Epitope Mapping of Series of Monoclonal Antibodies Against the Hepatocellular Carcinoma‐associated Antigen HAb18G/CD147

The hepatocellular carcinoma‐associated antigen HAb18G/CD147, a member of CD147 family, could promote tumour invasion and metastasis via inducing the secretion of matrix metalloproteinases (MMP). Anti‐CD147 monoclonal antibodies (MoAb) have exhibited obvious inhibitory effect on MMP induction. However, none of the epitopes of these MoAb has been reported. We previously prepared five MoAb against HAb18G/CD147, named HAb18, 3B3, 1B3, 5A5 and 4D2. To map the epitopes of these MoAb, a series of truncated fragments of extracellular region of HAb18G/CD147 was expressed in Escherichia coli and the MoAb‐binding affinity to these fragments was examined with an enzyme‐linked immunosorbent assay and Western blot. The residues 39LTCSLNDSATEV50, 36KILLTCS42 and 22AAGTVFTTVEDL33 were determined to be the epitopes of HAb18, 3B3 and 1B3, respectively, which were further proved by a dot‐blot analysis with synthesized peptides and bioinformatics epitope prediction. The binding regions of MoAb 5A5 and 4D2 were located at residues E120–R203. Then we constructed and expressed full‐length HAb18G/CD147 and truncated HAb18G/CD147 without residues A22–V50 in COS‐7 cells. Gelatin zymography and Boyden chamber assay showed that the COS‐7 cells expressing truncated HAb18G/CD147 failed to induce MMP production and enhance the cells’ invasive potential, compared with the cells expressing full‐length HAb18G/CD147. Taken together with the obviously inhibitory effects of HAb18 on the function of full‐length HAb18G/CD147, these findings suggest that residues 22AAGTVFTTVEDLGSKILLTCSLNDSATEV50 may play a critical role in the functions of HAb18G/CD147 on MMP secretion and tumour invasion. These key residues can be used as potential drug target in cancer therapy.

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