-degrading enzyme 15-hydroxyprostaglan-din-dehydrogenase. Also living human precision cut lung slices (PCLS) infected with SARS-CoV-2 display upregulated COX-2. Regular exercise in aged individuals lowers PGE2 serum levels, which leads to increased Paired-Box-Protein-Pax-5 (PAX5) expression, a master regulator of B-cell survival, proliferation and differentiation also towards long lived memory B-cells, in human pre-B-cell lines. Moreover, PGE2 levels in serum of COVID-19 patients lowers the expression of PAX5 in human pre-B-cell lines. The PGE2 inhibitor Taxifolin reduces SARS-CoV-2-induced PGE2 production. In conclusion, SARS-CoV-2, male sex, old age, and sedentary life style increase PGE2 levels, which may reduce the early anti-viral defense as well as the development of immunity promoting severe disease courses and multiple infections. Regular exercise and Taxifolin treatment may reduce these risks and prevent severe disease courses.
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| 3. |
- Carriquí-Madroñal, Belén, et al.
(författare)
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The matrix metalloproteinase ADAM10 supports hepatitis C virus entry and cell-to-cell spread via its sheddase activity
- 2023
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Ingår i: PLoS Pathogens. - : Public Library of Science (PLoS). - 1553-7366 .- 1553-7374. ; 19:11
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Tidskriftsartikel (refereegranskat)abstract
- Hepatitis C virus (HCV) exploits the four entry factors CD81, scavenger receptor class B type I (SR-BI, also known as SCARB1), occludin, and claudin-1 as well as the co-factor epidermal growth factor receptor (EGFR) to infect human hepatocytes. Here, we report that the disintegrin and matrix metalloproteinase 10 (ADAM10) associates with CD81, SR-BI, and EGFR and acts as HCV host factor. Pharmacological inhibition, siRNA-mediated silencing and genetic ablation of ADAM10 reduced HCV infection. ADAM10 was dispensable for HCV replication but supported HCV entry and cell-to-cell spread. Substrates of the ADAM10 sheddase including epidermal growth factor (EGF) and E-cadherin, which activate EGFR family members, rescued HCV infection of ADAM10 knockout cells. ADAM10 did not influence infection with other enveloped RNA viruses such as alphaviruses and a common cold coronavirus. Collectively, our study reveals a critical role for the sheddase ADAM10 as a HCV host factor, contributing to EGFR family member transactivation and as a consequence to HCV uptake.
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| 4. |
- Lasswitz, Lisa, et al.
(författare)
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The tetraspanin CD81 is a host factor for Chikungunya virus replication
- 2022
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Ingår i: mBio. - : ASM International. - 2161-2129 .- 2150-7511. ; 13:3
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Tidskriftsartikel (refereegranskat)abstract
- Chikungunya virus (CHIKV) is an arthritogenic reemerging virus replicating in plasma membrane-derived compartments termed "spherules." Here, we identify the human transmembrane protein CD81 as host factor required for CHIKV replication. Ablation of CD81 results in decreased CHIKV permissiveness, while overexpression enhances infection. CD81 is dispensable for virus uptake but critically required for viral genome replication. Likewise, murine CD81 is crucial for CHIKV permissiveness and is expressed in target cells such as dermal fibroblasts, muscle and liver cells. Whereas related alphaviruses, including Ross River virus (RRV), Semliki Forest virus (SFV), Sindbis virus (SINV) and Venezuelan equine encephalitis virus (VEEV), also depend on CD81 for infection, RNA viruses from other families, such as coronaviruses, replicate independently of CD81. Strikingly, the replication-enhancing function of CD81 is linked to cholesterol binding. These results define a mechanism exploited by alphaviruses to hijack the membrane microdomain-modeling protein CD81 for virus replication through interaction with cholesterol.IMPORTANCE: In this study, we discover the tetraspanin CD81 as a host factor for the globally emerging chikungunya virus and related alphaviruses. We show that CD81 promotes replication of viral genomes in human and mouse cells, while virus entry into cells is independent of CD81. This provides novel insights into how alphaviruses hijack host proteins to complete their life cycle. Alphaviruses replicate at distinct sites of the plasma membrane, which are enriched in cholesterol. We found that the cholesterol-binding ability of CD81 is important for its function as an alphavirus host factor. This discovery thus broadens our understanding of the alphavirus replication process and the use of host factors to reprogram cells into virus replication factories.
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| 5. |
- Mhlekude, Baxolele, et al.
