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Authors:CA Stewart et al 

Journal/ Pre-PrintbioRxiv Preprint 

Tags: Bioinformatics, Molecular biology 

Research Highlights

  1. ACE2 and TMPRSS2 are highly expressed in a small number of cells in the respiratory tract, particularly tuft cells (epithelial cells in the respiratory tract characterised by POU2F3 expression)Cells expressing ACE2 are positively correlated with GLUL expression, which encodes glutamine synthetase, thus priming the cells for rapid replication.  

  1. Tumour cell lines can be used more widely in SARS-CoV2 research. In both NSCLC and TCGA tumour samples, ACE2 expression correlates with higher levels of epithelial proteins (E-cadherin, RAB25a and beta-catenin) compared to mesenchymal proteins (such as ZEB1)Computational investigations suggest direct involvement of ZEB1 on ACE2 expression via putative ZEB1 binding sites in the ACE2 promoter.  

  1. NSCLC cell lines (A549 and Calu-3) infected with SARS-CoV2 undergo an EMT-like transcriptional change, characterised by upregulation of ZEB1 and AXL, and downregulation of EPCAM, GLUL and ACE2.  

Summary 

This study observes that in aerodigestive and respiratory cancer cell lines and tumour specimens, that ACE2 expression is positively correlated with the epithelial phenotype and upon mesenchymal transition of these cells, ACE2 is downregulated. Expectedly, ZEB1, a mesenchymal marker, is upregulated and computational analysis suggests ZEB1 directly represses ACE2 expression by binding to its promoter.  

SARS-CoV2 infection of NSCLC cell lines (A549 and Calu-3) resulted in upregulation of ZEB1 and AXL and downregulation of EPCAM, GLUL and ACE2, indicating an EMT-like transcriptional change occurs. The authors suggest that the loss of ACE2 may increase the risk of acute respiratory distress syndrome in COVID-19 patients and that therapeutics targeting ZEB1 or AXL inhibition may be of benefit.  

Impact for SARS-CoV2/COVID19 research efforts  

Clinical symptoms and pathogenesis of SARS-Cov2/COVID19 

  • This paper highlights a novel pathogenesis mechanism of SARS-CoV2 infection. Once SARS-CoV2 has infected epithelial cells expressing ACE2 and TMPRSS2 in the aerodigestive or respiratory tract, the infected cells undergo an EMT-like transcriptional change, characterised by high ZEB1 and AXL expression and decreased ACE2 expression, which may increase the risk of Acute Respiratory Distress Syndrome in COVID-19 patients.  

Study Type  

  • In silico study / bioinformatics study 

  • In vitro study  

Strengths and limitations of the paper 

Novelty:  

  • Describes a novel model of SARS-CoV2 pathogenesis, whereby SARS-CoV2 infects highly epithelial cells in the aerodigestive or respiratory tract, leading to an EMT-like change, characterised by high ZEB1 and AXL expression, with a resultant down-regulation of ACE2 expression.  

  • Highlights that potentially using cancer cell lines and malignant tissues as models for SARS-CoV2 investigation could be beneficial as EMT plays a role in the pathogenesis of the infection and is much better defined in cancer cells than normal tissue.  

Standing in the field: 

  • The authors argue that decreased ACE2 expression may increase ARDS, but the relationship between ACE2 expression, viral infection, and pathogenesis has not been established in the field. 

  • Previous data has suggested that after infection with SARS-COV1, ACE2 expression is downregulated (Kuba, K., et al. A crucial role of angiotensin converting enzyme 2 (ACE2) in SARS coronavirus‐ induced lung injury. Nat Med 11, 875‐879 (2005)). This study confirms that this is also the case in SARS-CoV2 infection.  

  • There are inconsistencies in the literature with regards to the association between ACE2 expression and epithelial differentiation. This study provides evidence that epithelial to mesenchymal phenotype changes are inversely correlated with ACE2 expression.  

Appropriate statistics: 

  • Yes  

Viral model used: 

  • Used data from previous study that infected NSCLC cell lines with SARS-CoV2 (isolate USA-WA1/2020) (Blanco‐Melo, D., et al. Imbalanced Host Response to SARS‐CoV‐2 Drives Development of COVID‐ 19. Cell (2020)).  

Translatability: 

  • This study suggests that EMT occurs in SARS-CoV2-infected cells and highlights ZEB-1 and AXL as therapeutic targets in COVID-19.  

  • Bemcentinib, an AXL kinase inhibitor, is already in an ACCORD UK Phase II clinical trial.  

Main limitations:  

  • Lack of consistency in cell lines used throughout. The authors use multiple cancer cell lines and both normal and tumour samples. It is at times unclear why different samples are being used.  

  • Data is limited to transcriptional changes so protein and functional studies will be needed to assess translatability.  

  • Further studies confirming the direct role of ZEB1 as a repressor of ACE2 expression would be useful, such as exploration of the repressor sites in the ACE2 promoter.