Résumé : Background: A 3-week short-course of adjuvant-free hydrolysates of Lolium perenne peptide (LPP) immunotherapy for rhinoconjunctivitis with or without asthma over 4 physician visits is safe, well tolerated, and effective. Objective: We sought to investigate immunologic mechanisms of LPP immunotherapy in a subset of patients who participated in a phase III, multicenter, randomized, double-blind, placebo-controlled trial (clinical.gov NCT02560948). Methods: Participants were randomized to receive LPP (n = 21) or placebo (n = 11) for 3 weeks over 4 visits. Grass pollen–induced basophil, T-cell, and B-cell responses were evaluated before treatment (visit [V] 2), at the end of treatment (V6), and after the pollen season (V8). Results: Combined symptom and rescue medication scores (CSMS) were lower during the peak pollen season (−35.1%, P =.03) and throughout the pollen season (−53.7%, P =.03) in the LPP-treated group compared with those in the placebo-treated group. Proportions of CD63 + and CD203c bright CRTH2 + basophils were decreased following LPP treatment at V6 (10 ng/mL, P <.0001) and V8 (10 ng/mL, P <.001) compared to V2. No change in the placebo-treated group was observed. Blunting of seasonal increases in levels of grass pollen–specific IgE was observed in LPP-treated but not placebo-treated group. LPP immunotherapy, but not placebo, was associated with a reduction in proportions of IL-4 + T H 2 (V6, P =.02), IL-4 + (V6, P =.003; V8, P =.004), and IL-21 + (V6, P =.003; V8, P =.002) follicular helper T cells. Induction of FoxP3 + , follicular regulatory T, and IL-10 + regulatory B cells were observed at V6 (all P <.05) and V8 (all P <.05) in LPP-treated group. Induction of regulatory B cells was associated with allergen-neutralizing IgG 4 -blocking antibodies. Conclusion: For the first time, we demonstrate that the immunologic mechanisms of LPP immunotherapy are underscored by immune modulation in the T- and B-cell compartments, which is necessary for its effect.