(författare)
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Pharmacological inhibition of bromodomain and extra-terminal proteins induces an NRF-2-mediated antiviral state that is subverted by SARS-CoV-2 infection
- 2023
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Ingår i: PLoS Pathogens. - : Public Library of Science (PLoS). - 1553-7366 .- 1553-7374. ; 19:9
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Tidskriftsartikel (refereegranskat)abstract
- Inhibitors of bromodomain and extra-terminal proteins (iBETs), including JQ-1, have been suggested as potential prophylactics against SARS-CoV-2 infection. However, molecular mechanisms underlying JQ-1-mediated antiviral activity and its susceptibility to viral subversion remain incompletely understood. Pretreatment of cells with iBETs inhibited infection by SARS-CoV-2 variants and SARS-CoV, but not MERS-CoV. The antiviral activity manifested itself by reduced reporter expression of recombinant viruses, and reduced viral RNA quantities and infectious titers in the culture supernatant. While we confirmed JQ-1-mediated downregulation of expression of angiotensin-converting enzyme 2 (ACE2) and interferon-stimulated genes (ISGs), multi-omics analysis addressing the chromatin accessibility, transcriptome and proteome uncovered induction of an antiviral nuclear factor erythroid 2-related factor 2 (NRF-2)-mediated cytoprotective response as an additional mechanism through which JQ-1 inhibits SARS-CoV-2 replication. Pharmacological inhibition of NRF-2, and knockdown of NRF-2 and its target genes reduced JQ-1-mediated inhibition of SARS-CoV-2 replication. Serial passaging of SARS-CoV-2 in the presence of JQ-1 resulted in predominance of ORF6-deficient variant, which exhibited resistance to JQ-1 and increased sensitivity to exogenously administered type I interferon (IFN-I), suggesting a minimised need for SARS-CoV-2 ORF6-mediated repression of IFN signalling in the presence of JQ-1. Importantly, JQ-1 exhibited a transient antiviral activity when administered prophylactically in human airway bronchial epithelial cells (hBAECs), which was gradually subverted by SARS-CoV-2, and no antiviral activity when administered therapeutically following an established infection. We propose that JQ-1 exerts pleiotropic effects that collectively induce an antiviral state in the host, which is ultimately nullified by SARS-CoV-2 infection, raising questions about the clinical suitability of the iBETs in the context of COVID-19.
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| 6. |
- Passos, Vania, et al.
(författare)
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Innate immune response to SARS-CoV-2 infection contributes to neuronal damage in human iPSC-derived peripheral neurons
- 2024
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Ingår i: Journal of Medical Virology. - : John Wiley & Sons. - 0146-6615 .- 1096-9071. ; 96:2
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Tidskriftsartikel (refereegranskat)abstract
- Severe acute respiratory coronavirus 2 (SARS-CoV-2) causes neurological disease in the peripheral and central nervous system (PNS and CNS, respectively) of some patients. It is not clear whether SARS-CoV-2 infection or the subsequent immune response are the key factors that cause neurological disease. Here, we addressed this question by infecting human induced pluripotent stem cell-derived CNS and PNS neurons with SARS-CoV-2. SARS-CoV-2 infected a low number of CNS neurons and did not elicit a robust innate immune response. On the contrary, SARS-CoV-2 infected a higher number of PNS neurons. This resulted in expression of interferon (IFN) λ1, several IFN-stimulated genes and proinflammatory cytokines. The PNS neurons also displayed alterations characteristic of neuronal damage, as increased levels of sterile alpha and Toll/interleukin receptor motif-containing protein 1, amyloid precursor protein and α-synuclein, and lower levels of cytoskeletal proteins. Interestingly, blockade of the Janus kinase and signal transducer and activator of transcription pathway by Ruxolitinib did not increase SARS-CoV-2 infection, but reduced neuronal damage, suggesting that an exacerbated neuronal innate immune response contributes to pathogenesis in the PNS. Our results provide a basis to study coronavirus disease 2019 (COVID-19) related neuronal pathology and to test future preventive or therapeutic strategies.
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| 7. |
- Zapatero-Belinchon, Francisco J., et al.
