Microfluidic chip systems for characterizing glucose-responsive insulin-secreting cells equipped with FailSafe...

Area of Science:

• Biotechnology
• Regenerative Medicine
• Diabetes Research

Background:

• Type 1 diabetes (T1D) treatment faces challenges with pluripotent cell-derived islet replacement therapy due to uncontrolled cell proliferation and tumorigenicity risks.
• A critical need exists for enhanced safety mechanisms in cell-based therapies for T1D.

Purpose of the Study:

• To establish a proof-of-concept for a glucose-responsive, insulin-secreting cell line with an integrated FailSafe kill-switch.
• To enhance the safety of pluripotent cell-derived insulin-secreting cells for T1D treatment.

Main Methods:

• Generated β cell-induced progenitor-like cells (βiPLCs) from mouse pancreatic β cells via interrupted reprogramming.
• Utilized CRISPR/Cas9 to create a FailSafe βiPLC line (FSβiPLCs) by linking the HSV-thymidine kinase (TK) kill-switch to the Cdk1 gene.
• Employed PDMS-based transcapillary microfluidic systems for evaluating kill-switch functionality and glucose-stimulated insulin secretion (GSIS) assays on-chip.

Main Results:

• FSβiPLCs demonstrated essential pancreatic β cell properties, including glucose-responsive insulin secretion.
• The integrated FailSafe kill-switch effectively ablated dividing cells in a dose-dependent manner with ganciclovir (GCV), while non-dividing cells remained resistant.
• Resistant, non-dividing FSβiPLCs exhibited enhanced insulin secretion and glucose responsiveness compared to proliferating cells.

Conclusions:

• This study provides a proof-of-concept for a FailSafe kill-switch system in glucose-responsive, insulin-secreting cells, addressing safety concerns in T1D cell replacement therapy.
• Microfluidic systems were instrumental in assessing the functionality and safety of engineered cells, highlighting the kill-switch's potential to mitigate tumorigenicity risks.
• The developed system offers a promising strategy for safer pluripotent cell-derived therapies for Type 1 diabetes.