IgM anti-ACE2 autoantibodies in severe COVID-19 activate complement and perturb vascular endothelial function
Livia Casciola-Rosen, David R. Thiemann, Felipe Andrade, Maria I. Trejo-Zambrano, Elissa K. Leonard, Jamie B. Spangler, Nicole E. Skinner, Justin Bailey, Srinivasan Yegnasubramanian, Rulin Wang, Ajay M. Vaghasia, Anuj Gupta, Andrea L. Cox, Stuart C. Ray, Raleigh M. Linville, Zhaobin Guo, Peter C. Searson, Carolyn E. Machamer, Stephen Desiderio, Lauren M. Sauer, Oliver Laeyendecker, Brian T. Garibaldi, Li Gao, Mahendra Damarla, Paul M. Hassoun, Jody E. Hooper, Christopher A. Mecoli, Lisa Christopher-Stine, Laura Gutierrez-Alamillo, Qingyuan Yang, David Hines, William A. Clarke, Richard E. Rothman, Andrew Pekosz, Katherine Z.J. Fenstermacher, Zitong Wang, Scott L. Zeger, Antony Rosen
Livia Casciola-Rosen, David R. Thiemann, Felipe Andrade, Maria I. Trejo-Zambrano, Elissa K. Leonard, Jamie B. Spangler, Nicole E. Skinner, Justin Bailey, Srinivasan Yegnasubramanian, Rulin Wang, Ajay M. Vaghasia, Anuj Gupta, Andrea L. Cox, Stuart C. Ray, Raleigh M. Linville, Zhaobin Guo, Peter C. Searson, Carolyn E. Machamer, Stephen Desiderio, Lauren M. Sauer, Oliver Laeyendecker, Brian T. Garibaldi, Li Gao, Mahendra Damarla, Paul M. Hassoun, Jody E. Hooper, Christopher A. Mecoli, Lisa Christopher-Stine, Laura Gutierrez-Alamillo, Qingyuan Yang, David Hines, William A. Clarke, Richard E. Rothman, Andrew Pekosz, Katherine Z.J. Fenstermacher, Zitong Wang, Scott L. Zeger, Antony Rosen
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Clinical Research and Public Health COVID-19

IgM anti-ACE2 autoantibodies in severe COVID-19 activate complement and perturb vascular endothelial function

Abstract

Background Some clinical features of severe COVID-19 represent blood vessel damage induced by activation of host immune responses initiated by the coronavirus SARS-CoV-2. We hypothesized autoantibodies against angiotensin-converting enzyme 2 (ACE2), the SARS-CoV-2 receptor expressed on vascular endothelium, are generated during COVID-19 and are of mechanistic importance.Methods In an opportunity sample of 118 COVID-19 inpatients, autoantibodies recognizing ACE2 were detected by ELISA. Binding properties of anti-ACE2 IgM were analyzed via biolayer interferometry. Effects of anti-ACE2 IgM on complement activation and endothelial function were demonstrated in a tissue-engineered pulmonary microvessel model.Results Anti-ACE2 IgM (not IgG) autoantibodies were associated with severe COVID-19 and found in 18/66 (27.2%) patients with severe disease compared with 2/52 (3.8%) of patients with moderate disease (OR 9.38, 95% CI 2.38–42.0; P = 0.0009). Anti-ACE2 IgM autoantibodies were rare (2/50) in non-COVID-19 ventilated patients with acute respiratory distress syndrome. Unexpectedly, ACE2-reactive IgM autoantibodies in COVID-19 did not undergo class-switching to IgG and had apparent KD values of 5.6–21.7 nM, indicating they are T cell independent. Anti-ACE2 IgMs activated complement and initiated complement-binding and functional changes in endothelial cells in microvessels, suggesting they contribute to the angiocentric pathology of COVID-19.Conclusion We identify anti-ACE2 IgM as a mechanism-based biomarker strongly associated with severe clinical outcomes in SARS-CoV-2 infection, which has therapeutic implications.FUNDING Bill & Melinda Gates Foundation, Gates Philanthropy Partners, Donald B. and Dorothy L. Stabler Foundation, and Jerome L. Greene Foundation; NIH R01 AR073208, R01 AR069569, Institutional Research and Academic Career Development Award (5K12GM123914-03), National Heart, Lung, and Blood Institute R21HL145216, and Division of Intramural Research, National Institute of Allergy and Infectious Diseases; National Science Foundation Graduate Research Fellowship (DGE1746891)

Authors

Livia Casciola-Rosen, David R. Thiemann, Felipe Andrade, Maria I. Trejo-Zambrano, Elissa K. Leonard, Jamie B. Spangler, Nicole E. Skinner, Justin Bailey, Srinivasan Yegnasubramanian, Rulin Wang, Ajay M. Vaghasia, Anuj Gupta, Andrea L. Cox, Stuart C. Ray, Raleigh M. Linville, Zhaobin Guo, Peter C. Searson, Carolyn E. Machamer, Stephen Desiderio, Lauren M. Sauer, Oliver Laeyendecker, Brian T. Garibaldi, Li Gao, Mahendra Damarla, Paul M. Hassoun, Jody E. Hooper, Christopher A. Mecoli, Lisa Christopher-Stine, Laura Gutierrez-Alamillo, Qingyuan Yang, David Hines, William A. Clarke, Richard E. Rothman, Andrew Pekosz, Katherine Z.J. Fenstermacher, Zitong Wang, Scott L. Zeger, Antony Rosen

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Figure 5

Features of anti-ACE2 IgM antibodies.

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Features of anti-ACE2 IgM antibodies. (A) Anti-ACE2 IgM antibodies from ...
(A) Anti-ACE2 IgM antibodies from patient CV-1 or control do not inhibit ACE2 activity. Positive and negative controls were ACE2 alone, and ACE2 plus ACE2 inhibitor, respectively (see Supplemental Figure 6A). (B) IgM antibodies to ACE2 activate complement. Purified IgMs from anti-ACE2 IgM–positive COVID-19 patients (n = 8) and healthy controls (n = 11) was used for C1q binding assays. Values are means from 2 independent experiments performed on different days. ***P < 0.0001, Mann-Whitney test. (C–G) Anti-ACE2 IgM affects the pulmonary endothelium. (C) Phase image of a pulmonary microvessel (top). Fluorescence images of microvessels exposed to anti-ACE2–negative IgM (HC) or anti-ACE2–positive IgM (CV) after perfusion with 10 kDa dextran (lower). Representative images across n = 3 to 6 independent experiments for each IgM condition are shown. (D) ACE2 and CD31 microvessel staining following 24-hour perfusion with IFN-α/γ. Representative images across n = 3 independent experiments are shown. (E) C3c staining after perfusion with IFN and anti-ACE2–positive or control IgM (3.33 μg/mL). Representative images across n = 3 independent experiments are shown. (F) Permeability of microvessels perfused with IFN and anti-ACE2–positive IgM (CV) or anti-ACE2 negative IgM (HC) (100 μg/mL). D.L., detection limit. A linear mixed effects model was used to test the effect of anti-ACE2–positive IgM on permeability. n = 3 to 5 for each IgM condition; each dot represents an independent replicate. (G) Inhibition of microvessel permeability to 10 kDa dextran in response to anti-ACE2 IgM is IFN dependent. Low CV-1: 3.33 μg/mL; affinity-purified anti-ACE2 IgM: 100 ng/mL. To compare each condition to control, a Kruskal-Wallis test followed by Dunn’s multiple-comparison test was performed. n = 4 to 6 for each IgM condition; each dot represents an independent replicate.