Reduced tumor growth in BALB/c mice after CEA mimotope immunization.

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Background and aims: The carcinoembryonic antigen (CEA) is a glycoprotein which is overexpressed on nearly 50% of all human tumors. At present, anti-CEA antibodies are being tested in clinical studies as passive immunotherpeutics. We aimed to establish an active immunotherapy by replacing the poorly immunogenic glycoprotein with highly immunogenic peptides.

Methods: Col-1 is a monoclonal antibody directed against CEA on tumor cells. To generate structural mimics of the Col-1 epitope, we used the Col-1 antibody and the biopanning method in order to select immunreactive peptide mimotopes from random phage libraries. Subsequently, immunogenicity of a selected mimotope was examined in BALB/c mice. We assessed antibody-dependent cytotoxicity (ADDC) and complement-dependent cytotoxicity (CDC) mediated by the induced antibodies on CEA expressing HT29 cells. Furthermore, after immunisation the BALB/c mice were transplanted subcutaneously with Meth-A/CEA tumor cells.

Results: All peptides selected were exclusively recognized by the anti-CEA antibody. In addition, true mimicry of the peptides with the natural antigen was confirmed by competition ELISA using purified human CEA. The best mimotope was chosen for synthesis in a multiple antigenic peptide (MAP) configuration rendering, therefore, a multiple antigenic mimotope (MAM). BALB/c mice immunized with the MAM developed antibodies against the immunogen and the original antigen CEA. In vitro, the anti-mimotope antibodies were capable of inducing high specific lysis of CEA-expressing HT29 cells via antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity. Moreover, after s.c tumor transplantation no tumor growth was seen in MAM immunized mice, compared to the groups receiving a control mimotope or the naïve group. The CEA-mimotope immunized group showed a capsuled necrotic region without tumor cells but granulocytes and limited vascularisation. The non treated group showed tumor cells in different proliferation stages and neovascularisation.

Conclusion: From our results we conclude that the Col-1 epitope of the glycoprotein CEA can be translated into an immunogenic peptide mimic. The mimotope-induced antibodies recognize CEA and do effectively inhibit growth of CEA positive tumors.

Based on these findings we suggest that generated mimotopes are candidates for active immunotherapy of CEA-expressing tumors.