Integrated Immunological Engineering Platform for Scalable Production of iPSC-Derived Megakaryocytes

Severe thrombocytopenia and impaired platelet function remain major clinical challenges in hematologic diseases, chemotherapy, bone marrow failure, surgery and obstetric haemorrhage. Conventional platelet transfusions are limited by donor dependency, short shelf life, storage constraints, and immune-mediated platelet refractoriness caused primarily by HLA incompatibility. Here, we present a scalable, donor-independent platform to generate universally compatible, low-immunogenic megakaryocytes (MKs) from induced pluripotent stem cells (iPSCs). Human iPSCs were genetically engineered by shRNA-mediated silencing of β2-microglobulin (β2M) and/or class II transactivator (CIITA). Differentiation of HLA-silenced iPSCs resulted in MKs with reduced HLA class I and II expression, capable to release functional platelets in vitro and sustain platelet production in humanised mice with physiological distribution. To enhance product safety, gamma irradiation effectively eliminated proliferative contaminants while preserving MK function. This platform integrates immune engineering with scalable iPSC-based manufacturing and safety optimization, addressing key limitations of conventional donor-derived platelet products. Together, these findings demonstrate that HLA-silenced iPSC-derived MKs represent a promising off-the-shelf source of functional platelets with reduced immunogenicity. By combining donor-independent production, targeted immune engineering, and scalable manufacturing, this promising approach has the potential to improve transfusion support, overcome platelet refractoriness associated with HLA incompatibility, and expand access to readily available platelet therapies across diverse clinical settings.