Junji Morimoto

Monoclonal antibodies against CA125-bearing antigenic molecule fragments; reactivity with mucinous ovarian tumours and lung cancers

We first established a cell strain, SHIN-3, from human ovarian serous cystadenocarcinoma, and performed antigen analysis for CA125 which appeared to be massively secreted by the SHIN-3 cells. Protein digestion analysis revealed that a low molecular weight peptide of 49 kDa showed antigen activity. In the present study, we describe a mouse monoclonal antibody against this low molecular peptide, presumed to be part of the CA125-bearing antigenic molecule. Purified antigen prepared from culture supernatant was adsorbed to nitrocellulose membranes and injected intrasplenically in mice. Of the obtained 398 clones, 10 clones were selected by screening in an ELISA test. Of the 10, two were further selected, i.e. SH-9. This hybridoma produces IgG1 monoclonal antibodies. Immunoblotting analysis revealed that SH-9 recognizes the low molecular peptide used as the immunogen. Immunohistological examination with the SH-9 MAb revealed that the antigen reacted with the bronchial epithelium, cervical glands of the uterus and other various normal tissues. Of tumorous tissues, the antibody mainly reacted with ovarian tumours, but positive reactions were also observed with pulmonary adenocarcinomas or squamous cell carcinomas. Surprisingly the positive rate was high in mucinous tumour of the ovary, while no positive reaction was observed in serous tumours. Dot-blot assay using SH-9 revealed that 17/19 (90%) sera of lung cancer patients were positive for the titre suggesting that SH-9 may be useful to set up a serum test for lung cancers.

Establishment and characterization of a new murine mammary tumor cell line, balb/c-MC

SummaryA new murine mammary tumor cell line (BALB/c-MC) was established from a spontaneous mammary tumor in a 17-mo.-old female mouse of the low mammary cancer strain BALB/cHe. The cell line was derived from a papillary adenocarcinoma. In monolayer culture the line exhibits a pavementlike arrangement of cells and forms “domes” or “hemicysts” as the cells become confluent. The cell line rapidly forms tumors when transplanted into young syngeneic BALB/cHe mice. The subcutaneous injection of 106 cells resulted in the development of mammary tumors (typical papillary adenocarcinomas) in 33 of 37 (87% recipients within 2 to 3 mo. after injection. These mammary tumors also metastasize to lung [14 of 33 (42%) of recipients] during this time. The number of chromosomes in this cell line is hyperdiploid (average of 43, range 39 to 44).

HISTOLOGICAL DISTRIBUTION OF MTV ANTIGEN IN MICE DETECTED BY IMMUNO‐PEROXIDASE STAINING

Histological localization of mammary tumor virus (MTV/) antigen was investigated using a variety of organs high (DD/Tbr, SHN, SLN, GR) and low (BALB/c) mammary cancer mice strains and immuno‐peroxidase staining with MTV antigen. Except for BALB/c strain mice, the mammary gland and mammary tumors were generally positive. Accessory male genital organs including the prostate, seminal vesicle, and coagulating gland also demonstrated a positive reaction, but the testis and female genital organs including uterus and ovaries did not. MTV antigen was also revealed in the serous acini of the salivary gland in both sexes. The site of positive reaction in the accessory male sex organs and salivary gland was located in the apical portion of the secretory epithelial cells and their secretory substance. Localization and intensity of antigenic expression of MTV detected histologically were comparable to results obtained by immunodiffusion test and radioimmunoassay. These evidences support the idea that MTV is transmitted horizontally via seminal fluid or saliva.

Polyclonal and monoclonal antibodies monospecific to MMTV LTR orf protein produced in E. coli.

Monoclonal and polyclonal antibodies specific to an open reading frame of the mouse mammary tumor virus long terminal repeat were generated using an open reading frame-beta-galactosidase fusion protein produced in E. coli. Both antibodies reacted with the open reading frame-beta-galactosidase fusion protein but not with beta-galactosidase alone using an immunoblotting technique. It is concluded that these antibodies were specific for the protein encoded by the open reading frame of the mouse mammary tumor virus long terminal repeat.

MORPHOLOGICAL STUDY OF NORMAL AND TUMOR CELLS IN THE HEAD AND NECK REGION USING IRRADIATED COLLAGEN GEL EMBEDDING TECHNIQUE: MORPHOLOGICAL STUDY USING COLLAGEN GEL

Fourteen specimens of normal, benign, or malignant tumor cells from the head and neck region were subjected for culture using 0.2% of irradiated collagen gel embedding technique. Re-differentiation of glands within the gel in serum-free medium was observed. There were marked differences in the growth patterns within the gel between normal or benign and malignant cells. Four normal glands and 5 out of 6 benign tumors showed branch-like growth patterns. On the other hand, malignant tumors showed no branch-like pattern, or could not grow at all. The results showed that the collagen gel technique could be useful for the differential diagnosis of malignancy in head and neck tumors.