Cell and Gene Therapy

In CAR-T cell therapy, autologous T lymphocytes are isolated, genetically modified to express a chimeric antigen receptor and re-infused into the patient. By the end of 2024, more than 30 cell and gene therapy products had been approved by the U.S. FDA, and over 2,200 cell and gene therapy candidates were in clinical or preclinical development worldwide, reflecting the rapid maturation of this therapeutic class.

Lentiviral vectors (LVV) and adeno-associated viral (AAV) vectors are the dominant gene delivery systems. All EMA- and FDA-approved CAR-T products to date rely on lentiviral or gamma-retroviral vectors for stable transgene integration, while AAV vectors enable durable, episomal expression for in vivo gene therapy. Vector production typically uses transient transfection of HEK293 cells in suspension culture.

An emerging class of cell therapies addresses the global blood supply by producing blood cells directly in bioreactors. Cultured red blood cells (cRBCs) generated from CD34⁺ hematopoietic stem cells or induced pluripotent stem cells (iPSCs) have already entered clinical trials — the UK-led RESTORE trial transfused lab-grown allogeneic RBCs into healthy volunteers. Platelets derived from iPSC megakaryocytes have been generated in clinical-scale 8 L turbulence-based stirred bioreactors, producing approximately 100 billion platelets per run.

Messenger RNA (mRNA) therapeutics, formulated in lipid nanoparticles (LNP), have emerged as a flexible platform for vaccines and in vivo cell engineering. mRNA avoids the risk of insertional mutagenesis associated with integrating viral vectors and shifts manufacturing from cell-based viral vector production toward cell-free in vitro transcription, expanding the toolbox of cell and gene therapy modalities.

Autologous therapies (one batch per patient) demand small-scale, highly reproducible processes, while allogeneic, off-the-shelf products — including donor-independent cRBCs, iPSC-derived platelets and universal CAR-NK cells — require scalable expansion of donor- or stem-cell-derived populations in stirred-tank bioreactors. Both modalities depend on tight control of viable cell density, dissolved oxygen, pH and shear stress to preserve cell phenotype, viability and potency.

Unlike protein biologics where glycosylation defines product quality, the critical quality attributes (CQAs) of cell therapies are cellular: viability, identity, purity, transduction efficiency, vector copy number, enucleation rate (for cRBCs), and functional potency such as cytotoxicity for CAR-T or hemostatic activity for platelets. Bioreactor process control directly determines whether these CQAs fall within specification. 

Once considered too shear-sensitive for immune cell culture, T cells have been successfully expanded in stirred-tank bioreactors with controlled agitation. Recent QbD-based studies in 250 mL stirred-tank perfusion systems achieved CAR-T cell densities greater than 21 × 10⁶ cells/mL by initiating perfusion early (48 h post-inoculation) and applying perfusion rates of approximately 1.0 vessel volume per day, without compromising critical quality attributes.

Process intensification through perfusion in 2 L single-use stirred-tank bioreactors has enabled consistent CAR-T cell yields of around 30 × 10⁶ cells/mL, corresponding to over 110 anti-CD19 CAR-T doses per batch within a 7-day expansion process — a clear path toward scalable allogeneic cell therapy manufacturing.

Manufacturing transfusable RBCs ex vivo requires expansion and terminal differentiation of erythroblasts into enucleated reticulocytes at very high cell densities (~10¹¹ cells per unit). Stirred-tank bioreactors with perfusion enable scalable erythroblast expansion from peripheral blood mononuclear cells or iPSCs in feeder-free, GMP-compatible media, while microcarrier-supported and 3D culture systems mimic the bone marrow niche to improve enucleation rates.

Platelet biogenesis in vivo is driven by shear forces in the bone marrow vasculature. Turbulence-controllable stirred bioreactors of 8 L scale have replicated this physical cue ex vivo, releasing platelets from iPSC-derived megakaryocytes at clinical-scale yields of approximately 10¹¹ platelets per run, with hemostatic function comparable to donor platelets — enabling the first in-human transfusion trials of iPSC-derived platelets.

Upstream gene therapy manufacturing is shifting from adherent HEK293 cultures toward high-density suspension cultures with perfusion. Combined with optimised single- and dual-plasmid transfection systems and next-generation transfection reagents, perfusion-based AAV production significantly increases viral titer and productivity per square metre of clean-room space.

Pre-sterilised single-use bioreactors reduce cleaning validation and minimise the risk of cross-contamination between patient batches. Combined with capacitance-based viable cell density sensing, dissolved oxygen and pH monitoring, and AI-supported bioprocess modelling, they enable real-time release strategies and predictive process control — directly addressing one of the largest historical bottlenecks in cell and gene therapy manufacturing: quality control turnaround time.