A Microfluidic Device to Enhance Viral Transduction Efficiency During Manufacture of Engineered Cellular Therapies

Abstract

The development and approval of engineered cellular therapies are revolutionizing approaches to treatment of diseases. However, these life-saving therapies require extensive use of inefficient bioprocessing equipment and specialized reagents that can drive up the price of treatment. Integration of new genetic material into the target cells, such as viral transduction, is one of the most costly and labor-intensive steps in the production of cellular therapies. Approaches to reducing the costs associated with gene delivery have been developed using microfluidic devices to increase overall efficiency. However, these microfluidic approaches either require large quantities of virus or pre-concentration of cells with high-titer viral particles. Here, we describe the development of a microfluidic transduction device (MTD) that combines microfluidic spatial confinement with advective flow through a membrane to efficiently colocalize target cells and virus particles. We demonstrate that the MTD can improve the efficiency of lentiviral transduction for both T-cell and hematopoietic stem-cell (HSC) targets by greater than two fold relative to static controls. Furthermore, transduction saturation in the MTD is reached with only half the virus required to reach saturation under static conditions. Moreover, we show that MTD transduction does not adversely affect cell viability or expansion potential.

Materials promoting viral gene delivery

“…Physical methods such as microinjection, 68,69 microfluidic, 70,71 sonication, 72,73 centrifugation, 74 cellular deformation, 75 laser irradiation 76,77 or electroporation 78,79 induce or facilitate cell entry of vectors by mechanical force, mainly through disrupting the plasma membrane. They are commonly applied in non-viral gene delivery because they transport the genetic information easily without the need for carriers and the otherwise low infectivity.…”

Viral Generation, Packaging, and Transduction on a Digital Microfluidic Platform

“…Thus, this limits the expandability for large-scale clinical settings in gene therapy. Hence, another group from the Biological Microsystems department in Cambridge has developed another microfluidic transduction device (MTD) that colocalizes target cells and lentiviruses together on a semi-permeable membrane to accomplish several-fold increases in transduction efficiency [59] . They achieve this by introducing both cells and viruses through a channel into a chamber where they are pinned against the semipermeable membrane which 'increases the rate of interaction between' both bodies.…”

Scaffold‐Mediated Static Transduction of T Cells for CAR‐T Cell Therapy

“…[18,19] Because of the short distance the retrovirus can travel in solution by Brownian motion (less than 600 µm within one halflife), [20,21] activated T cells and retrovirus must be brought into contact either by spinoculation in the presence of transductionpromoting agents such as retronectin or polybrene or by using microfluidic based transduction devices. [22][23][24] Transductionpromoting agents like polybrene [25] and protamine sulfate [26,27] as well as microfluidic transduction devices have been used with modest success, [28] while recombinant fibronectin fragment CH296 (Retronectin) represents the most widely clinically used transduction-promoting reagent for retrovirus. [29] Retronectin binds retrovirus through its heparin-binding domain and T cells via CS-1/RGD domains, bringing cells and retrovirus together and facilitating gene transfer.…”