In Vitro and in Vivo Antitumor Effects of Murine Monoclonal Antibody NCC-ST-421 Reacting with Dimeric Lea (Lea/Lea) Epitope1

A murine monoclonal antibody (MAb), NCC-ST-421 (IgG3), was raised by using a human gastric cancer xenograft St-4 as immunogen. Immunization was achieved by transferring immunocompetent normal BALB/c mouse spleen cells into BALB/c-nu/nu mice bearing St-4 tumors. Hybridomas were produced from spleen cells of the mice after rejection of the tumors and were screened for preferential reactivity with cancers on formalin-fixed paraffin sections, as described previously for establish ment of MAb NCC-ST-439 (M. Watanabe et al., Jpn. J. Cancer Res., 76:43-52, 1985). The immunobiological and immunochemical properties of the new MAb NCC-ST-421 are described here. The MAb is essentially directed to a structure with dimeric Le" (V4III4Fuc2LcÂ«Cer)epitope (Gal/31->3|Fucal ->4|GlcNAc/91 ->3Gal/31 ->3|FucÂ«l->4|GlcNAc/31 -Â» 3Gal/31-Â»4Glcj81-Â»lCer). It cross-reacts with Le" but does not show any effect on Le'-positive RBC in vitro or on Le'-positive tissue loci in vivo. ST-421 strongly induced antibody-dependent cellular cytotoxicity using human peripheral blood leukocytes as effector cells with a variety of human tumor cells, using the short-term "'( r release assay. It also showed striking complement-dependent cytotoxicity with a human complement source and was able to produce lysis of a variety of human cancer cell lines, supporting its observed ability to cause cytotoxic suppression of tumor growth in nude mice. In another series of experiments, i.p. injection of ST-421 completely inhibited growth of human tumor xenografts in nut nu mice, and this inhibitory activity was closely dependent on expression of the dimeric Le* antigen on the cell surface. While Le* antigen was expressed in the kidneys of nu/nu mice, infusion of ST-421 in these mice did not cause histolÃ³gica! change in kidney tissue. This finding suggests that the MAb does not damage normal cells or tissues which contain cross-reacting Le*antigen. These results demonstrate that ST-421 exerts a significant antitumor effect in vitro as well as in vivo, does not affect Le* antigen expressed on normal tissues, and therefore has potential application in therapy of certain types of human cancer which express the dimeric Le*antigen. such MAbs may be related to their ADCC and CDC activity. According to previous studies, IgG3 MAbs are partic ularly efficient in suppression of tumor growth in vivo (8) and, in human cancers, mediate ADCC with human effector cells, as well as CDC with a human complement source (4-7). A surprisingly large number of tumor-associated antigens defined by MAbs have been identified as carbohydrate epitopes, carried by either proteins (glycoproteins) or sphingolipids (glycosphin-golipids) (9). Watanabe, Hirohashi, and associates previously developed a novel procedure to establish mouse MAb ST-4-39, using a human gastric cancer xenograft grown in nude mice as immunogen (10). Immunization was achieved by transferring immunocompetent mouse spleen cells into nude mice bearing human tumors, and hybridomas were produced by standard technique and screened by immunohistological reactivity. In the present study, an identical procedure was used to obtain a new MAb (NCC-ST-421) which belongs to the IgG3 subclass, recognizes a novel dimeric Lea antigen (Ref. 11; see Table 1), and shows ADCC and CDC in vitro and in vivo. In this paper, we describe the immunochemical and immunobiological prop erties of ST-421 and its effect on tumors and on normal tissues where cross-reacting Lea is present. immunohistochemical an HistolÃ³gica! normal