ABSTRACT
Background: Despite recent advances, the management of COVID19 is complicated by vaccine availability, the modest efficacy of existing treatments, and the potential for viral resistance. Therefore, there is a pressing need for new prophylactic and therapeutic agents. Modifying the expression of the SARS-CoV-2 entry receptor ACE2 could prevent viral infection and limit disease progression. Here, we identify that ACE2 expression is controlled by the transcription factor farnesoid X receptor (FXR) and demonstrate that ACE2 downregulation through FXR antagonism, using approved drugs, such as ursodeoxycholic acid (UDCA), could represent a novel therapeutic strategy to complement current approaches. Methods: Primary cholangiocyte, pulmonary and intestinal organoids were propagated using established protocols. Marker expression was assessed using singlecell RNA sequencing, QPCR, immunofluorescence and flow cytometry. FXR binding on DNA was assessed with chromatin immunoprecipitation. SARS-CoV-2 was isolated from bronchoalveolar lavage of a COVID19 patient. Viral load was measured via QPCR. Human livers not used for transplantation were perfused ex-situ using the metra (OrganOx) normothermic perfusion device. Serum ACE2 activity was measured with commercial kits. Patient data from the COVID-Hep and SECURE-Liver registries were compared using propensity score matching. Results: FXR activation directly upregulated ACE2 transcription in organoids from COVID19 affected tissues, including the biliary, gastrointestinal and respiratory systems. Conversely, FXR antagonism with z-guggulsterone or UDCA, had the opposite effect. Importantly, both drugs reduced susceptibility to SARS-CoV-2 infection in lung, cholangiocyte and gut organoids. Furthermore, systemic administration of UDCA in human organs perfused ex-situ downregulated ACE2 and reduced SARS-CoV-2 infection ex-vivo. Oral UDCA rapidly reduced serum ACE2 in vivo. Registry data showed a correlation between UDCA administration and better clinical outcomes in COVID19 patients, including hospitalisation, ICU admission, mechanical ventilation and death. Conclusion: We discovered FXR as a novel therapeutic target against SARS-CoV-2 and we identified approved FXR inhibitors which could be repurposed to potentially treat COVID19, paving the road for future clinical trials to validate these results.
ABSTRACT
Introduction The management of COVID19 is complicated by vaccine availability, the modest efficacy of existing treatments, and the potential for viral resistance. Therefore, there is a pressing need for new prophylactic and therapeutic agents. The viral receptor ACE2 is an ideal target as it is required for SARS-CoV-2 entry in host cells. Modifying ACE2 expression could prevent infection and/or limit disease progression. Nevertheless, the mechanisms controlling ACE2 expression remain elusive. Aims To identify pathways controlling the transcriptional regulation of ACE2, and exploit them to reduce SARS-CoV-2 infection. Methods Organoids from primary biliary, intestinal and pulmonary epithelia were derived and cultured as previously described. Single-cell RNA sequencing, QPCR, immunofluorescence and flow cytometry were used to assess marker expression. Chromatin immunoprecipitation was used to assess FXR binding on DNA. Bronchoalveolar lavage SARS-CoV-2 patient isolates were used for infection experiments. Human livers not used for transplantation were connected to the metra (OrganOx) normothermic perfusion device and perfused ex-situ using therapeutic doses of UDCA for 12 hours. ACE2 activity was measured following manufacturer's instructions. Patient data from the COVID-Hep and SECURE-Liver registries were compared using propensity score matching for sex, age and Child-Turcotte-Pugh score. Results We first demonstrated that cholangiocytes are susceptible to SARS-CoV-2 infection in vivo and in organoid culture. We then used cholangiocyte organoids to identify FXR as a transcriptional regulator of ACE2. We validated our results in pulmonary and intestinal organoids, showing that ACE2 regulation by FXR represents a broad mechanism present in multiple COVID19-affected tissues. We then demonstrated that approved FXR inhibitors, such as ursodeoxycholic acid (UDCA) and z-guggulsterone (ZGG), decrease ACE2 levels and reduce viral infection in vitro in primary biliary, intestinal and pulmonary organoids. We interrogated the impact of systemic UDCA administration in human livers perfused ex-situ, demonstrating reduced ACE2 levels and SARS-CoV-2 infection. Furthermore, we showed that commencing UDCA treatment lowers ACE2 levels in primary biliary cholangitis (PBC) patients. Finally, we identified a correlation between UDCA treatment and better clinical outcome in COVID-19 patients, including hospitalisation, ICU admission, mechanical ventilation and death, using registry data. Conclusion We identified FXR as a novel master regulator of ACE2 expression. Using a bench-to-bedside approach we combined in vitro, ex-vivo and patient data to demonstrate the efficacy of ACE2 downregulation against SARS-CoV-2 infection and identified approved and inexpensive drugs (UDCA, ZGG) which could be repurposed as prophylactic and therapeutic agents against SARS-CoV-2 infection, paving the road for future clinical trials.