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 (39)LTCSLNDSATEV(50), (36)KILLTCS(42) and (22)AAGTVFTTVEDL(33) 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 E(120)-R(203). Then we constructed and expressed full-length HAb18G/CD147 and truncated HAb18G/CD147 without residues A(22)-V(50) 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 (22)AAGTVFTTVEDLGSKILLTCSLNDSATEV(50) 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.
Ku
X-M
XM
Cell Engineering Research Center & Department of Cell Biology, State Key Laboratory of Cancer Biology, The Fourth Military Medical University, Xi'an, China.
Liao
C-G
CG
Li
Y
Y
Yang
X-M
XM
Yang
B
B
Yao
X-Y
XY
Wang
L
L
Kong
L-M
LM
Zhao
P
P
Chen
Z-N
ZN
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000378:metabolism* / Q000627:therapeutic use*; D000970:Antineoplastic Agents / Q000378:metabolism / Q000627:therapeutic use*; D051926:Basigin / Q000276:immunology* / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000201:enzymology / Q000276:immunology / Q000378:metabolism / Q000628:therapy*; D018604:Epitope Mapping / Q000379:methods*; D000939:Epitopes / Q000276:immunology / Q000378:metabolism; D006801:Humans; D008113:Liver Neoplasms / Q000201:enzymology / Q000276:immunology* / Q000378:metabolism / Q000628:therapy*; D045726:Metalloproteases / Q000037:antagonists & inhibitors / Q000378:metabolism; D008969:Molecular Sequence Data
D000911:Antibodies, Monoclonal; D000970:Antineoplastic Agents; C497886:BSG protein, human; D000939:Epitopes; D051926:Basigin; D045726:Metalloproteases