A let-7 microRNA-RALB regulatory axis links the immune properties of iPSC-derived megakaryocytes in platelet production

We previously conducted the first-in-human clinical trial of human induced pluripotent stem cell-derived platelets (iPSC-PLTs), which were generated from immortalized megakaryocyte progenitor cell lines (imMKCLs). The reverse translational insights gained from this trial indicated the presence of immune subsets within the imMKCL population that have unknown effects on the quality of the imMKCLs and PLT production. Here, we observed heterogenous let-7a-5p and let-7g-5p activity among imMKCLs during the proliferation stage and demonstrated that let-7 microRNA-switch biotechnology can be used to enrich immune-skewed subpopulations in imMKCLs. We identified RAS like proto-oncogene B (RALB) as a downstream target of let-7a-5p and a crucial factor in determining immune-biased transcriptional signatures. We further found that the induced immune-related signaling by imMKCLs led to arrested proliferation and deficient PLT production. Our findings suggested that the let-7 microRNA-RALB axis is important for modulating immune-skewed properties in imMKCLs and that RALB may serve as a potential predictor for the quality control of iPSC-PLT production. Overall, the study provides important insights into the immune properties of imMKCLs and their implications for ex vivo iPSC-PLT manufacturing towards clinical applications.