(författare)
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Characterization of the Filovirus-Resistant Cell Line SH-SY5Y Reveals Redundant Role of Cell Surface Entry Factors
- 2019
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Ingår i: Viruses. - : MDPI. - 1999-4915. ; 11:3
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Tidskriftsartikel (refereegranskat)abstract
- Filoviruses infect a wide range of cell types with the exception of lymphocytes. The intracellular proteins cathepsin B and L, two-pore channel 1 and 2, and bona fide receptor Niemann-Pick Disease C1 (NPC1) are essential for the endosomal phase of cell entry. However, earlier steps of filoviral infection remain poorly characterized. Numerous plasma membrane proteins have been implicated in attachment but it is still unclear which ones are sufficient for productive entry. To define a minimal set of host factors required for filoviral glycoprotein-driven cell entry, we screened twelve cell lines and identified the nonlymphocytic cell line SH-SY5Y to be specifically resistant to filovirus infection. Heterokaryons of SH-SY5Y cells fused to susceptible cells were susceptible to filoviruses, indicating that SH-SY5Y cells do not express a restriction factor but lack an enabling factor critical for filovirus entry. However, all tested cell lines expressed functional intracellular factors. Global gene expression profiling of known cell surface entry factors and protein expression levels of analyzed attachment factors did not reveal any correlation between susceptibility and expression of a specific host factor. Using binding assays with recombinant filovirus glycoprotein, we identified cell attachment as the step impaired in filovirus entry in SH-SY5Y cells. Individual overexpression of attachment factors T-cell immunoglobulin and mucin domain 1 (TIM-1), Axl, Mer, or dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) rendered SH-SY5Y cells susceptible to filovirus glycoprotein-driven transduction. Our study reveals that a lack of attachment factors limits filovirus entry and provides direct experimental support for a model of filoviral cell attachment where host factor usage at the cell surface is highly promiscuous.
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| 8. |
- Zapatero-Belinchón, Francisco J., et al.
(författare)
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Fluvastatin mitigates SARS-CoV-2 infection in human lung cells
- 2021
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Ingår i: iScience. - : Elsevier. - 2589-0042. ; 24:12
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Tidskriftsartikel (refereegranskat)abstract
- Clinical data of patients suffering from COVID-19 indicates that statin therapy, used to treat hypercholesterolemia, is associated with a better disease outcome. Whether statins directly affect virus replication or influence the clinical outcome through modulation of immune responses is unknown. We therefore investigated the effect of statins on SARS-CoV-2 infection in human lung cells and found that only fluvastatin inhibited low and high pathogenic coronaviruses in vitro and ex vivo in a dose-dependent manner. Quantitative proteomics revealed that fluvastatin and other tested statins modulated the cholesterol synthesis pathway without altering innate antiviral immune responses in infected lung epithelial cells. However, fluvastatin treatment specifically downregulated proteins that modulate protein translation and viral replication. Collectively, these results support the notion that statin therapy poses no additional risk to individuals exposed to SARS-CoV-2 and that fluvastatin has a moderate beneficial effect on SARS-CoV-2 infection of human lung cells.
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| 9. |
- Zapatero-Belinchón, Francisco J., et al.
(författare)
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Interdependent Impact of Lipoprotein Receptors and Lipid-Lowering Drugs on HCV Infectivity
- 2021
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Ingår i: Cells. - : MDPI. - 2073-4409. ; 10:7
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Tidskriftsartikel (refereegranskat)abstract
- The HCV replication cycle is tightly associated with host lipid metabolism: Lipoprotein receptors SR-B1 and LDLr promote entry of HCV, replication is associated with the formation of lipid-rich membranous organelles and infectious particle assembly highjacks the very‑low-density lipoprotein (VLDL) secretory pathway. Hence, medications that interfere with the lipid metabolism of the cell, such as statins, may affect HCV infection. Here, we study the interplay between lipoprotein receptors, lipid homeostasis, and HCV infection by genetic and pharmacological interventions. We found that individual ablation of the lipoprotein receptors SR‑B1 and LDLr did not drastically affect HCV entry, replication, or infection, but double lipoprotein receptor knock-outs significantly reduced HCV infection. Furthermore, we could show that this effect was neither due to altered expression of additional HCV entry factors nor caused by changes in cellular cholesterol content. Strikingly, whereas lipid‑lowering drugs such as simvastatin or fenofibrate did not affect HCV entry or infection of immortalized hepatoma cells expressing SR-B1 and/or LDLr or primary human hepatocytes, ablation of these receptors rendered cells more susceptible to these drugs. Finally, we observed no significant differences between statin users and control groups with regards to HCV viral load in a cohort of HCV infected patients before and during HCV antiviral treatment. Interestingly, statin treatment, which blocks the mevalonate pathway leading to decreased cholesterol levels, was associated with mild but appreciable lower levels of liver damage markers before HCV therapy. Overall, our findings confirm the role of lipid homeostasis in HCV infection and highlight the importance of the mevalonate pathway in the HCV replication cycle.
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