Plant cell suspension cultures present a promising alternative to traditional bioreactor-based systems for the production of recombinant therapeutic proteins. While some currently used platforms lack the ability to perform complex glycosylation or are susceptible to contamination with mammalian pathogens, plant cell cultures offer advantages of biosafety, controllability, reproducibility, conformance to cGMP requirements, and ability to grow in simple, low-cost, chemically defined, animal component-free medium.<br/>Here we demonstrate the use of a transgenic rice cell suspension culture for efficient, semicontinuous production of butyrylcholinesterase (BuChE), a complex and highly glycosylated human enzyme that can function as a bioscavenger against organophosphorus nerve agents. In this platform, the target protein is expressed under the control of a metabolically regulated promoter and tagged with a secretion signal peptide that enables secretion out of the cell. These features enable the use of a semicontinuous operational strategy that allows for independent optimization of growth and production phases, reduces the need for long seed trains, and minimizes turn-around time, CIP and SIP operations, chemicals, and energy.<br/>After design of the gene construct and transformation of the cells, over 300 cell lines from independent transformation events were screened at increasing scale for high growth rate and BuChE expression. After selection of a single cell line, the culture was scaled up to a 5L lab scale bioreactor to determine growth, BuChE production, and sugar consumption kinetics under controlled conditions of temperature, agitation, and dissolved oxygen. These data were then used to determine an efficient operational strategy to maximize volumetric BuChE productivity through multiple cycles of growth and expression. Finally, the experimental data obtained were used to perform preliminary technoeconomic analyses of the process to demonstrate cost-effectiveness of the process at the industrial scale.

Semicontinuous bioreactor production of a recombinant therapeutic protein in metabolically regulated transgenic rice cell cultures

251st National Meeting (2016)

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