**3. Production: mammalian cells, plants and beyond**

Production platforms for monoclonal antibodies determine the cost effectiveness and hence their viability as a therapeutic product. Due to the need to transcribe and assemble 4 components (IgA heavy & light chain, J chain and secretory component), SIgA production is a complex multi-step process which has been attempted in different protein production systems.

Mammalian cells, specifically CHO (Chinese hamster ovary cells), are the industry standard in therapeutic IgG monoclonal antibody production. SIgA manufacture has been achieved either through multiple gene transfection or in vitro reconstitution (ie. dimeric IgA incubated with secretory component) [22]. However, the technology is associated with very high production costs which is reflected in the expensive price of monoclonal antibody therapy, even for IgGs [31]. This problem is exacerbated for SIgA as the yields in mammalian cells are still low.

Plants have emerged as an attractive alternative platform for SIgA production. Plant-derived therapeutics are coming of age—previously against Ebola, Gaucher's disease and more recently the plant-based SARS-CoV-2 vaccine Covifenz® has been approved for use by Health Canada [32–35]. Plants are well suited to produce SIgA by expressing the four components either transiently or by the sequential crossing of plant lines stably expressing each component [36]. A plant-specific issue, however, is the apparent cleavage of the IgA Fc tailpiece required for J chain incorporation which may be due to differential glycosylation in plants [37]. Efforts to circumvent this include the co-expression of the N-glycosylation facilitating enzyme oligosaccharyltransferase from *Leishmania major* [38].

Furthermore, plant-associated glycans (non-human modifications) on recombinant antibodies present a challenge to therapeutic advancement. For example, plants lack branched and sialylated N-glycosylation, and produce plant-specific xylose and fucose residues [39]. Plant O-glycosylation also differs from humans with the presence of complex arabinogalactans on hydroxyproline residues (extensively found on the IgA1 long hinge [40]) and galactosylated serine which does not appear in humans [41]. This is being addressed by using glycoengineered plant lines—these can produce antibodies which function as well as a their CHO-produced counterpart, but with more controlled and homogenous glycan profiles [42, 43].

Overall, SIgA production in different production platforms carry specific advantages and disadvantages—the overarching issue being the production of multiple assembly intermediates (single chains, monomers and dimers), and it is troublesome to isolate the desired fully assembled SIgA complex [16, 44]. In order to increase the therapeutic potential of SIgA, it is necessary to increase yield (expression levels) and optimise downstream processing.
