CD3 downregulation identifies high-avidity, multipotent SARS-CoV-2 vaccine- and recall antigen-specific Th cells with distinct metabolism.

Functional avidity is supposed to critically shape the quality of immune responses, thereby influencing host protection against infectious agents including SARS-CoV-2. Here we show that after human SARS-CoV-2 vaccination, a large portion of high-avidity spike-specific CD4+ T cells lost CD3 expression after in vitro activation. The CD3- subset was enriched for cytokine-positive cells, including elevated per-cell expression levels, and showed increased polyfunctionality. Assessment of key metabolic pathways by flow cytometry revealed that superior functionality was accompanied by a shift toward fatty acid synthesis at the expense of their oxidation, whereas glucose transport and glycolysis were similarly regulated in SARS-CoV-2-specific CD3- and CD3+ subsets. As opposed to their CD3+ counterparts, frequencies of vaccine-specific CD3- T cells positively correlated with both the size of the naive CD4+ T cell pool and vaccine-specific IgG levels. Moreover, their frequencies negatively correlated with advancing age and were impaired in patients under immunosuppressive therapy. Typical recall antigen-reactive T cells showed a comparable segregation into functionally and metabolically distinct CD3+ and CD3- subsets but were quantitatively maintained upon aging, likely due to earlier recruitment in life. In summary, our data identify CD3- T helper cells as correlates of high-quality immune responses that are impaired in at-risk populations.