Effect of plant produced Anti-hIL-6 receptor antibody blockade on pSTAT3 expression in human peripheral blood mononuclear cells

Abstract

As a response to invasion by pathogens, the secretion of interleukin 6 (IL-6) which is a cytokine, activates IL-6/JAKs/STAT3 intracellular signaling via., phosphorylation. Over expression of pSTAT3 induces IL-6 positive feedback loop causing cytokine release syndrome or cytokine storm. Plants have gained momentum as an alternative expression system. Hence, this study aims to produce mAb targeting human IL-6 receptor (hIL-6R) in Nicotiana benthamiana for down regulating its cellular signaling thus, decreasing the expression of pSTAT3. The variable regions of heavy and light chains of anti-hIL-6R mAb were constructed in pBYK2e geminiviral plant expression vector and transiently co-expressed in N. benthamiana. The results demonstrate the proper protein assembly of anti-hIL-6R mAb with highest expression level of 2.24 mg/g FW at 5 dpi, with a yield of 21.4 µg/g FW after purification. The purity and N-glycosylation of plant produced antibody was analyzed, including its specificity to human IL-6 receptor by ELISA. Additionally, we investigated the effect to pSTAT3 expression in human PBMC’s by flow cytometry wherein, the results confirmed lower expression of pSTAT3 with increasing concentrations of plant produced anti-hIL-6R mAb. Although, further in vivo studies are key to unveil the absolute functionality of anti-hIL-6R, we hereby show the potential of the plant platform and its suitability for the production of this therapeutic antibody.