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Emerging SARS-CoV-2 variants have created concern in the general public and the scientific community for their potential to evade the immunity elicited against the original Wuhan strain. To date, several SARS-CoV-2 variants have been identified around the world. The CDC has classified several variants such as B.1.1.7 (United Kingdom), B.1.351 (South Africa), P.1 (Brazil), B.1.427 (California), and B.1.429 (California) as variants of concern. This is because of their increased transmissibility and reduced susceptibility to neutralization. Nevertheless, thus far, neutralizing responses induced by infections or vaccines have been only minimally to moderately diminished by these variants.
Taking a Closer Look at T cell Responses against SARS-CoV-2
Neutralizing antibodies block viral cell-entry by targeting the Spike protein. Therefore, studies evaluating immunity to SARS-CoV-2 have been heavily focused on qualifying and quantifying neutralizing antibody responses. Nevertheless, beyond B cell-mediated antibody responses, SARS-CoV-2 infection elicits both CD4+ and CD8+ T cell responses. Among these, CD4+ T cell responses are more robust and target most SARS-CoV-2 proteins. It has been shown that the magnitude of the CD4+ T cell response correlates well with viral protein abundance, being more prominent against Spike, M, and nucleocapsid proteins (Sette and Crotty, 2021).
Specific and robust T cell responses against SARS-CoV-2 help controlling infections. Particularly, CD4+ T cell responses play a critical role in reducing disease severity (Rydyznski Moderbacher et al. 2020). Therefore, with the emergence of new SARS-CoV-2 variants, it is crucial to evaluate the resilience of T cell responses present in vaccinated or COVID-19 convalescent individuals. To this end, a new study takes advantage of synthetic peptides to specifically address how T cell immunity to the original Wuhan strain may fare against new viral variants.
Peptide Library Design Tools: Overlapping peptide libraries can be used for different applications such as mapping antibody epitopes, identifying T-cell epitopes, and pinpointing binding sites within a protein of interest.
Accordingly, Tarke et al. 2021 generated synthetic peptides that spanned the entire sequence of the original SARS-CoV-2 Wuhan strain or that of variants such as B.1.1.7, B1.351, P.1, and CAL20C. Basically, creating libraries of overlapping peptides, each consisting of 15 mers and overlapping by 10 amino acids. Tarke and colleagues proceeded to use peptide pools for each SARS-CoV-2 protein of interest to enable testing of T cell reactivities.
T cell responses were analyzed through in vitro assays using peripheral blood mononuclear cells (PBMCs) derived from COVID-19 convalescent or vaccinated individuals (i.e., Moderna mRNA-1273 or Pfizer/BioNTech BNT162b2). To evaluate T cell activation, Tarke and colleagues relied on a flow cytometry-based activation induced cell marker (AIM) assay. AIM assays support the specific and sensitive detection of antigen responses. In this assay, isolated PBMCs are cultured in the presence of viral peptide pools followed by immunostaining against specific T cell identity and activation markers, such as CD3 (total T cells), CD4, CD8, CD137, OX40, and CD69 (Grifoni et al. 2020).
By following this approach, Tarke et al. showed mostly conserved CD4+ and CD8+ T cell reactivities against Spike peptide pools from viral variants compared to those derived from the original SARS-CoV-2 strain. Only a mild decrease in T cell reactivity was seen against Spike peptide pools from the B.1.351 variant when analyzing PBMCs from immunized individuals.
Additionally, T cell responses were functionally evaluated through FluoroSpot assays by measuring the secretion of IFNγ. Tarke et al. found a moderate decrease in IFNγ secretion following T cell activation with Spike peptides from the B.1.1.7, P.1, and CAL.20C variants compared to control peptide pools.
Lastly, mutations in SARS-CoV-2 variants span sequences beyond the Spike protein. Tarke and colleagues probed convalescent T cell reactivities using peptide pools covering the entire viral proteome of the original Wuhan strain and variants. These assays revealed that T cell responses were essentially unaffected by variant specific mutations.
Overall, analysis of previously identified SARS-CoV-2 CD4+ and CD8+ T cell epitopes agree with these functional findings, demonstrating that most T cell epitopes are unaffected by variant associated mutations (Tarke et al. 2021).
Peptide Libraries for Viral Diseases
Grifoni, A. et al. Targets of T Cell Responses to SARS-CoV-2 Coronavirus in Humans with COVID-19 Disease and Unexposed Individuals. Cell (2020) doi:10.1016/j.cell.2020.05.015.
Rydyznski Moderbacher, C. et al. Antigen-Specific Adaptive Immunity to SARS-CoV-2 in Acute COVID-19 and Associations with Age and Disease Severity. Cell (2020) doi:10.1016/j.cell.2020.09.038.
Tarke, A. et al. Comprehensive analysis of T cell immunodominance and immunoprevalence of SARS-CoV-2 epitopes in COVID-19 cases. Cell Reports Med. (2021) doi:10.1016/j.xcrm.2021.100204.
Tarke, A. et al. Negligible impact of SARS-CoV-2 variants on CD4 + and CD8 + T cell reactivity in COVID-19 exposed donors and vaccinees. bioRxiv Prepr. Serv. Biol. (2021) doi:10.1101/2021.02.27.433180.