ABSTRACT Pancreatic islet transplantation offers long-term treatment for Type 1 diabetes, however the shortage of donors and the need for immunosuppressive drugs restrict its therapeutic utility. Islet-like organoids generated from human pluripotent stem cells (PSCs) are an attractive alternative. Moreover, given the increasing appreciation for the role of multi-cellular organization in the functional maturation and maintenance of organs, islet-like organoids have the potential for superior functionality compared to β cells alone. The goal of this project is to develop the next generation of human islet-like organoids (HILOs) from stem cells for efficient and immune evasive transplantation. The underlying hypothesis is that a combination of a novel 3D differentiation protocol, modulation of cell surface signaling, and anti-inflammation transcriptional machinery will enable HILOs to survive long-term and function in an immune competent environment in vivo. To achieve this goal, Aim 1 proposes to establish the long-term efficacy and safety profile of HILOs, of which the function has been validated extensively in vitro. Aim 2 proposes to develop immune-tolerant HILOs by engineering the expression of PD-L1 and demonstrating the efficacy in humanized immune-competent diabetic mice. To further extend protection of transplanted HILOs, Aim 3 proposes to apply pharmacological activation of vitamin D signaling to alleviate cytokine stress on transplanted HILOs. The goal is to determine whether the incorporation of these strategies to improve survival as well as minimize allo-rejection of transplanted HILOs will result in an unlimited supply of therapeutically viable engineered islets for treating diabetes.