Immunological effects of subcutaneous and sublingual immunotherapy in house dust mite-allergic adults: a nine-month prospective pilot study

House dust mite (HDM) allergy contributes to allergic rhinitis and asthma worldwide. Allergen-specific immunotherapy (AIT) is the only disease-modifying treatment, with immunoglobulin G4 (IgG4) and regulatory T cells (Tregs) mediating immune tolerance. Comparative immunological effects of subcutaneous (SCIT) versus sublingual (SLIT) therapy remain underexplored. To evaluate immunological changes induced by SCIT and SLIT and their association with clinical improvement in HDM-allergic patients. In this prospective cohort, 43 adults with HDM-sensitized allergic patients received SCIT (n = 22) or SLIT (n = 21) for nine months. Clinical outcomes were assessed using the Asthma Control Test (ACT) and Rhinitis Control Assessment Test (RCAT). Serum IgG4 and total IgE were measured by ELISA and electrochemiluminescence, respectively, and CD4⁺CD25⁺FoxP3⁺ Tregs were analyzed by flow cytometry. Responders were defined as patients achieving ≥ 20% improvement in ACT or RCAT. Wilcoxon signed-rank, Mann-Whitney U, and Spearman correlation tests were used. AIT increased IgG4 (320 → 920 ng/mL; p < 0.001) and Tregs (3.4% → 6.3%; p < 0.001), with a non-significant decrease in IgE. Responders had higher IgG4 and Tregs and lower IgE than non-responders. SCIT elicited higher IgG4 levels (median 1035 vs 705 ng/mL) and a trend toward greater Treg expansion compared with SLIT, although clinical improvement was similar between groups. IgG4 correlated with ACT (p < 0.001) and RCAT (p = 0.002), and Tregs correlated positively with IgG4 (p = 0.003) and inversely with IgE (p = 0.010). Both SCIT and SLIT improve clinical outcomes in HDM-allergic patients via total IgG4 elevation and Treg expansion. SCIT may induce stronger systemic immunological responses, supporting the use of these biomarkers for early monitoring and personalized therapy. These findings should be interpreted within the context of a pilot study with a relatively small sample size.