Construction of scFv derived from a tumor-associated monoclonal antibody having tumoricidal activity on human hepatocellular carcinoma.

A mouse monoclonal antibody (Mab-HepTAA43), classified as an anti-tumor-associated antigen, was raised by immunizing BALB/c mice with the Thai human hepatocellular carcinoma S102 (HCC-S102) cell line cells using hybridoma techniques. The Mab-HepTAA43 reacted with and markedly inhibited the growth of human hepatocellular carcinoma cell lines as well as a tumor mass in an animal model. Human hepatoma transplanted into nude mice did not show metastasis after 20 injections amounting to a total of about 4 mg of the Mab over 1-month period. A single-chain variable fragment (scFv) molecule derived from the Mab was constructed by phage display method. DNA sequence analysis of the active variable regions of both heavy- and light-chains of the cDNA clone was subsequently performed. The scFv43 molecule contains a V(L) kappa type and a unique V(H) sequence having 88% amino acid homology to that of Mab-MAK B raised against tumor-associated antigen. Immunohistochemical staining on frozen sections of paired hepatoma (NCI-I) and normal liver tissue from the same individual showed that both scFv43 and Mab-HepTAA43 antibodies reacted with hepatoma but not with normal liver tissue. The results suggest that scFv43 may be useful in the immunotherapy of hepatocellular carcinoma.

Tumor suppressive monoclonal antibody belonging to the V(H) 7183 family directed to the oncodevelopmental carbohydrate antigen on human hepatocellular carcinoma.

Hep27 monoclonal antibody (Hep27 Mab) was raised by immunizing BALB/c mice with cells of the Thai human hepatocellular carcinoma (HCC) cell line HCC-S102 using hybridoma technology. The Hep27 Mab recognizes oncofetal development antigens by reacting with many HCC, other cancers, fetal and newborn liver but not adult liver. The Hep27 Mab alone markedly inhibits the growth of hepatocellular carcinoma cell lines (65% viability on the third day), suggesting its clinical usefulness. Moreover, complementary DNA (cDNA) for active variable regions of both heavy and light chains of the antibody has been cloned. Sequence analysis of the variable region of the Hep27 Mab revealed that the V(H) and V(L) genes belong to the V(H) 7183 and V(K) families, respectively. We have also characterized the reactivity of the Hep27 Mab to synthetic carbohydrate epitopes and 1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP)-treated HCC-S102 cells. The results showed that the Hep27 Mab recognizes a neoglycolipid containing a mucin core unit and PDMP treatment reduced Hep27 Mab binding activity to HCC-S102 cells, indicating that the Hep27 Mab recognizes a glycolipid antigen on HCC-S102 cells. This Mab may be potentially useful for studying antigenic expression in hepatocellular carcinoma and as a targeting agent for radioimmunodetection and immunoconjugated therapy.