Table 3

Challenges impeding progress toward the development of effective T-cell biomarkers in T1D

ChallengesPotential solutions and technological advances needed
 High repertoire diversity and low precursor frequency of autoreactive T cells in peripheral bloodDevelop or improve assays capable of measuring the complex mixture of autoreactive T cells
Implement new technologies and approaches for identifying pathogenic signatures, including high-dimensional flow cytometry, mass cytometry, and barcoded antibodies or pMHC multimers for use in scRNA-Seq approaches
Develop sensitive molecular biomarkers capable of detecting signatures of autoreactive T cells, including TCR immunosequencing
 Large numbers of genetic risk variants impacting cellular functionCreate isogenic cellular systems to identify causative SNPs and elucidate their impact on T-cell function
Employ well-characterized biobanks with genotype-selectable donor samples
 High degree of heterogeneity in T-cell phenotypes among subjects with T1DConduct functional testing on subjects with defined phenotypic profiles
Define and control for covariates leading to heterogeneity in T-cell responses
Design and conduct interventional trials using targeted populations with mechanistic outcomes
 Transient or variable autoreactivity over the natural history of the diseaseBuild robust longitudinal and interventional cohorts with sufficient clinical samples
 Low sample volumes in peripheral blood of pediatric samplesWork toward miniaturizing functional assays
Develop surrogate markers of autoreactivity that do not require large sample volumes
 Need for measures that correlate T-cell autoreactivity with endogenous β-cell mass and/or functionDevelop assays capable of detecting signals from autoreactive T cells in circulation reflective of ongoing pathology within T1D islets
Characterize the degree of overlap between tissues and peripheral blood signatures
 Need to understand the pathogenic potential of T-cell subsets or reactivitiesCreate biomimetic devices to model the islet:immune microenvironment
Employ new technologies to test the function of antigen-specific T cells in viable pancreatic tissue sections
 Need for assay reproducibility and interoperabilityEmploy independent validation cores and sample resources capable of repeating assays to test reproducibility and robustness
Incentivize replication testing
 Focus on limited epitopes from known autoantigensConsider nonnative peptides, hybrid peptides, posttranslationally modified peptides
Implement novel high-throughput unbiased peptide screens
Implement novel computational approaches to model peptides capable of activating T cells through the TCR:MHC complex
 Consider alternate concepts to explain origins of autoreactivityImprove understanding of endogenous stress response and host response to commensal bacteria and viral agents, for example
 Focus on classical T1D pathogenesisBroaden studies to include longitudinal studies of T cells in cancer subjects receiving immune checkpoint inhibitors
Understand autoreactivity emanating from rare genetic variants with high penetrance of T1D
 T-cell–centric approachesBroaden studies to better understand T cell:B cell and T cell:APC interactions
Understand exogenous signals that can break T-cell tolerance
 Heavy focus on the pathogenic features of T-cell autoreactivity in subjects with known genetic riskBetter understand the principles related to the mechanisms by which the MHC class II haplotype of DR15-DQ6 influences the T-cell repertoire and leads to dominant protection from disease
  • APC, antigen-presenting cell; pMHC, peptide MHC; scRNA-Seq, single-cell RNA sequencing; SNPs, single nucleotide polymorphisms.