Scalable microfluidics that can address the challenge of delivery of large transgenes to therapeutic cells

To accelerate the manufacturing innovation, the goal of this project is to validate a cell engineering technology that uses a microfluidic transfection for delivery of therapeutic transgenes to four cell types of importance to the pharma industry.

Industry Need

Cost of manufacturing genetically engineered cells for therapies is significant, with high process complexity, and variable cell product quality.

Approach

Validate a cell engineering technology that uses microfluidic transfection for delivery of therapeutic transgenes.

Impacts

Demonstrate mechanoporation transfection of iPSCs with large CRISPR-correction plasmids

Develop a non-viral, mechanical approach to transfection of gene editing cargo, including CRISPR/Cas9, DNA, and mRNA

Demonstrate high efficiency T cell knockout editing using mechanoporation

Demonstrate clinical scale processing (>1B cells) of T cells

Value Statement/Outcomes

By implementing a GMP-compliant, reagent-free microfluidic mechanoporation platform for transgene delivery as demonstrated by this project, an organization will reduce gene modification costs by eliminating viral vectors and cutting electroporation-related cell loss (improving viability from ~50% to >80%), enabling scalable processing from 1 million to 1 billion cells with 2–3× higher transfection efficiency

Outputs/Deliverables

Developed consumable transfection chip for T cells and iPSCs that was delivered to partners

Demonstrated first mechanoporation transfection of iPSCs with large CRISPR-correction plasmids

Demonstrated high efficiency T cell knockout editing using mechanoporation

Demonstrated clinical scale processing (>1B cells) of T cells

Publications

Loo, J., Sicher, I., Goff, A., Kim, O., Clary, N., Alexeev, A., Sulchek, T., Zamarayeva, A., Han, S., & Calero-Garcia, M. (2021). Microfluidic transfection of mRNA into human primary lymphocytes and hematopoietic stem and progenitor cells using ultra-fast physical deformations. Scientific Reports, 11(1). https://doi.org/10.1038/s41598-021-00893-4

Stone, N. E., Voigt, A. P., Mullins, R. F., Sulchek, T., & Tucker, B. A. (2021). Microfluidic processing of stem cells for autologous cell replacement. Stem Cells Translational Medicine, 10(10), 1384-1393. https://doi.org/10.1002/sctm.21-0080

Presentations

New mechanical approaches to perform cell transfections, Cellicon Valley 2021 – Viral Vector Free Delivery, May 6, 2021

Scalable microfluidics that can address the challenge of delivery of large transgenes to therapeutic cells (PC2.2-122), NIIMBL Member Forum, Virtual, August 26, 2021.

Tong Yu, Peter Shankles, Noah Kramer, Shaylyn Grier, Todd Sulchek, “Fabrication and characterization of a rigid microfluidic mechanoporation device with high pressure tolerance and high cell transfection”, IEEE 35th International Conference on Micro Electro Mechanical Systems Conference (MEMS) 2022, Online