Characterisation of the Immune Response to SARS-CoV-2 Infection / COVID-19 in Type 1 Diabetes
Condition: SARS-CoV2 Infection; Diabetes Mellitus, Type 1
MeSH terms: Infections; Communicable Diseases; COVID-19; Diabetes Mellitus; Diabetes Mellitus, Type 1
Study type: Observational

Summary: 
      Emerging clinical details of the current SARS-CoV-2 pandemic have illustrated that there are
      multiple clinical presentations and outcomes of this viral infection. People with an
      infection have been reported to have a spectrum of disease from severe acute respiratory
      distress requiring ventilation, to mild respiratory or gastrointestinal symptoms and
      asymptomatic presentations. The SARS-CoV-2 pandemic has been accompanied with a substantial
      increase in the number of individuals presenting with new onset type 1 diabetes [1]. Most
      individuals presenting with type 1 diabetes since the start of the COVID-19 pandemic are
      SARS-CoV-2 antibody positive. These findings suggest that SARS-CoV-2 infection can cause type
      1 diabetes. Investigators have identified that many individuals presenting with type 1
      diabetes since the start of the COVID-19 pandemic are SARS-CoV-2 positive by swab or blood
      test. Researchers have also observed that T cells in patients who have had COVID recognise
      some of the peptides in the pancreatic islet cells, which are responsible for production of
      insulin. These findings suggest that SARS-CoV-2 infection may be associated with new onset of
      type 1 diabetes.

      The aim of this project is to understand the host immune response to infection with
      SARS-CoV-2 over time in convalescent newly diagnosed patients with type 1 diabetes, including
      acquired immune responses, gene expression profiling in peripheral blood and to identify host
      genetic variants associated with disease progressions or severity. Participants will have
      Type 1 diabetes and will have had a diagnosis of COVID-19 (confirmed by a positive
      nasopharyngeal swab PCR test and/or SARS-CoV-2 antibody test) and have recovered from
      COVID-19. Samples will be processed and analysed to explore the molecular mechanisms by which
      SARS-CoV-2 infection might precipitate immune attack on insulin-producing cells resulting in
      autoimmune diabetes.
    

Detailed description: 
      AIMS AND OBJECTIVES The objective of this project is to understand the host immune response
      to infection with SARS-CoV-2 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. If the reverse is true, it would provide a plausible mechanism
      for the development of T1D following SARS-CoV-2 infection. We would like to ascertain why
      some people develop T1D after SARS-CoV-2 infection.

      Primary objectives are to determine whether individuals that develop T1D following SARS-CoV-2
      infection bear the HLA A*0201 and HLA*2402 genes known to be associated with T1D and to show
      that SARS-CoV-2-specific killer T-cells in these individuals are capable of directly
      destroying insulin-producing pancreatic beta cells. The ability of SARS-CoV-2-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 might induce T1D.

      Methodology will examine whether convalescent COVID-19 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 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 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=20). 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,
      Londonwho 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.

      A Lay Description:

      The blood will be separated into constituent parts using a thick density gradient solution
      and a centrifuge. The white blood cell layer, or PBMCs, is isolated from other parts of the
      blood and retained. All other parts are discarded in a bleach solution PBMCs are further
      processed to isolate the T-cells, these cells can then go down one of five routes:

        1. The cells are treated to extract the DNA/RNA, and the DNA/RNA is analysed to gain
           information about the T-cell receptors and other associated molecules. The cells are
           killed during DNA/RNA extraction.

        2. The cells will undergo genetic editing to provide information about the processes
           involved in normal T-cell function. Cells will be cultured for 28 days post gene
           editing, will undergo multiple rounds of division, and become non-relevant in terms of
           the Human Tissue Authority (HTA) act 2004.

        3. The cells will be used in an assay where they are exposed to overlapping bits of
           SARS-CoV-2 proteins, they will then be grown in culture for 14 days after which time the
           cells will have undergone division and will be no longer HTA relevant. Cells that
           respond to SARS-CoV-2 proteins will be tested to see whether they have capacity to
           destroy insulin-producing pancreatic beta cells.

        4. Using an automated sorter and fluorescent markers, the cells are sorted and analysed.
           Cells will die during this process which takes place on the day cells are processed in
           the lab.

        5. Cells will be used in plate-based assays to learn about the processes involved in T-cell
           functions. Cells will be disposed after the assay which can take between 1 and 3 days
           depending on type of assay used.

      The date and time of the participants positive SARS-CoV-2 swab/blood test will be captured
      and linked to the participant study record using the unique study number by a member of the
      research team in the direct clinical care teams These details will be stored on a secure NHS
      server and in the site file at each hospital site in a secure, lockable, fireproof container.
      All participants will be given a participant information sheet to take away with them. This
      data set will not be accessible to laboratory-based researchers at any of the connected
      universities.

      This methodology was designed following Patient and Public Involvement, which were conducted
      remotely due to social distancing.
    

Intervention name(s): venous blood sample
Intervention type(s): Procedure
Intervention description(s): Phlebotomy procedure for a venous blood draw.
Other intervention name(s): blood tests
Observational model: Cohort
Time perspective: Prospective
Sampling method: Non-Probability Sample
Study population: 
        Patients diagnosed with new Type 1 diabetes during the COVID pandemic.
      
Biospecimen description: 
      whole venous blood including peripheral blood mononuclear cells and serum.