Expression, glycosylation and function of recombinant anti‐colorectal cancer mAb CO17‐1A in SfSWT4 insect cells

Abstract

Colorectal cancer is a commonly diagnosed cancer in the world. Monoclonal antibody (mAb) CO17‐1A recognizes the tumor‐associated antigen GA733, a cell surface glycoprotein highly expressed in colorectal carcinoma cell, which is considered to be applicable for diagnosis and therapeutic treatment against colorectal cancer. In addition antibodies are glycoproteins, efficiently recognize and eliminate specific pathogenic and disease antigens. We have currently established baculovirus insect cell expression system to produce anti‐colorectal cancer mAb CO17‐1A. In this study, mAb CO17‐1A was expressed in the transgenic insect cell line SfSWT4, in which glycosylation processing pathway has been humanized. Immunoblot confirmed that mAb CO17‐1A properly expressed in SfSWT4 insect cells. mAb CO17‐1A was purified using protein G affinity column. In addition, MALDI‐TOF verified that the mAb CO17‐1A fused to KDEL, endoplasmic reticulum (ER) retention signal (mAb CO17‐1AK) had high mannose type of glycan structure. Migration assay showed that insect cell‐derived mAb CO17‐1AK (mAbI CO17‐1AK) with high mannose type of glycan structure was effective as mammalian‐derived mAb CO17‐1A (mAbM CO17‐1A) in inhibition of metastasis. Kinetic analysis of antigen‐antibody interaction using Surface Plasmon Resonance (SPR) confirmed that mAbI CO17‐1AK is efficient to interact with antigen GA733 as mAbM CO17‐1A. These results suggest that the insect cell expression system with the SfSWT4 possibly can be used as a useful alternative way to produce full‐size mAb for cancer immunotherapy.

Murine response studies of insect cell (Sf9) expressed recombinant colorectal cancer vaccine candidate using surface plasmon resonance studies

“…; Park et al . , ; Lee & Ko ; Qiao et al . ) because insect cells are eukaryotic and they are capable of glycosylation.…”

“…Baculovirus in insect cells can efficiently produce recombinant glycoproteins (Deparis et al 2003;Song et al 2010;Park et al 2011Park et al , 2014Lee & Ko 2014;Qiao et al 2015) because insect cells are eukaryotic and they are capable of glycosylation. Glycan structure affects the biological functions of glycoproteins, including immunogenicity, stability and ligand-receptor interactions (Balen & Krsnik-Rasol 2007;Kamionka 2011).…”

Baculovirus titration method based on MOI values for optimizing recombinant protein expression of the anti‐cancer vaccine candidate GA733‐Fc using Sf9 insect cells

“…The baculovirus expression vector system (BEVS) has been proven useful for the production of recombinant proteins in eukaryotic insect cells (Deparis et al 2003;Song et al 2010;Park et al 2011Park et al , 2014Park et al , 2015Lee & Ko 2014;Kim et al 2015;Qiao et al 2015;Lim et al 2015aLim et al , 2015bLee et al 2016). BEVS has many advantages, including scalability using high-density suspension cultures, capacity for expressing multiple genes at high expression levels, and the possibility of post-translational modifications (Deparis et al 2003;Malde & Hunt 2004;Kost et al, 2005).…”

“…BEVS has many advantages, including scalability using high-density suspension cultures, capacity for expressing multiple genes at high expression levels, and the possibility of post-translational modifications (Deparis et al 2003;Malde & Hunt 2004;Kost et al, 2005). In the present study, we used the BEVS technology to express diverse recombinant proteins, including anti-colorectal cancer monoclonal antibody CO17-1A (Song et al 2010;Park et al 2011Park et al , 2014Lee & Ko 2014 and anti-cancer vaccine candidate GA733-Fc (Kim et al 2015;Park et al 2015;Qiao et al 2015;Lim et al 2015aLim et al , 2015bLee et al 2016).…”

“…Thus, in this study, the Sf9 cell line, which was derived from the parental Spodoptera frugiperda cell line, was used for virus titration (Lee & Ko 2014;Park et al 2014;Qiao et al 2015). We then investigated the effect of the KDEL sequence on the expression of the colorectal cancer vaccine candidate GA733-Fc [a human epithelial cell adhesion molecule (EpCAM)] fused to the human IgG Fc region.…”