Background/Purpose: The mechanism by which HLA-B27 induces Ankylosing Spondylitis (AS) and related disease is unknown, but a leading hypothesis is that it presents ‘arthritogenic epitopes’ to CD8 T-lymphocytes. Supporting this hypothesis, we have previously demonstrated clonal expansions of CD8 TRB clones adhering to the CASSVG(V/I/L)(Y/F)STDTQYF CDR3 motif in AS patients (PMID: 32162785), which have also been reported by 2 recent independent studies in AS, as well as in samples from patients with bacterial-induced reactive arthritis (PMID: 10201900; 12472659; 28002888). The presence of those clones exclusively in HLA-B*27+ve disease points to a common antigen engaged by receptors of similar structure. The aim of our study was to obtain the paired αβTCR carried by these cells, characterize their phenotype, and, furthermore, to determine whether such clones reside at the normal enthesis.

Methods: We used PBMCs of 2 AS donors screened in our previous study and flow-sorted memory CD8 T-lymphocytes carrying the relevant TRBV genes (TRBV9 and TRBV27) to enrich for our low-frequency target population. Using single-cell RNA sequencing, the whole transcriptome and targeted VDJ sequences were obtained. Healthy enthesal tissue (n=3) was obtained from patients undergoing elective spinal surgery and the peri-enthesal bone (PEB) and the enthesis soft tissue (EST) were disaggregated by mechanical and enzymatic digestion. PEB and EST single-cell suspensions were analyzed by flow cytometry.

Results: The analysis of the enriched memory CD8 T cells from AS patients captured 8 different clonal expansions containing the same TRB consensus CDR3 sequence CASSVGL(Y/F)STDTQYF, and all carried the TRBV9 TCRB. These clones all paired with the same TRAV gene and presented similar TRA-CDR3 sequences. These cells were characterized by the expression of KLRB1, LGALS3, DUSP1, GNLY, ID2, FOS, and JUN whilst lacking HLA-II expression, a marker of activation. Higher expression of inflammatory genes was also observed, including the upregulation of CD69, NFKBIA, TNFAIP3, and PDE4D, an AS-relevant therapeutic target. The analysis of the healthy enthesal tissue confirmed the presence of TRBV9 carriers amongst the memory CD8 and CD4 T cell populations in both the PEB and the EST.

Conclusion: This study confirms the identification of expanded CD8 clonotypes in AS patients, carrying highly conserved TCRB CDR3 sequences, and also demonstrates restriction of the clonotypic TCRA carriage to a single TRAV gene in patients studied thus far. This is consistent with AS being driven by exposure to a common antigen presented by HLA-B27 ie the ‘arthritogenic peptide’ theory of causation of AS. On-going analysis will determine whether the AS-associated clonal expansions resemble the T cell clones that populate the healthy spinal enthesis PEB and EST.

« Back to ACR Convergence 2021

ACR Meeting Abstracts - https://acrabstracts.org/abstract/characterization-of-%ce%b1%ce%b2tcr-of-hla-b27-restricted-cd8-t-cell-clones-associated-with-spondyloarthropathies/