Transition from static culture to stirred tank bioreactor for the allogeneic production of therapeutic discogenic cell spheres

Abstract

Background Culturing cells as cell spheres results in a tissue-like environment that drives unique cell phenotypes, making it useful for generating cell populations intended for therapeutic use. Unfortunately, common methods that utilize static suspension culture have limited scalability, making commercialization of such cell therapies challenging. Our team is developing an allogeneic cell therapy for the treatment of lumbar disc degeneration comprised of discogenic cells, which are progenitor cells expanded from human nucleus pulposus cells that are grown in a sphere configuration. Methods We evaluate sphere production in Erlenmeyer, horizontal axis wheel, stirred tank bioreactor, and rocking bag format. We then explore the use of ramped agitation profiles and computational fluid dynamics to overcome obstacles related to cell settling and the undesired impact of mechanical forces on cell characteristics. Finally, we grow discogenic cells in stirred tank reactors (STRs) and test outcomes in vitro (potency via aggrecan production and identity) and in vivo (rabbit model of disc degeneration). Results Computation fluid dynamics were used to model hydrodynamic conditions in STR systems and develop statistically significant correlations to cell attributes including potency (measured by aggrecan production), cell doublings, cell settling, and sphere size. Subsequent model-based optimization and testing resulted in growth of cells with comparable attributes to the original static process, as measured using both in vitro and in vivo models. Maximum shear rate (1/s) was maintained between scales to demonstrate feasibility in a 50 L STR (200-fold scale-up). Conclusions Transition of discogenic cell production from static culture to a stirred-tank bioreactor enables cell sphere production in a scalable format. This work shows significant progress towards establishing a large-scale bioprocess methodology for this novel cell therapy that can be used for other, similar cell therapies.

Allogeneic Disc Progenitor Cells Safely Increase Disc Volume and Improve Pain, Disability, and Quality of Life in Patients With Lumbar Disc Degeneration—Results of an FDA-Approved Biologic Therapy Randomized Clinical Trial

“…This includes the important proteoglycan, aggrecan, which has been measured in a protein assay. 15 Discogenic cells are allogeneic, as they are derived from adult human organ donors. However, we have observed that discogenic cells appear to be immune-privileged.…”

“…Second, discogenic cells lack expression of co-stimulatory molecules CD40, CD80, and CD86, which are required for effector T cell induction. 13,15 The cells do not form tumors in vitro or in vivo. Studies testing human IDCT in animal models of disc degeneration have shown IDCT to be safe and bioactive.…”

“…CD90, and human leukocyte antigen (HLA)-ABC but do not express CD24, CD34, or HLA-DR/DQ/DP. 13,15 This surface marker pattern is tested for and must be present on every batch of discogenic cells. Discogenic cells are multipotent for mesenchymal lineages, including having a strong chondrogenic potential.…”

Emerging perspectives on 3D printed bioreactors for clinical translation of engineered and bioprinted tissue constructs

“…It is important to optimize the hydrodynamic profile (agitation rate) by understanding the effect of maximum shear stress on the cells. 24,25 Although these bioreactors are very helpful in the mass production of therapeutic cells and their products, the major limitation is the inability to support nutrient supply to the core regions of the tissue engineered scaffolds. For example, mass transfer of nutrients to the periphery of the bone scaffold was found to be higher when compared to the core regions.…”

Multiple Cell Cultures for MRI Analysis

“…A work by Rodriguez-Granrose et al demonstrated the transition from static culture to a stirred-tank bioreactor. The study assessed the production of beads in the Erlenmeyer system and of discogenic cells for the development of cell therapy for the treatment of lumbar disc degeneration [ 169 ]. On the other hand, Manstein et al combined STBR with the power of in silico process modeling for the best results in a human pluripotent stem cell (hPSC) culture [ 170 ].…”