In the editor:
Mutations in the coronavirus 2 ear protein of severe acute respiratory syndrome (SARS-CoV-2) that cause the escape of neutralizing antibodies may occur in immunocompromised patients with prolonged infection.1,2 It is hypothesized that this viral escape contributes to the emergence of global variants. of concern.3 In the absence of effective immune responses, selective pressures such as monoclonal antibody treatment may lead to the appearance of immunologically important mutations.
To understand the selective pressures driving the SARS-CoV-2 evolution of the host, we examined the relationship between this evolution and endogenous immune responses and exogenous antibody treatment in convenience samples obtained from five patients with cell deficiencies. B. (Details of each patient’s medical history are provided in the supplementary appendix, available with the full text of this letter at NEJM.org.) All patients had SARS-CoV-2 infections ranging from last between 42 and 302 days after a first positive test (day 0) (Fig. S1 and table S1 in the supplementary appendix). The study was approved by the Institutional Review Committee of Emory University. Informed consent was obtained from patients who donated whole blood samples for research (patients 2, 4, and 5).
Figure 1. Figure 1. Neutralizing antibody titers, effector T cell responses, and ear mutations in five immunocompromised patients.
Panel A shows the neutralizing antibody titers in the patient’s serum against Wuhan-Hu-1, the reference SARS-CoV-2 pseudovirus, at various times after infection. These titers represent the reciprocal serum dilution at which pseudovirus neutralization was observed at the maximum half. The data show the geometric means of two to five independent experiments; Bars 𝙸 indicate standard deviations. The dotted line represents the lower limit of detection. Panels B and C show remaining background frequencies of CD4 + or CD8 + T cells expressing CD154, interferon-γ, tumor necrosis factor (TNF), or interleukin-2 as a percentage of non-naive cells (i.e. , effectors or memory) in response. to the stimulation of peripheral blood mononuclear cells with a megapool of peptides containing 15-mer of the open spike reading frame (ORF) and a megapool of peptides containing CD8 + T cell epitopes predicted d ‘ORF included, respectively. Frequencies were determined by flow cytometry in patients 4 and 5, as well as in a healthy control donor (HC2) and two patients of the same age hospitalized with Covid-19 (Covid 1 and 2). Panel D shows mutations in the gene encoding the SARS-CoV-2 ear protein compared to the Wuhan-Hu-1 strain, depending on the patient’s identifier and time point. Shading denotes the frequency of mutation. For each mutation, the observed nucleotide variant is listed above the graph and the amino acid mutation is listed below the graph.
Patient 1 received no antibody treatment and was negative for neutralizing antibodies on day 37. Patients 2 and 3 were treated with the monoclonal antibody bamlanivimab on days 4 and 8, respectively. Its serum potently neutralized the reference pseudovirus (Wuhan-Hu-1) on day 33 (patient 2) and day 55 (patient 3) and retained high titers of neutralizing antibodies during days 77 and 83, respectively (Figure 1A). Patient 4 received convalescent plasma on days 0 and 104 and had undetectable neutralizing antibodies on days 82 and 101. Patient 5 received convalescent plasma on day 200 and had low neutralizing antibody titers on day 204. ‘IgG in the ear protein reflected serum neutralization. titles (Fig. S2). All but one patient (patient 2) eventually recovered. Patients 2, 4, and 5 provided peripheral blood samples for the immunophenotype. All three patients had a low lymphocyte count and low to undetectable CD19 + B cell frequencies (0.19% in patient 2, 0.01% in patient 4, and 0.01% in patient 5) compared with healthy controls. and hospitalized by age. patients with coronavirus disease 2019 (Covid-19) (Fig. S3). Patient 3 had clinically low levels of T and B cells. Thus, the reference antibody responses to SARS-CoV-2 in patients 2, 3, and 5 were likely due to exogenous treatments. SARS-CoV-2-specific effector T cell responses were detected in patients 4 and 5, with CD8 + T cells secreting antiviral interferon-γ and tumor necrosis factor, but were only detected at the level of background to patient 2 (Figure 1B and 1C). and figures S4, S5 and S6).
Sequencing of SARS-CoV-2 (Table S2 and Figures S7 and S8) revealed the evolution of spike protein in patients 2 and 3 (Figure 1D and Figure S9); both patients who had been treated with bamlanivimab were deficient in T and B cells. in the N-terminal domain (NTD), regions that have been associated with immune leakage.4 In contrast, no RBD or NTD mutations were found. found in patient 1, who received no antibodies, or in patients 4 and 5, who received convalescent plasma and had intact T cell responses to SARS-CoV-2.
