A 3D culture platform enables development of zinc-binding prodrugs for targeted proliferation of β cells

Abstract

Advances in treating β cell loss include islet replacement therapies or increasing cell proliferation rate in type 1 and type 2 diabetes, respectively. We propose developing multiple proliferation-inducing prodrugs that target high concentration of zinc ions in β cells. Unfortunately, typical two-dimensional (2D) cell cultures do not mimic in vivo conditions, displaying a markedly lowered zinc content, while 3D culture systems are laborious and expensive. Therefore, we developed the Disque Platform (DP)—a high-fidelity culture system where stem cell–derived β cells are reaggregated into thin, 3D discs within 2D 96-well plates. We validated the DP against standard 2D and 3D cultures and interrogated our zinc-activated prodrugs, which release their cargo upon zinc chelation—so preferentially in β cells. Through developing a reliable screening platform that bridges the advantages of 2D and 3D culture systems, we identified an effective hit that exhibits 2.4-fold increase in β cell proliferation compared to harmine.

A therapeutic convection–enhanced macroencapsulation device for enhancing β cell viability and insulin secretion

“…GSIS. The analysis was conducted following previously described procedure (48). The ceMEDs were loaded 150 μL of MIN6 cells or SC-βCs at a seeding density with 3 million cells or 2,000 IEQ.…”

Construction of 3D hierarchical tissue platforms for modeling diabetes

“…3(a) ]. 46 The developed DP included core extracellular matrix (ECM) components such as laminin and collagen type IV that are involved in human islet development. Consequently, they observed higher expression rates of key transcription factors (e.g., PDX1 and NKX6.1) and junctional structures (e.g., E-cadherin and connexin 36) in the 3D reconstituted beta cells inside the DP compared to 2D monolayer conditions.…”

Harnessing Proliferation for the Expansion of Stem Cell-Derived Pancreatic Cells: Advantages and Limitations

“…Screening primary human b-cells for mitogens that had been previously identified to initiate proliferation in rodent b-cells revealed that many factors were unable to initiate significant replication, with the dual-specificity tyrosine phosphorylation regulated kinase 1A (DYRK1A)-inhibiting compound harmine being the only mitogen that induced significantly increased replication with treatment (31). Indeed, harmine is unique in that it has been repeatedly reported in the literature to induce bcell and other pancreatic cell replication (32)(33)(34)(35)(36)(37). Differing effects on b-cell replication through shared signaling mechanisms between rodent and human b-cells may be due to additional cellular factors that inhibit proliferation, such as the high accumulation of p16 in adult human b-cells (38)(39)(40).…”

“…With this tool, it is possible to screen for chemical or genetic regulators that provide control over proliferation in pancreatic cells. In addition to using compound-based treatments previously mentioned, such as harmine (34,35,37), another method for inducing replication could be through viral delivery of controllable systems (70)(71)(72). Once proliferative drivers have been identified, rigorous examination of the expanded population must confirm that no aberrant mutations have been acquired during the process and that expanded cells retain the ability to form b-like cells.…”

“…The study utilized β-like cells in an in vitro 3D platform to screen for molecules that targeted β-like cell replication. This novel platform allowed for the development of a harmine-carrying zinc-binding prodrug that was more effective than harmine alone at expanding β-like cells ( 37 ). Therefore, further development of compounds that induce targeted proliferation provide promising therapeutic avenues for both inducing and regulating β-like cell replication.…”