We established an anti-ErbB-2 mouse-human chimeric monoclonal antibody (MoAb), CH401, which was able to kill cancer cells overexpressing the ErbB-2 protein in vitro. The analysis of the killing mechanism indicated that MoAb CH401 might be the first anti-ErbB-2 mouse-human chimeric MoAb which can induce the apoptosis of cancer cells, since morphological changes and DNA fragmentation were recognized in MoAb CH401-treated cells. The ErbB-2 receptor appears to have two opposing functions: acting as a receptor both for a growth factor and for an apoptotic factor. Our results indicate that MoAb CH401 treatment may prove to be very useful for cancer therapy. Key words:ErbB-2 -Apoptosis -G 1 arrest -Chimeric monoclonal antibody -Growth factor receptorThe human erbB-2 gene product, which encodes a growth factor receptor with intrinsic tyrosine kinase activity, is expressed in a variety of adenocarcinomas.1-5) Many anti-ErbB-2 monoclonal antibodies (MoAbs) have been developed.6-8) ErbB-2 protein has the following advantages as a target molecule: [1] it is a receptor-type molecule expressed on the cancer cell surface; [2] the expression level in normal adult human tissues is very low; [3] the shedding level of this antigen as well as the incidence of positivity for circulating ErbB-2 antigen in gastrointestinal malignant diseases is very low. Accordingly, the blocking effect of circulating antigen on the function of anti-ErbB-2 MoAb should be negligible. 9)Overexpression on cancer cells and extracellular accessibility of the ErbB-2 protein enable it to be a potential target, so anti-ErbB-2 MoAbs seem promising for immunotherapy.Four anti-ErbB-2 mouse MoAbs have been established in our laboratory and characterized.9) The epitopes recognized by these MoAbs are in an extracellular domain within amino acids 292-370 of the human c-erbB-2 molecule. In vitro anti-tumor activity by these MoAbs has been measured and one of them has been selected and chimerized. A mouse-human chimeric MoAb, CH401, for ErbB-2 protein was established by a procedure using a heavy chain loss mouse mutant hybridoma and a human immunoglobulin expression vector, as described elsewhere. 9, 10)Here we show that the anti-tumor activity of this MoAb is due to the induction of apoptosis of cancer cells. MATERIALS AND METHODS MoAbsThe anti-ErbB-2 mouse MoAbs E907 and E401, and the mouse-human chimeric MoAb CH401, each of which recognizes an epitope existing within amino acids 292-370 of the extracellular domain of the human ErbB-2 receptor, 9, 10) were used. These antibodies were of the IgG1 subclass. We developed a mouse-human chimeric form (CH401) of mouse MoAb E401.10) The affinities of mouse MoAb E401 and chimeric MoAb CH401 were similar. The specificity of mouse MoAb E401 and that of chimeric MoAb CH401 have also been confirmed to be the same.10) The epitopes recognized by mouse MoAbs E907 and E401 are different.9, 10) The mouse-human chimeric anti-intercellular adhesion molecule-1 (ICAM-1) MoAb chHA58 of class IgG1 was used as a control.

Anti‐idiotypic antibodies are a new type of useful tools for the possible treatment of cancer patients, since some act as antigen specific immunomodulators. Anti‐idiotypic monoclonal antibody (anti‐Id MAb) D704 (Ab2) was established which bore the internal image of the determinant defined by MAb M2590 (Ab1) against a sialic acid residue on GM3 ganglioside. In an in vivo syngeneic tumor system, anti‐Id MAb D704 was more effective in preventing tumor progression, as compared with anti‐GM3 MAb or no treatment. Significant suppression of tumor growth and prolongation of survival by administration of anti‐Id MAb D704 in an animal group inoculated with 1 × 104/mouse melanoma cells were seen, but not in a group inoculated with 5 × 104 cells/mouse. In an active specific immunotherapy protocol utilizing Ab2, the activity of anti‐anti‐Id antibodies (Ab3) specific for GM3 (antigen) which has a weak immunogenicity only, was maintained for more than 3 months. Ab2 generated cellular anti‐tumor immune responses, including delayed type hypersensitivity (DTH) reaction. Immunohistological analysis indicated a marked infiltration of CD4 and CD8 positive cells into the DTH sites. Our results suggest that internal image bearing anti‐Id MAbs have a therapeutic potential against tumors if the number of melanoma cells is relatively low or if hosts are at an early stage of melanoma progression. Int. J. Cancer 76:345–353, 1998. © 1998 Wiley‐Liss, Inc.

Despite the advent of novel therapies and improvements in survival, multiple myeloma (MM) remains an incurable disease. Thus, new treatment strategies including immunotherapies are needed for MM patients with stable disease after induction chemotherapy as well as for disease control in patients with advanced disease.

In order to analyze the epitope structure of carcinoembryonic antigen (CEA) and the idiotype network system, seven anti-idiotypic monoclonal antibodies (anti-Id MoAbs) were generated from a BALB/c mouse immunized with anti-CEA MoAb P1-356, which recognized a synthetic peptide P1 of CEA, domain I. These MoAbs were divided into four groups. The anti-Id MoAbs specifically reacted with MoAb P1-356, but not with any MoAb, and inhibited the binding of MoAb P1-356 to CEA, indicating that all of these anti-Id MoAbs recognized private idiotopes at the combining sites of MoAb P1-356. Polyclonal anti-anti-idiotypic antiserum (Ab3) generated with anti-Id MoAbs M315 (Ab2), blocked the binding of MoAb P1-356 (Ab1) to CEA and reacted with CEA(Ag) and synthetic peptide P1. The analysis of serological assays suggested that it contains "Ab1-like Ab3." Therefore, we prepared anti-anti-Id MoAbs using anti-Id MoAb M315. Among 13 candidates, anti-anti-Id MoAb 11B2 was selected, because it competed with MoAb P1-356 (Ab1) binding to CEA. A direct binding assay using MoAb11B2 showed that it reacted with purified CEA and with CEA synthetic peptide P1. In addition, MoAb 11B2 (Ab3) reacted with both CEA-producing cultured cells and colonic cancerous tissues in immunostaining. These results indicate that anti-anti-Id MoAb 11B2 is "Ab1-like Ab3." Therefore, it is suggested that anti-Id MoAb M315 bears an "internal image" of the MoAb P1-356-defined epitope on CEA.

It is known that the expression levels of intercellular adhesion molecule‐1 (ICAM‐1) in adult T cell leukemia(ATL) cells are high, whereas those in T‐lymphoid cells are not. In order to investigate the factors that influence the induction of ICAM‐1 molecules, Northern blot analysis to measure the expression level of ICAM‐1 mRNAs and Southern blot hybridization to analyze the integration of human T‐cell‐leukemia virus type 1 (HTLV‐1) provirus were done. The levels of ICAM‐1 mRNA expression of ATL cells were generally higher than those of T‐lymphoid cells. However, ILT‐mat cells and ATL16T(‐) cells, although they were ATL cells, showed rather low surface ICAM‐1 expression and ICAM‐1 mRNA expression. Southern blot hybridization showed that only two and four bands were found in ILT‐mat and ATL16T(‐) cells, respectively, whereas >10 bands were detected in other ATL cells. These results suggest that monoclonal integration of HTLV‐1 provirus to the genome of T cell, especially the number of integration sites, is one of the factors for induction of ICAM‐1 molecules. J. Clin. Lab. Anal. 11:186–189, 1997. © 1997 Wiley‐Liss, Inc.