Introduction
Growing global demand and public spotlight on the biopharmaceutical industry is driving increased importance on production costs. This spotlight also exacerbates the importance of viral contamination control (Aranha, 2011). These external pressures position the industry to consider alternatives to microbial fermentation and mammalian cell culture production systems.
Plant cell suspension cultures have demonstrated promise as an alternative. Plant cells are higher eukaryotes, able to produce a wide array of complex protein products through a versatile set of expression and processing techniques (Huang & McDonald, 2012; Nandi & McDonald, 2014). Plant cell cultures are relatively inexpensive to operate due to their simple, often chemically-defined culture medium free from animal-derived components (Häkkinen et al., 2018). They have been used at the commercial manufacturing scale for production of multiple drug products, including the secondary metabolite paclitaxel (Tabata, 2006) and the recombinant human enzyme glucocerebrosidase produced by Protalix Biotherapeutics (Ratner, 2010). Currently, Protalix is the only company with an FDA approved recombinant biologic produced in plant cell suspension culture (Tekoah et al., 2015), and they have several more products in clinical development (Almon et al., 2017; Schiffmann et al., 2019). Protalix’s process, which has paved the way for regulatory approval of this technology, serves as an excellent guide for design of future plant cell culture processes.
We have recently demonstrated the promise of plant cell culture technology for production of a challenging recombinant human therapeutic, the human enzyme butyrylcholinesterase (BChE). BChE is a bioscavenger agent that protects against organophosphorus compounds that are used in chemical warfare and also used as agricultural pesticides. The previously reported cell culture system is able to produce BChE in a metabolically-regulated transgenic rice culture (referred to as rice recombinant BChE or rrBChE) over multiple cycles in a stirred tank bioreactor (Corbin et al., 2016) and can operate semicontinuously for >6 months with no decrease in the rrBChE production (unpublished data). Using a combination of scalable, commonly used operations including tangential flow filtration and column chromatography, rrBChE can be purified to >95% with a 41% overall process recovery at laboratory scale. Furthermore, rrBChE has shown comparable structure, activity, and in vitroorganophosphate inhibition efficacy to native human BChE (hBChE) (Corbin et al., 2018). These factors indicate that manufacturing-scale implementation of this technology could lead to effective and affordable production of this important drug.
Despite the promise of plant cell cultures for biopharmaceutical production and their demonstrated efficacy and ease of use by Protalix, manufacturing scale use of these cultures has been limited. Due to the high cost of entry into the pharmaceutical manufacturing business, novel processes are often viewed as too risky for development. To mitigate risk associated with adoption of a new process, risk severity and probability must both be considered.
Techno-economic analysis is one method to reduce economic uncertainty of manufacturing costs and gauge risks. It can also be helpful to assess process operation strategies and predict theoretical costs to identify process and economic parameters with the highest impact on manufacturing costs. This can be done using “back-of-the-envelope” calculations, spreadsheets, computer modeling, and simulation tools such as SuperPro Designer® (Petrides, Carmichael, Siletti, & Koulouris, 2014).
Several traditional biopharmaceutical manufacturing processes have been studied using SuperPro Designer® and other process simulation tools, including tissue plasminogen activator (Rouf, Douglas, Moo-Young, & Scharer, 2001; Rouf, Moo-Young, Scharer, & Douglas, 2000) and monoclonal antibody (Xenopoulos, 2015) production in transgenic mammalian cells. Other studies have focused on whole plant-based biopharmaceutical processes, including lactoferrin (Nandi et al., 2005) and lysozyme production in transgenic rice (Wilken & Nikolov, 2012), and transient expression of monoclonal antibody (Mir‐Artigues et al., 2019; Nandi et al., 2016), recombinant BChE (Tusé, Tu, & McDonald, 2014), antimicrobial proteins (McNulty et al., 2019), and Griffithsin (Alam et al., 2018) in Nicotiana benthamiana plants. These studies suggest that plant-based protein expression can produce high quality recombinant proteins with a substantial cost savings, though the magnitude of this savings depends on the specific molecule, as well as the production and processing system.
However, to our knowledge, no such analyses have been performed for a plant cell culture-based biomanufacturing process. In this work, we present a techno-economic model, simulation, and analysis of a large-scale version of the process our group has developed for semicontinuous production of rrBChE in rice cell suspension culture. Our design inputs draw from laboratory-scale process data we have generated and demonstrate the potential cost savings that can be obtained by implementing this process for production of a challenging human biopharmaceutical. The base case facility is designed to produce 25kg of purified rrBChE/year at >95% purity as bulk drug substance with single-use bioreactors used in the seed train and stainless steel bioreactors used for production. The rrBChE was assumed to be cell-associated, extracted from the rice biomass, and purified using tangential flow filtration and chromatographic operations. An additional goal of this model development is to create a tool that can be easily modified, adapted, and broadly applicable to similar processes. To the best of our knowledge, this work represents the first techno-economic analysis reported for production of recombinant protein in plant cell culture and the first facility simulation model for semicontinuous bioreactor operation over long time frames (~6 months). We believe this analysis can be considered as a general model, and the simulation tool can be used for widespread evaluation of semicontinuously-operated cell culture platforms for production of moderate-volume biopharmaceutical products.