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Glycomodification and characterization of anti-colorectal cancer immunotherapeutic monoclonal antibodies in transgenic tobacco

Abstract

Anti-colorectal cancer mAb CO17-1A (IgG2a) recognizes the antigen GA733, which is highly expressed on the surface membrane of human colorectal carcinoma cells. In this study, a transgenic tobacco system for the production of mAb CO17-1A was developed. The mAb construct included a KDEL sequence, an endoplasmic reticulum (ER) retention signal attached to the C-terminus of the heavy chain, to target accumulation of mAb into ER. An immunoblot showed significantly enhanced levels of expression of the plant-derived mAbK (mAbPK) CO17-1A compared to mAbP CO17-1A mAb without the KDEL sequence. An ELISA assay using human colorectal carcinoma cells confirmed that expression of mAbPK was also significantly higher than that of mAbP. Glycosylation analysis revealed that mAbP had plant-specific glycans; whereas, mAbPK primarily had oligomannose glycans. FACS showed that the Fc domains of both mAbPK and mammalian-derived mAb (mAbM) had similar binding activity to the FcγRI receptor (CD64). However, the Fc domains of the mAbP had slightly lower binding activity to the FcγRI receptor than both mAbPK and mAbM. The antibody-dependent cell cytotoxicity of mAbPK, against human colorectal cancer cells, was as efficient as mAbM; whereas mAbP was very low. These results suggest that KDEL localized and accumulated mAbP in the ER and eventually enhanced the expression of mAbP with oligomannose glycan and similar anti-cancer biological activity to the parental mAbM.

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Abbreviations

ADCC:

Antibody-dependent cell cytotoxicity

ER:

Endoplasmic reticulum

FACS:

Fluorescense activated cell sorting

mAb:

Monoclonal antibody

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Acknowledgments

The authors wish to thank Dr. Hilary Koprowski for suggestions and expertise. This research was supported by a grant from the Korean Rural Development Administration (Code# PJ007492); Korea Institute of Planning and Evalution for Technology of Food, Agriculture, Forestry and Fisheries (iPET-Code# 111096-03-1-SB010); the Next-Generation BioGreen 21 Program (SSAC, grant# PJ00906), Rural Development Administration.

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Correspondence to Kisung Ko.

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So, Y., Lee, KJ., Kim, DS. et al. Glycomodification and characterization of anti-colorectal cancer immunotherapeutic monoclonal antibodies in transgenic tobacco. Plant Cell Tiss Organ Cult 113, 41–49 (2013). https://doi.org/10.1007/s11240-012-0249-z

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Keywords

  • ADCC
  • Anti-cancer antibody
  • Glycosylation
  • Oligomannose
  • Transgenic plant
  • Human colorectal carcinoma