Developments in stem cell-derived islet replacement therapy for treating type 1 diabetes

Abstract

The generation of islet-like endocrine clusters from human pluripotent stem cells (hPSCs) has the potential to provide an unlimited source of insulin-producing β cells for the treatment of diabetes. In order for this cell therapy to become widely adopted, highly functional and well-characterized stem cell-derived islets (SC-islets) need to be manufactured at scale. Furthermore, successful SC-islet replacement strategies should prevent significant cell loss immediately following transplantation and avoid long-term immune rejection. This review highlights the most recent advances in the generation and characterization of highly functional SC-islets as well as strategies to ensure graft viability and safety after transplantation.

Keywords: CRISPR; SC-islets; bioreactors; clinical trials; critical quality attributes; cryopreservation; diabetes; differentiation; distribution; encapsulation; immune tolerance; insulin; pancreas; safety; scale-up; single-cell sequencing; stem cells; stress; therapy; transplantation.

Figure 1.. Key pillars of a successful SC-islet therapy for treating T1D.

The remaining challenges limiting the application of SC-islets as a treatment option for diabetes cover a broad range of considerations. The first major component of a successful SC-islet therapy is the ability to produce a highly functional, uniform cell-based product for transplantation. Further optimization of differentiation protocols should aim limit the generation of off-target cell populations and improve the gene expression and chromatin accessibility profiles to match those of human adult islets. These advancements would ideally improve the amount of insulin secreted by SC-β cells and thus reduce the number of cells required per patient. Furthermore, improved transplant survival and a reduced, or eliminated, need for immunosuppressive drugs can be achieved by applying bioengineering strategies. Specifically, using immunomodulating biomaterials or genetically engineered hypoimmune SC-islets could maximize the efficacy of transplanted grafts while minimizing the immune response commonly associated with the transplantation of allogeneic materials. Finally, regulatory standards must be established to maximize the safety and efficacy of SC-islet transplantation. This includes improving methods for large-scale manufacturing of SC-islets with sufficient quality control characterizations, such as avoiding mutations that pose safety risks, and developing standardized methods for storage and distribution. Additionally, as these challenges are addressed and SC-islet products are improved, guidelines to optimize dosing, transplantation site, and graft survival must be established.

Figure 2.. Growth factors and small molecules used during the multistage differentiation of hPSCs to SC-islets.

The generation of SC-islets from hPSCs is a multistage process that involves the temporal application of small molecules and growth factors (highlighted in red). At each stage, specific developmental pathways are targeted to mimic human pancreatic development. While multiple differentiation protocols have been reported, each follows the same general differentiation trajectory by first specifying hPSCs into definitive endoderm using Activin A (AA) and WNT pathway activation, most often with CHIR99021. Next, definitive endoderm cells are guided into a primitive gut tube state using keratinocyte growth factor (KGF), also known as FGF7. Notably, some protocols include additional compounds that target WNT (e.g., IWP2) and TGF-β signaling pathways to improve primitive gut tube specification. Generating pancreatic progenitor cells requires additional activation of protein kinase C with TPPB and retinoic acid signaling pathways while simultaneously inhibiting BMP signaling with LDN193189 and SHH signaling with SANT1. Epidermal growth factor (EGF) and nicotinamide have also been shown to improve the generation of PDX1⁺/NKX6-1⁺ pancreatic progenitors. Interestingly, it has been reported that the use of a selective BET inhibitor can maintain pancreatic progenitors cells in a proliferative and expandable state. This can potentially accelerate SC-islet manufacturing by providing an intermediate point from which to generate SC-islets. During the later stages of SC-islet differentiation, reported protocols begin to diversify both in nomenclature of cell populations and compounds applied. For example, latrunculin A (LatA) has been used to specify endocrine cells in planar culture. Additional nomenclature differences arise with some protocols describing the initial SC-islet product as “immature” and include an additional stage to generate “mature” SC-islets using an aurora kinase inhibitor (ZM447439) or WNT analogs.

Figure 3.. Critical quality attributes of SC-islets.

Despite making up a very small percentage of the pancreas, primary islets function to maintain blood glucose levels within a tightly regulated range. SC-islets have been able to replicate some of the key attributes of native islet function but fall short in some areas. Critical quality attributes that have been achieved in SC-islets are indicated in [black], while those that remain unresolved are indicated in [red].