To assess whether viruses obtained from patients 1, 2, and 3 had been neutralized by autologous serum, we constructed infectious pseudoviruses expressing variant peaks (Fig. S10). Serum from patients 1, 2, and 3 did not neutralize pseudoviruses with variant peaks, although serum from patients 2 and 3 neutralized the reference pseudovirus (Fig. S11). Thus, peak mutations in patients 2 and 3 conferred resistance to neutralization to bamlanivimab.
Our results underscore the potential importance of selective pressures such as the use of monoclonal antibodies, in combination with the lack of an effective endogenous immune response, to promote the onset of SARS-CoV-2 escape mutations. These findings highlight the need to better understand the ramifications of different therapies in immunocompromised patients. Our results also corroborate the findings of previous studies in which it was found that patients with B cell deficiencies caused effector T cells, 5 a result that may indicate an important role for T cells in the infection control.
Erin M. Scherer, Ph.D., D.Phil.Ahmed Babiker, MB, BSMax W. Adelman, MDBrent Allman, BAAutum Key, MSJennifer M. Kleinhenz, BSRose M. Langsjoen, Ph.D.Phuong-Vi Nguyen, BSIvy Onyechi, MSJacob D. Sherman, BSTrevor W. Simon, MSHannah SoloffEmory University, Atlanta, GA [email protected]
Jessica Guide, MPHEmory Healthcare, Atlanta, GA
Jay Varkey, MDAndrew S. Webster, MDEmory University, Atlanta, GA
Daniela Weiskopf, Ph.D.La Jolla Institute for Immunology, La Jolla, CA.
Daniel B. Weissman, Ph.D.Yongxian Xu, MDJesse J. Wagoner, MDKatia Koelle, Ph.D.Nadine Rouphael, MDStephanie M. Pouch, MDAnne Piantadosi, MD, Ph.D.Emory University, Atlanta, GA [email protected]
With the support of a contract (75D30121C10084 under BAA ERR 20-15-2997, to Drs Babiker, Wagoner, Koelle and Piantadosi) of the Centers for Disease Control and Prevention; through a grant (5UM1AI148576-02, to Dr. Rouphael and Dr. Scherer) from the National Institutes of Health (NIH); for a Simons Foundation Researcher Award in Mathematical Modeling of Living Systems (Dr. Weissman); and the Pediatric Research Alliance Center for Childhood Infections and Vaccines (Dr. Piantadosi) and Children’s Healthcare in Atlanta and the Emory Woodruff Health Sciences Center Covid-19 Urgent Research Engagement (CURE) Center with support from the O. Wayne Foundation Rollins and the William Randolph Hearst Foundation (to Dr. Piantadosi and Dr. Wagoner). The research reported in this letter was supported by a grant (K08AI139348, to Dr. Piantadosi) from the NIH National Institute of Allergy and Infectious Diseases and a contract (75N9301900065, to Dr. Weiskopf) from NIH. The Jolla Institute for Immunology has applied for patent protection for various aspects of the design of T-cell vaccines and epitopes.
The disclosure forms provided by the authors are available with the full text of this letter at NEJM.org.
The views expressed in this letter are those of the authors and do not necessarily represent the official views of the National Institutes of Health.
This letter was published on June 8, 2022 on NEJM.org.
5 References
-
1. Choi B, Choudhary MC, Regan J, et al. Persistence and evolution of SARS-CoV-2 in an immunocompromised host. N Engl J Med 2020; 383: 2291-2293.
-
2. Greaney AJ, Loes AN, Crawford KHD, et al. Comprehensive mapping of mutations in the SARS-CoV-2 receptor binding domain that affect recognition by polyclonal human plasma antibodies. Cell Host Microbe 2021; 29 (3): 463-476.e6.
-
3. Cele S, Karim F, Lustig G, et al. Prolonged SARS-CoV-2 infection during advanced HIV disease develops a major immune breakdown. Cell Host Microbe 2022; 30 (2): 154-162.e5.
-
4. McCarthy KR, Rennick LJ, Nambulli S, et al. Recurrent deletions of SARS-CoV-2 ear glycoprotein lead to antibody leakage. Science 2021; 371: 1139-1142.
-
5. Gaitzsch E, Passerini V, Khatamzas E, et al. COVID-19 in patients receiving CD20-depleting immunochemotherapy for B-cell lymphoma. 201;