Characterisation of the Immune Response to SARS-CoV-2 and Other Common Infections in Type 1 Diabetes

AIMS AND OBJECTIVES The objective of this project is to understand the host immune response to infection with SARS-CoV-2 and other common infections over time in convalescent patients that develop type 1 diabetes (T1D). Preliminary data show that diabetogenic T-cells in patients that developed T1D before 2019 can recognise SARS-CoV-2 and other common human pathogens. If the reverse is true, it would provide a plausible mechanism for the development of T1D following SARS-CoV-2 or some common infections. We would like to ascertain why some people develop T1D after infection.

Primary objectives are to determine whether individuals that develop T1D following infection bear the HLA A*0201 and HLA*2402 genes known to be associated with T1D and to show that SARS-CoV-2/pathogen-specific killer T-cells in these individuals are capable of directly destroying insulin-producing pancreatic beta cells. The ability of pathogen-specific T-cells to recognise diabetogenic T-cell epitopes will be examined using peptide-HLA multimers. Cross-reactive T-cell receptors will be manufactured for biophysical and structural analyses to enable an understanding of how SARS-CoV-2 and other common infectious agents might induce T1D.

Methodology will examine whether convalescent patients that develop T1D predominantly express the human leukocyte antigens (HLAs) HLA A*02 and HLA*24 by staining a small part of the sample with antibodies for these HLA. Investigators anticipate that there could be a large enrichment for these disease risk HLA (>75% of patients expressing one of these disease-risk alleles) [2]. T-cell library [3] and peptide-HLA multimer [4-6] approaches will be used to isolate T-cells that recognise epitopes from SARS-CoV-2 and other common pathogens via HLA A*02 and HLA*24. The monoclonal T-cell populations generated will be tested for recognition of known diabetogenic epitopes present on the surface of human pancreatic beta cells. Where such T-cell cross-reactivity [7] is identified the team will sequence the complementarity determining region of the cognate T-cell receptor so that it can be manufactured as a soluble protein [8] for further biophysical and structural studies [9,10] aimed at understanding how SARS-CoV-2 and other common pathogens might induce T1D in a minority of individuals.

The study will be prospective, observational study. Participants will be convalescent patients with recent new diagnosis of type 1 diabetes (n=30). Eligible participants will be identified and referred from Consultant Paediatric Endocrinologists at The Department of Child Health at Cwm Taf Morgannwg University Health board, The Department of Child Health, Cardiff and the Vale health board and Department of Child Health, St, Mary's Hospital, London who are all part of the clinical research study team and direct clinical care team. All participants and their parents /guardians will receive a participant information sheet and have time and privacy to read and understand the content. A member of the study team trained in Good Clinical Practice will be available to provide support and answer any questions. Informed consent will then be gained in writing on a consent form, or in the case of paediatric participants on an assent form and a Parent/Guardian consent form. Participants will be issued with a unique study number.

A sample of blood will be taken by a trained phlebotomist, 50mls (or less in children according to the participant's weight). The sample will be anonymised and issued with the unique study number. We will require peripheral blood mononuclear cells (PBMC) from of blood on the day of recruitment. Anonymised blood samples will be transferred for T cell research and Cwm Taf University Health board laboratory for serology tests (Roche SARS-CoV-2 IgM and IgG assay at a UKAS accredited laboratory) with appropriate UN3373 packaging. Samples will be used for immunological, virological and host genetic assays. Bloods will be processed to isolate serum and PBMC, including host RNA and DNA. T-cell libraries will be established using the PBMC. T-cell clones generated will be tested for cross-reactivity with established diabetogenic T-cell epitopes. The T-cell receptors (TCRs) will be manufactured from cross-reactive T-cells for further study. PBMC samples will also be stained with cognate peptide-HLA multimers using flow cytometry. This will generate information about their phenotype and activation status.