Development of Plant Cell Culture Processes to Produce Natural Product Pharmaceuticals: Characterization, Analysis, and Modeling of Plant Cell Aggregation
Date of Award
Open Access Dissertation
Doctor of Philosophy (PhD)
Susan C. Roberts
Michael A. Henson
Plant derived natural products represent some of the most effective anti-cancer and anti-infectious disease pharmaceuticals available today. However, uncertainty regarding the feasibility of commercial supply due to the limited availability of many plants in nature has resulted in a dramatic reduction in the use of natural products as leads in modern drug discovery. Plant cell suspension culture, consisting of dedifferentiated plant cells grown in vitro and amenable to large scale industrial biotechnology processes, is a production alternative which promises renewable and economical supply of these important drugs. The widespread application of this technology is limited by low product yields, slow growth rates, challenges in scale-up, and above all, variability in these properties, which is poorly understood. Plant cells grow as aggregates in suspension cultures ranging from two to thousands of cells (less than 100 micron to well over 2 mm). Aggregates have long been identified as an important feature of plant cell culture systems, as they create microenvironments for individual cells with respect to nutrient limitations, cell-cell signaling, and applied shear in the in vitro environment. Despite its purported significance, a rigorous engineering analysis of aggregation has remained elusive. In this thesis, aggregation was characterized, analyzed, and modeled in Taxus suspension cultures, which produce the anti-cancer drug paclitaxel. A technique was developed to reliably and routinely measure aggregate size using a Coulter counter. The analysis of aggregate size as a process variable was then used to evaluate the effect of aggregation on process performance, and the analysis of single cells isolated from different sized aggregates was used to understand the effect of aggregation on cellular metabolism and heterogeneity. Process characterization studies indicated that aggregate size changed over a batch cycle as well as from batch to batch, so a population balance equation model was developed to describe and predict these changes in the aggregate size distribution. This multi-scale engineering approach towards understanding plant cell aggregation serves as an important step in the development of rational strategies aimed at controlling the process variability which has heretofore limited the application of plant cell culture technology.
Kolewe, Martin, "Development of Plant Cell Culture Processes to Produce Natural Product Pharmaceuticals: Characterization, Analysis, and Modeling of Plant Cell Aggregation" (2011). Open Access Dissertations. 467.