Design and Development of a Perfusion Bioreactor with Internal Volume Expansion and Cell Retention for Cell and Gene Therapy Manufacturing

Economically producing patient-specific cell therapies at commercial levels remains a challenge as it requires manufacturing separate batches per patient where the production system must be scaled-out rather than scaled-up to supply more patient doses. The resulting high production cost and consequent price-point can limit patient access. Specifically, the current cell expansion processes are highly manual and laborious, with the number and complexity of operations and components increasing the opportunities for failure and contamination. This work therefore aims to design a mechanically simple bioreactor that is better suited for scale-out by consolidating the cell expansion process to fewer components, vessels, and operations. I propose and develop a novel perfusion bioreactor design with internal culture volume expansion and cell retention capability achieved by a multi-tier airlift reactor design with internal inclined settlers. I developed and employed multiphase and transient computational fluid dynamic (CFD) models in combination with prototype testing to design, characterize, and optimize the bioreactor design. Specifically, I used the CFD models to simulate the gas-liquid flow patterns and predict fluid velocities, pressure, mixing, oxygen mass transfer (kLa), turbulence dissipation, and shear rate for multiple design geometries and aeration rates. I underwent two major phases of design optimization during cell culture prototype testing using CHO cells for the initial proof of concept studies and Jurkat T-cells to demonstrate capability for human cells. First, I resolved flow dynamics at the reactor bottom to establish the geometry and operating conditions for maintaining the cells in suspension and expanding them at rates matching suspension batch cultures. I then corrected the resulting flow disruptions in the settling zone with modifications to reduce momentum transfer from the bulk culture zone. The final design presented herein was a 1L reactor capable of attaining CHO cell densities exceeding 40 million cells/mL with 96% viability and 99% cell retention. Here, the purely pneumatic agitation, passive separation mechanism, and overflow removal of cell-clarified culture enable dynamic perfusion operation without requiring mechanical agitation, outlet pumping, or a separate cell retention unit operation. This design can also be scaled-up for larger volume allogeneic and gene therapy applications