Tumor Suppressive Monoclonal Antibody Belonging to the VH 7183 Family Directed to the Oncodevelopmental Carbohydrate Antigen on Human Hepatocellular Carcinoma

Abstract

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.

Construction and high cytoplasmic expression of a tumoricidal single-chain antibody against hepatocellular carcinoma

“…In our previous study [1], we found that the viability of HCC-S102 cells was reduced to 85 %, 83 % and 65 % on the first, second, and third day, respectively, after treatment with 5 μg/ml of Hep27 Mab. No significant change could be found in negative control (HCC-S102 treated with Hep16 Mab, an Ab secreted from hybridoma clone without tumuricidal activity).…”

“…Hep27 Mab showed no reactivity to normal liver and low binding activity to HCC-S102 treated with PDMP, an inhibitor of glycolipid synthesis. Furthermore, Hep27 Mab alone can inhibit tumor cell growth [1]. …”

“…Although we did not study how scFv works on the same cells without tumor antigen such as normal liver tissue, our previous study showed Hep27 Mab reacted against HCC-S102, fetal and newborn livers but not against normal adult livers. In addition, the reactivity of Hep27 Mab against HCC-S102 treated with PDMP, an inhibitor of glycolipid synthesis, was decreased compared to untreated cells [1]. Therefore, we assumed that Hep27scFv consisting of a nucleotide sequence encoding for the variable region of its parental antibody (Hep27 Mab) would not recognize normal liver.…”

“…Moreover, as determined by FACS, the binding activity of Hep27 Mab to HCC cells treated with PDMP (1-phenyl-2-decanoylamino-3-morpholino-1-propanol, an inhibitor of glycolipid synthesis) was decreased; indicating that epitope recognized by Hep27 Mab should be a glycolipid. Hep27 Mab alone inhibits the growth of an HCC-S102 cell line (65% viability) without effector cells [1]. The mechanism of tumor suppression is not known exactly, but it may concerns the regulation of tumor cell proliferation by binding Hep27 Mab to the growth factor receptor on the cell.…”

Identification of an Oligopeptide Binding to Hepatocellular Carcinoma

“…For this reason, research aimed at finding gene therapy vectors that could be targeted to HCC cells using specific monoclonal antibodies is being actively conducted[16]. Because monoclonal antibodies such as AF-20 [17] and Hep27 [18] bind to the antigen uniformly expressed in HCC-derived cell lines and human tumors, including those with distant metastasis, it is likely that they would have high specificity and efficiency as HCC-targeting gene transfer vectors. In addition, single-chain antibody fragments may have potential application for tumor targeting [19].…”