The NIAID national protocol intends to enroll at least 1000 COVID19 patients in a study designed to track infected patients through tracking, over time, their immune responses, viral load, and a variety of multi-omic analytes that can provide deep insights into how infection by SARS-CoV-2 is revealed in host defense responses and disease- perturbed networks. At the heart of this study is the establishment of high quality biorepositories that can be used to quantitatively assess viral load, quantitatively interrogate viable PBMCs, and permit direct comparisons between different patients and different time points of disease progression. The nature of the infection, with highly differential patient outcomes, will eventually require significant computational efforts that can account for confounding factors such as co-morbidities, the influence of various therapies that are being broadly tested in these patients, etc. It will also certainly require both broadly available immune characterization tools that can be applied on all patient samples, but also specialized tools that can be used to inform the interpretation of the general analytics. In this project, we propose to integrate two sets of immune cell characterizations into the national NIAID effort. Those characterizations include single cell, functional phenotyping of select immune cell classes via an analysis designed to quantitate the levels of 35 secreted proteins from up to 2000 single cells of a given immune cell type. The second characterization is based upon reducing proteins from the SARS-CoV-2 into peptide antigen-major histocompatibility complex (pMHC) libraries that can be used to identify SARS-CoV- 2 antigen-specific CD8+ and CD4+ T cell populations from isolated, viable PBMCs. These assays provide deep and complementary information that will significantly inform the interpretation of the immune phenotyping assays that constitute the COREs of the NIAID study.
A proposal to link an Institute for Systems Biology/Swedish Medical Center ongoing investigation of COVID19 patients to the larger NIAID nationwide COVID19 patient immunophenotyping study is described. The primary aim is to introduce two single-cell based phenotyping assays into the study. The first is to establish the polyfunctionality of designated immune cell populations via the analysis of a panel of 35 secreted proteins from each of a couple thousand cells per immune cell-type, with an initial focus on CD4+ and CD8+ T cells, but potentially broaded to include additional cell types as indicated via a broad survey scan of patient PBMCs using sc-RNA seq that incorporates a large panel of DNA-labeled antibody surface markers. The second immunophenotyping assay builds upon recently developed technologies to pair SARS-CoV-2 antigen specific T cell populations with T cell receptor genes for select, common HLA alleles. The basic technologies underlying these antigen-specific T cell assays are described, along with preliminary results from library construction.
