3D Islet Assay Development and SAR of Chromenones for Beta-Cell Replenishment

Abstract

Diabetes poses a global health crisis affecting individuals across age groups and backgrounds, with a prevalence estimate of 700 million people worldwide by 2045. Current therapeutic strategies primarily rely on insulin therapy or hypoglycemic agents, which fail to address the root cause of the disease - the loss of pancreatic insulin-producing beta-cells. Therefore, bioassays that recapitulate intact islets are needed to enable drug discovery for beta-cell replenishment, protection from beta-cell loss, and islet-cell interactions. Standard cancer insulinoma beta-cell lines MIN6 and INS-1 have been used to interrogate beta-cell metabolic pathways and function but are not suitable for studying proliferative effects. Screening using primary human/rodent intact islets offers a higher level of physiological relevance to enhance diabetes drug discovery and development. However, the three-dimensionality of intact islets has presented challenges in developing robust, high-throughput assays to detect beta-cell proliferative effects. Established methods rely on either dissociated islet cells plated in 2D monolayer cultures for imaging or reconstituted pseudo-islets formed in round bottom plates to achieve homogeneity. These approaches have significant limitations due to the islet cell dispersion process. To address these limitations, we have developed a robust, intact pancreatic islet imaging platform in 384-well format that is capable of detecting diabetes-relevant endpoints including beta-cell proliferation, chemoprotection, and islet spatial morphometrics. We were then able to apply our method to a preliminary in vivo study of a new scaffold for beta-cell modulation. Islets were isolated from the pancreata of mice after a week of exposure to our compound, GNF-9228. This method proved to be a drastic improvement over classical histological analysis for in vivo beta-cell proliferation and encouraged us to pursue a structure-activity relationship campaign to improve the physical properties of our lead compound. Subsequent work led to the evaluation of over seventy compounds in bio various assays. Additionally, we were able to leverage existing scRNA seq data from Type 2 diabetes samples to evaluate our compound’s effects on the human kinome.