RPS26 Expression as a Predictive Biomarker and Functional Modulator in Sublingual Immunotherapy for Japanese Cedar...

Japanese cedar pollinosis (JCP) is highly prevalent in Japan, and sublingual immunotherapy (SLIT) remains one of the few effective disease-modifying treatments. While many patients have significant benefits, biomarkers predicting SLIT responsiveness are lacking. Single-cell RNA sequencing (scRNA-seq) of peripheral blood mononuclear cells (PBMCs) identified the ribosomal protein gene RPS26 as a candidate associated with treatment response. This study aimed to determine whether RPS26 expression contributes to SLIT efficacy and to define its involvement in T-cell dynamics, cytokine production, and epigenetic regulatory pathways.

PBMCs from 35 patients with JCP were analysed before SLIT, and paired samples from 26 patients were evaluated after 1 year of SLIT. scRNA-seq was performed in four responders and three non-responders. Changes in T-cell subsets and interleukin (IL)-5 and IL-10 secretion were evaluated during SLIT. Functional relevance was examined through small interfering RNA-mediated RPS26 knockdown followed by allergen stimulation. The relationship between RPS26 and the ten-eleven translocation family (TET2 and TET3) expression was assessed by quantitative PCR.

scRNA-seq revealed significant RPS26 upregulation in responders. Although bulk RPS26 expression did not differ between responders and non-responders, all subjects with high RPS26 expression (≥1,000 relative units) were responders. SLIT increased follicular regulatory and type 1 regulatory T cells in responders, whereas IL-10 elevation occurred exclusively in high-RPS26 responders. RPS26 knockdown enhanced early apoptosis and cell death, and reduced IL-10 expression following allergen challenge. RPS26 expression was strongly correlated with the epigenetic regulators TET2 and TET3, both essential for Treg-cell stability.

RPS26 expression was enriched in clinical responders to SLIT and was associated with enhanced IL-10 induction, expansion of regulatory T (Treg)-cell subsets, resistance to antigen-induced apoptosis, and strong correlations with the epigenetic regulators TET2 and TET3. Rather than serving as a universal predictive biomarker, RPS26 appears to define a responder-enriched molecular phenotype linked to tolerance-associated immune programming. Larger prospective studies and mechanistic investigations will be required to validate its clinical utility and clarify causal pathways.