Work

Optimization of Human Umbilical Cord-Derived Mesenchymal Stem Cell Growth and Expansion Using a Closed-System Hollow Fiber Bioreactor

Year: 2019

Type: article

Abstract: The Marcus Center for Cellular Cures (MC3) manufactures human umbilical cord (hUC) mesenchymal stem cells (MSCs) for phase I and II clinical trials to treat children with autism and cerebral palsy under Investigational New Drug numbers 17,313 and 18,414. To meet cellular demands of future trials and streamline our current manufacturing process, we are optimizing the use of an automated, closed-system hollow fiber bioreactor to improve our current manual, open-system, flask-based method. We pursued optimal methods for operating the hollow fiber bioreactor to generate MSC yields that were better or comparable to our current Passage 2 (P2) expansion process using Corning HYPERFlasks, and that met all of our potency and release specifications. More than 20 MSC process development runs were completed in the bioreactor. We optimized media/supplements, seeding densities, attachment protocols, feed rates, wash-outs, harvest protocols, and time in the bioreactor. To support these development runs, a proprietary algorithm using lactate production, glucose consumption, and doubling time was developed to use real time bioreactor data to calculate feed rates, predict growth rates, determine day and time of cell harvest, and estimate yields at harvest. Supplementation of P2 expansion medium, xeno-free and serum-free, with human platelet lysate gave a 70% increase in yield. Seeding densities ranging from 45 to 1,300 cells/cm2 were tested, with yields ranging from 800 million to more than 2 billion cells. Population doublings ranged from 5.5 to 10 over the length of the culture. Glucose and lactate concentrations were kept stable by wash-outs and feed rate adjustments. The proprietary algorithm was used to control these parameters, conserve media, and determine harvest time. We determined the optimum seeding density range and medium composition to achieve high cell yields and rapid growth of hUC-MSCs in the hollow fiber bioreactor. Monitoring metabolite levels, using our algorithm, enabled us to maintain healthy cultures and determine harvest time without cell visualization. Cord-to-cord variation in cell growth rates remains a large source of variation in final cell yields. Our development runs have allowed us to begin formal comparability studies with the hollow fiber bioreactor and our current clinical manufacturing process, during which we will assess functionality, potency, and release criteria specifications between the two methods.

Source: Stem Cells Translational Medicine

DOI: 10.1002/sctm.12560

Authors: Lynn A. Cheatham, Pamela Noldner, Barbara Ressler, Joanne Kurtzberg

Institutions: Duke University, Duke Medical Center

Language: English

Topic: Mesenchymal stem cell research

Subfield: Genetics

Field: Medicine

Domain: Health Sciences

SDG: Good health and well-being

Open Access status: gold

Estimated APC paid: $1,750