Plant cell suspension cultures have the potential to be a disruptive technology due to their inherent biosafety, controllability, reproducibility, conformance to cGMP requirements, capacity to perform complex glycosylation, and ability to grow in simple, low-cost, chemically defined, animal component-free medium. We have demonstrated use of this alternative expression platform by producing a recombinant form of butyrylcholinesterase (BChE) in a transgenic rice cell suspension culture. BChE is a complex and highly glycosylated human enzyme that has been studied as a therapeutic and prophylactic treatment against organophosphate poisoning and cocaine toxicity. However, widespread use of BChE has been limited because of the difficulty in producing such a large (~340kDa) and complex tetrameric molecule, as well as the prohibitively high costs of its upstream and downstream bioprocessing. The plant cell suspension culture is well-suited to address these specific needs at both the laboratory and industrial scale.<br/>In this transgenic rice cell suspension culture, BChE is expressed under the control of a metabolically regulated promoter along 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 longer seed trains, and minimizes turn-around time, CIP and SIP operations, chemicals, and energy. Our data demonstrate that BChE can be produced through multiple cycles of growth and BChE expression at varying process scales.<br/>We are developing a downstream processing scheme including techno-economic analysis that illustrates how recombinant proteins produced in plant cell suspension culture can be harvested and purified using standard, linearly scalable methods (including tangential flow filtration, DEAE, and affinity chromatography) to obtain high purities, yields and cost effectiveness. We will also present some selected data on characterization of the recombinant BChE and compare it to its native BChE counterparts. These data include the relationship between recombinant BChE activity, temperature, and pH, amino acid sequence coverage, and the structure and abundance of N-glycans.

Integrated bioprocess for production of recombinant butyrylcholinesterase from transgenic rice cell suspension cultures

253rd National Meeting (2017)

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