Combined B2M Knockout and Alginate Encapsulation Reduces Immune Rejection of iPSC-Derived Pancreatic Progenitors

Allogeneic cell replacement therapy for diabetes using induced pluripotent stem cell (iPSC)-derived pancreatic progenitors (PPs) is currently limited by immune rejection. Alginate encapsulation and the genetic ablation of HLA-I via β2-microglobulin (B2M) knockout represent distinct strategies to reduce immune recognition and to protect the transplanted cells. We generated encapsulated B2M-knockout pancreatic progenitors (IPS-∆B2M-PPs) and evaluated their immunogenicity in vitro using each of: peripheral blood mononuclear cell (PBMC) co-culture, natural killer cell (NK cell) degranulation, T-cell activation, and rat macrophage adhesion. In vivo studies were also performed to assess capsule biocompatibility and structural stability, following transplantation into rat omentum. While the B2M knockout did not compromise differentiation to pancreatic lineage, encapsulation largely prevented NK cell degranulation regardless of the encapsulated cell type, although T-cells did show residual activation after 7 days of co-culture with encapsulated progenitors, albeit significantly reduced in comparison with non-encapsulated controls. IPS-∆B2M-PP-containing capsules showed reduced macrophage adhesion in vitro compared to wild-type progenitors, while Interleukin-1β (IL-1β) secretion by the macrophages remained negligible. In vivo, all capsules formed a myofibroblastic zone without giant foreign-body cells; IPS-∆B2M-PP implants exhibited the thinnest fibrotic capsules and minimal lymphocytic infiltration. No capsule deformation was observed at 2 weeks or 1 month, indicating preserved structural stability. Although this characterizes both the local tissue response to the biomaterial and capsule stability, it does not establish immune evasion or reduced immunogenicity of the encapsulated cells in vivo. Overall, this study demonstrates the feasibility of combining B2M knockout with alginate encapsulation and suggests a potential additional protective role for B2M deletion under conditions of compromised capsule integrity that requires further investigation.