Rethinking Cell Engineering for Cell Therapy Manufacturing

Rethinking Cell Engineering for Cell Therapy Manufacturing

December 13, 2023PAO-11-23-CL-03

Novel microfluidics cell engineering technology from CellFE provides superior delivery of the largest range of complex payloads to cells for cell therapy development from research scale to commercial manufacturing. The rapid nature of the process, which is readily scalable, reduces cell therapy vein-to-vein manufacturing time and leads to improved cell health and final product quality.

Increasing Access to Cell Therapies

Despite the tremendous transformative potential of cell therapies, only a fraction of the patients who could benefit have access to these lifesaving therapies. This is a consequence of several challenges, including the limited number of global locations that provide cell therapies, the long vein-to-vein time associated with manufacturing, and the high costs of manufacturing.

Driven by a motivation to expand access to these curative therapies, CellFE addresses the latter two issues through manufacturing innovations using its microfluidic cellular engineering technology. The CellFE platform offers the following advantages over existing technologies: gentle processing of cells, rapid cell recovery that enables complex gene-editing with high transfected cell yields, and a simple process that leads to higher quality of genetically engineered cells that are critical to robust and durable therapies.

From Academic Studies to Commercial Products

CellFE began as an academic endeavor to explore cell mechanical responses to large deformations occurring within short timescales. Our studies led to a foundational hypothesis that cells could withstand large deformations for very short periods of time with no impact on the biology. While investigating this hypothesis, we discovered a practical application of delivering large and complex payloads into cells for engineering cell therapies.

Delivering a payload into a cell requires two things: a pore in the cell membrane (i.e., a transient and reversible permeabilization) and a driving force for transporting the payload into the cell interior. The most common approaches for delivering molecules into cells for cell therapy are the use of viral vectors and electroporation. Viral vectors are the predominant platform for integrating a functional transgene into the cell, though their use results in high manufacturing costs, cumbersome logistics, and risks associated with random genetic integration. Electroporation creates pores in the cell membrane by applying repeated electrical fields, after which molecules passively diffuse or are electrophoresed into cells before the temporal pores reseal. Exposure of cells to repeated electrical pulses can cause irreversible damage to cells that, along with other harmful effects, hinder cell proliferation. With the CellFE technology, my research team found that, when cells are compressed abruptly at short timescales (<1 milliseconds), they act much like sponges in that they quickly reduce in volume and return to their original shape and thereby actively transport exogenous material into their interior. Anything contained in the cell medium is carried by this flow into the cells, providing a powerful mechanism to deliver larger, more complex gene-editing payloads.

Several applications of this technology have been investigated. With medical partners at Emory University, we found chimeric antigen receptor (CAR)-T cells could be generated, and at greater yield, a result that indicated distinct and improved outcomes from other delivery methods. In addition to the delivery of gene-editing payloads, we also studied the delivery of nanoparticles and other modifications to cells. With medical partners from Stanford University, we found that high concentrations of iron oxide nanoparticle payloads could be delivered to T cells to study the homing of CAR-T cells to the tumor in vivo. This preliminary work helped us elucidate the key features needed for an effective device that provides the best results for cell engineering. Recognizing the commercial potential of this technology to impact patients’ lives, CellFE was founded.

A Brief Overview of Microfluidics

Microfluidics refers to the control of fluids at small scales within microchannels. It is an ideal technology for achieving fine control over forces applied to cells within liquids. With the CellFE microfluidic technology, results obtained with a one-channel scenario can be quickly scaled to many channels without further need for optimization. This unlocks a range of possibilities for a rapid and seamless transition from laboratory experiments to clinical, large-volume applications. The key to success is appropriate, fit-for-purpose engineering of microfluidic devices. We believe this unparalleled seamless scalability is a game-changer as it will provide significant time and cost efficiencies in development and manufacturing of cell therapies.

CellFE’s Solution for Cell Therapy

CellFE’s technology offers a solution that optimizes delivery and cell health, representing an unmet need in the cell therapy field. Within the microfluidic chamber, cells are compressed via a sudden constriction, which induces a temporary decrease in volume, followed by their re-expansion. This results in a transfer of surrounding payload into the cells. This brief compression can also be designed to minimize perturbation to the nucleus, which could result in unwanted cell damage. In addition, throughput is increased by allowing the cells to move in a plane to pass through the constriction in parallel without being forced into a single-file flow leading to cellular roadblocks.

