(69c) The Effect of Mechanical Shear On the Growth and Aggregation Dynamics of Taxus Suspension Cultures

Paclitaxel, derived from the Taxus Pacific Yew Tree, is a mitotic inhibitor used in cancer chemotherapeutics. Currently, paclitaxel is being produced using plant cell suspension cultures, but the process is limited by low and variable product yields. One possible cause for this variability is the tendency of plant cells to grow in a range of aggregates sizes, creating subpopulations throughout the culture due to varying oxygen and nutrient diffusion profiles. The Roberts’ lab has found that cultures with smaller aggregate sizes have increased paclitaxel production levels. As a result, this project aims to investigate the effect of mechanical shearing on growth, viability and aggregation of Taxus suspension cultures. Different shearing methods (pipetting and vortexing) were investigated in order to determine the most efficient and effective way to break aggregates. Manual pipetting of cultures through a glass pipet was found to be the most effective method to decrease the aggregate size. The aperture tip, where most of the breakage occurs, plays an important role in determining the final culture aggregate size. Along with the aperture tip, the flow rate of pipetting affects the rate of particle breakage and determines the terminal diameter of the aggregates. The culture density had no effect on determining the terminal particle size when sheared through pipetting, which is important for shearing of cultures throughout a growth cycle. The effect of mechanical shear was then investigated on the culture growth and aggregation over multiple generations. The culture mean aggregate size was effectively reduced without inhibiting culture growth over three generations. Controlling aggregate size through mechanical shearing by pipetting could lead to cultures with reduced variability and increased paclitaxel production levels.