Expression of Colorectal Cancer Antigenic Protein Fused to IgM Fc in Chinese Cabbage (Brassica rapa)

Abstract

The epithelial cell adhesion molecule (EpCAM) is a tumor-associated antigen and a potential target for tumor vaccine. The EpCAM is a cell-surface glycoprotein highly expressed in colorectal carcinomas. The objective of the present study is to develop an edible vaccine system through Agrobacterium-mediated transformation in Chinese cabbage (Brassica rapa). For the transformation, two plant expression vectors containing genes encoding for the EpCAM recombinant protein along with the fragment crystallizable (Fc) region of immunoglobulin M (IgM) and Joining (J)-chain tagged with the KDEL endoplasmic reticulum retention motif (J-chain K) were constructed. The vectors were successfully transformed and expressed in the Chinese cabbage individually using Agrobacterium. The transgenic Chinese cabbages were screened using genomic polymerase chain reaction (PCR) in T-0 transgenic plant lines generated from both transformants. Similarly, the immunoblot analysis revealed the expression of recombinant proteins in the transformants. Further, the T-1 transgenic plants were generated by selfing the transgenic plants (T-0) carrying EpCAM-IgM Fc and J-chain K proteins, respectively. Subsequently, the T-1 plants generated from EpCAM-IgM Fc and J-chain K transformants were crossed to generate F-1 plants carrying both transgenes. The presence of both transgenes was validated using PCR in the F-1 plants. In addition, the expression of Chinese cabbage-derived EpCAM-IgM Fc x J-chain K was evaluated using immunoblot and ELISA analyses in the F-1 plants. The outcomes of the present study can be utilized for the development of a potential anti-cancer vaccine candidate using Chinese cabbage.