Single-cell immunoprofiling after immunotherapy for allergic rhinitis reveals functional suppression of pathogenic TH2 cells and clonal conversion

Background: Allergic rhinitis is a growing problem worldwide. Currently the only treatment that can modify the disease is antigen-specific immunotherapy, but its mechanism of action is not fully understood. Objective: We comprehensively investigated the role and changes of antigen-specific T cells before and after sublingual immunotherapy (SLIT) for Japanese cedar pollinosis. Methods: We cultured peripheral blood mononuclear cells obtained both before and 1 year after initiating SLIT and used a combination of single-cell RNA sequencing and repertoire sequencing. To investigate biomarkers, we used cells frompatients participating a phase 2/3 trial of SLIT tablets for Japanese cedar pollinosis and cells from outpatients with good and poor response. Results: Antigen-stimulated culturing after SLIT led to clonal expansion of T(H)2 and regulatory T cells, and most of these CD4(+) T cells retained their CDR3 regions before and after treatment, indicating antigen-specific clonal responses and differentiation resulting from SLIT. However, SLIT reduced the number of clonal functional T(H)2 cells but increased the trans-type T(H)2 cell population that expresses musculin (MSC), TGF-beta, and IL-2. Trajectory analysis suggested that SLIT induced clonal differentiation of the trans-type T(H)2 cells differentiated into regulatory T cells. Using real-time PCR, we found that the MSC levels increased in the active SLIT group and those with good response after 1 year of treatment. Conclusion: The combination of single-cell RNA sequencing and repertoire analysis helped reveal part of the underlying mechanism: SLIT promotes the expression of MSC on pathogenic T(H)2 cells and suppresses their function. MSC may be a potential biomarker of SLIT for allergic rhinitis.