Regulatory T-cell (Treg) immunotherapy has emerged as a promising and highly effective strategy to combat graft-versus-host disease (GvHD) after allogeneic hematopoietic cell transplantation (allo-HCT). Both naturally occurring Treg and induced Treg populations have been successfully evaluated in trials illustrating the feasibility, safety, and efficacy required for clinical translation. Using a non-mobilized leukapheresis, we have developed a good manufacturing practice (GMP)-compatible induced Treg product, termed iG-Tregs, that is enriched in cells expressing the potent immunosuppressive human leucocyte antigen-G molecule (HLA-G+). To assess the safety and the maximum tolerable dose (MTD) of iG-Tregs, we conduct a phase I–II, two-center, interventional, dose escalation (3 + 3 design), open-label study in adult patients undergoing allo-HCT from an HLA-matched sibling donor, which serves also as the donor for iG-Treg manufacturing. Herein, we present the clinical protocol with a detailed description of the study rationale and design as well as thoroughly explain every step from patient screening, product manufacturing, infusion, and participant follow-up to data collection, management, and analysis (registered EUDRACT-2021-006367-26).