SARS-CoV-2 cell entry depends on ACE2 and TMPRSS2 and is blocked by a clinically proven protease inhibitor
Authors: M. Hoffman, H. Kleine-Weber, S. Schroeder, N. Krüger, S. Erichsen, Schiergens, T.S., et al.
Journal: Cell, 181, 1-10, 25 Feb 2020.
SARS-CoV-2 uses its viral spike (S) glycoprotein for host cell receptor binding and requires its proteolytic cleavage (priming) for viral fusion and successful host cell entry.
These authors are one of the first to demonstrate that SARS-CoV-2 uses the same ACE-2 receptor for entry and the same serine TMPRSS2protease for S protein priming, as SARS-CoV utilizes.
A TMPRSS2 inhibitor approved for clinical use in Japan reduced viral entry of clinical isolate Munich 929 SARS-CoV-2 in a human lung cell line and SARS-CoV-2 S-pseudotyped particles in primary lung cells.
Sera from convalescent SARS-patients efficiently inhibits SARS-CoV and partially cross-neutralizes SARS-COV-2 pseudotype viral entry in the susceptible Vero cell line.
- SARS-CoV-2 uses the SARS-CoV receptor ACE2 for host cell entry
- The spike protein of SARS-CoV-2 is primed by TMPRSS2
- Antibodies against SARS-CoV S may offer some protection against SARS-CoV-2
For inhibition of COVID19 transmission (✔)
- Some evidence that sera from convalescent SARS-CoV patients has a partially protective effect against a SARS-CoV-2 S pseudotype in a monkey cell line.
For treatment of SARS-COV-2 positive individuals (✔)
- Identify two molecules important in the host cell entry, ACE2 receptor and serine protease TMPRSS2, that could be targeted therapeutically to reduce host cell infection (particularly as TMPRSS2 has previously been shown to be dispensable for development and homeostasis (Kim et al., 2006, Mol Cell Biol.)). Moreover, show that a drug currently in clinical use (serine protease inhibitor camostat mesylate) reduced entry of virus in cell lines and in primary human lung cells.
Amino acid sequence analysis to identify sequence similarity of receptor binding domain in SARS-CoV-2 and other coronaviruses.
The commonalities between SARS-CoV and SARS-CoV-2 viral entry were probed mainly using VSV particles carrying GFP and luciferase and pseudotyped with respective S glycoproteins in combination with various cell lines. Key experiments are also performed with clinical isolate SARS-CoV-2 (Munich 929) and on primary human airway epithelial cells.
SARS-CoV-2 isolate Munich 929 was propagated in VeroE6 cells (passage 1) after primary isolation from patient material on Vero-TMPRSS2 cells (passage 0).
Convalescent human anti-SARS-CoV sera was obtained from the serum collection of the national consiliary laboratory for coronavirus diagnostics at Charité, Berlin, Germany or the Robert Koch Institute, Berlin, Germany. All sera were previously tested positive using a recombinant S-based immunofluorescence test.
Strengths and weaknesses of the paper
- Second paper implicating ACE2 as receptor.
- First to identify TMPRSS2 (and to lesser extent cathepsin B/L) as being involved in S glycoprotein priming in SARS-CoV-2 specifically.
- First in vitro testing of camostat in context of SARS-CoV-2.
- First to look at sera of SARS-CoV convalescent patients in cross-neutralization of SARS-CoV-2.
- Replicate findings in paper published month before (ACE2 data – Zhou et al., 2020).
- Use mean of three experiments across key experiments
- Use both model SARS-2-S-based VSV pseudotype and “authentic” SARS-CoV-2. Also use cell lines and primary human lung cells.
- Findings easily translated – use drug that has already been used in a clinical setting
- However, low efficacy of cross-neutralizing anti-SARS-CoV antibodies for treating SARS-CoV-2 infection
- Some controls not shown, which would have been helpful for interpretation of TMPRSS2 data, particularly in determining the relative contributions of TMPRSS2 and cathepsin B/L to S protein priming (e.g. figure S3 – do not show pseudotype entry in untreated cells)
- Obvious limitation in that it is only in vitro experiments → next step would be to look at ACE2 and TMPRSS2, and any targeting of these proteins, in animal models.