The phenotype and functionality of spike-specific CD4⁺ T cells after COVID-19 vaccination associates with time after transplantation in transplant recipients

A primary series of two mRNA-1273 COVID-19 vaccinations did not induce robust antibody and T cell responses in a large proportion of kidney (KTR) and lung (LTR) transplant recipients. Interestingly, some of these transplant recipients showed spike-specific T cell responses without detectable antibodies. In order to improve the immunogenicity of vaccines in this vulnerable population, this finding warrants in-depth investigation of the spike-specific CD4⁺ T cell phenotype and functionality in these patients. In this in-depth study, we isolated peripheral blood mononuclear cells (PBMCs) from 17 KTR, 13 LTR, and 20 controls, who were previously classified as T cell responders based on IFN-γ ELISpot. The phenotype of the spike-specific CD4⁺ T cells was investigated using AIM assays, and the spike-specific cytokine secretion was measured in cell-culture supernatant following spike-specific stimulation. Twenty-eight days post-second vaccination, a lower frequency of spike-specific CD4⁺ T cells was observed in both KTRs and LTRs compared to controls. In all groups, these spike-specific CD4⁺ T cells were predominantly of the central and effector memory phenotype. Unsupervised hierarchical clustering revealed three distinct clusters of cytokine secretion profiles in the culture supernatants. The cluster with the lowest cytokine diversity had a higher frequency of terminally differentiated spike-specific CD4⁺ T cells. This cluster contained transplant recipients who were classified as antibody non-responders, were significantly shorter after transplantation, and more often received triple immunosuppressive therapy. The KTR with a restricted cytokine secretion profile also remained mostly antibody non-responders after a third vaccination. In conclusion, we show an association between the phenotype and functionality of spike-specific CD4⁺ T cells and the time after transplantation in transplant recipients. This knowledge aids the identification of transplant patients in need of alternative vaccination strategies and possible targets to improve efficacy of vaccines in these patients.