The unique design of CellFE’s technology has been used to deliver large DNA templates that are 15 kilobases in size into cells — approximately twice the size of a payload that a virus can carry. In one example, our partner at the University of Iowa used CellFE’s technology to simultaneously deliver a >10-kb DNA construct and CRISPR/Cas9 reagents to perform homology-directed repair (HDR) in patient-derived induced pluripotent stem cells (iPSCs) to correct a mutation of an inherited disease that causes blindness, recently published as a first-in-microfluidics application.

Key Performance Differentiators

CellFE’s technology offers several key advantages over other delivery methods, such as viral vectors, electroporation, lipofection, or other microfluidic-based approaches. One key differentiator is the simplicity of the process. Cells do not need to be placed in a special medium or buffer; they can be taken directly from cell culture and placed into the transfection device along with the payload. A second key differentiator is the high proliferative viability of the cells so that they can readily grow and proliferate following transfection or even undergo multiple editing events sequentially, opening new possibilities for performing complex edits.

A full appreciation of the benefits of the CellFE platform may be contingent on which metrics are prioritized to assess manufacturing outcomes. Conventionally, developers remain focused on legacy metrics that can be easily measured immediately after processing, including transfection efficiency and necrotic viability. However, such metrics may not be the best indicators of the yield or quality (persistence, potency) of the cells, which ultimately drive product manufacturing efficiencies. For instance, a high transfection efficiency that results in the majority of the cells being lost or apoptotic, and thus failing to expand, negates any benefits of high transfection efficiencies. In contrast, the advantages of CellFE’s microfluidic technology are clear when viewed through the lens of metrics pertinent to therapeutic success. A CAR-T researcher recently compared the CellFE platform to electroporation. This researcher was able to use the CellFE platform to generate over six-fold more CAR-T cells five days after editing, with further increases in yield observed during the 12-day monitoring period. Stable integration of a CAR transgene without using virus and with robust cell growth post-editing can open new paths to rapid manufacturing workflows.

Importantly, CellFE’s microfluidics technology has the potential to optimize the potency and durability of cell therapies by unlocking higher cell yields, enabling shorter manufacturing processes than currently required, and lowering the vein-to-vein time. Many CAR-T cell therapy manufacturing times are measured in many days or weeks, risking further decline in the patient’s health during that waiting period. Moreover, the quality of the manufactured cells plummets with long ex vivo expansion. Researchers at the University of Pennsylvania and elsewhere have found that reduced culture time leads to a higher percentage of stem memory (versus effector) T cells, among the best predictors of cell therapy success to blood cancers. CellFE’s technology shortens the current manufacturing timeline by preventing the damage caused by conventional methods that hinders edited cells’ ability to expand and grow. The impact is a greater therapy potency and durability, which further underscores the unmet need to improve cell manufacturing.

Addressing Scalability

Another challenge posed by cell therapies is the need to engineer cells with a wide range of production scales throughout process development and clinical and commercial manufacturing. Some cell therapies, such as CAR-T cell therapies, are personalized medicines in which one batch must be produced for each patient. Newer CAR-T treatments might involve clinical doses of just 10 million edited cells, while more typical products comprise a few hundred million or up to a billion edited cells. Production batches for allogeneic cell therapies require even greater numbers of cells — several billion edited cells.

CellFE’s technology is designed for simple scalability — scaling is elegantly achieved by only increasing the number of channels. Most importantly, it is engineered to ensure that all cells move through the channels in an identical manner to ensure the highest-quality results. As CellFE’s technology becomes more automated and integrated with other operations, reduced costs will also follow. The simplicity of operation of the microfluidics solution will facilitate a more distributed manufacturing approach, in which manufacturing facilities need not be centralized but can be located closer to the point of care, another key step in expanding access to cell therapies.

Future Vision—Focusing on the State of the Cell

CellFE’s technology is transformative because its manufacturing workflow results in rapid editing of cells delivering exceptional quality and high proliferative yield. We are continuing to develop innovative and powerful workflows, including complex editing of cells in a manner that retains cell quality while exhibiting lower genotoxicity and fewer risks of chromosomal abnormalities. The CellFE technology allows new ways of performing multiple edits, with each edit performed at a different timepoint and avoiding negative impacts to cell quality. Since gene editing requires the introduction of double-strand DNA breaks, co-existing breaks introduce the possibility of unintended genetic rearrangements or translocations with mutagenic risks and additional toxicities to the cell. CellFE’s technology reduces the risk of translocations by providing optionality to when those edits occur, making it an attractive solution for manufacturers developing allogenic cell therapies or cell therapies targeting solid tumors, both of which require multiple edits.

The long-term vision for CellFE is to become the backbone of next-generation cell therapies by enabling higher-quality genetic-engineered cells that result in more robust and durable therapies.

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