Objective: To probe better immunization dose and interval of anti-idiotypical minibody (6B11 minibody) in animals and to guide the clinical trials of ovarian epithelial cancer treatment.
Methods: Minibody was induced with isopropylthiogalactoside (IPTG) in E.coli and analyzed with direct enzyme linked immunosorbent assay (ELISA). Normal BALB/c mice were randomly divided into groups A, B, C, D, E, F, immunized with the murine monoclonal anti-idiotypic antibody 6B11 minibody 150 microg, 100 microg, 50 microg, and mice IgG 100 microg, human IgG 100 microg, phosphate buffered saline (PBS) 100 microl at d(1), d(14), d(28) respectively. Anti-anti-idiotypic antibody (Ab3) level was tested and antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) effect of mouse spleen cells to human ovarian carcinoma SKOV3 cells were measured using (51)Cr-release assay. UV absorption values were tested at 490 nm.
Results: 6B11 minibody was successfully expressed on E.coli and could react with a murine monoclonal antibody COC166-9. Ab3 could not be detected until one week after d(14)'s boost, its level gradually elevated and peaked one week after d(28)'s boost, and maintained at high level for about six weeks then declined dramatically seven weeks after d(28)'s boost. When boosted again at seven weeks after d(28)'s boost, the Ab3 level elevated quickly one week after, and maintained at this level at least for two weeks. The Ab3 level of group B was higher than groups A and C, mean A (absorption) value during six weeks after d(28)'s boost was 1.05 in group A, 1.06 in group B, 0.94 in group C, and there was no significant difference (P > 0.05). At four different effector cell/target cell (E/T) ratios, the ADCC was highest in group A and lowest in group C; when the E/T ratio was 1:125, (51)Cr releasing rate was 23% in group A, 17% in group B, and 12% in group C, the difference between groups A, B, C had statistical significance (P < 0.05). When serum was diluted at 1:50, the CDC effects were higher in group A and B than in group C ((51)Cr releasing rate was 47%, 39%, 26% respectively; P < 0.05), but there was no significant difference between groups A and B (P > 0.05).
Conclusion: 6B11 anti-idiotypic minibody of ovarian carcinoma can be used as tumor vaccine, the suitable immunization protocol in animals may be: prime at d(1), d(14) and d(28) at a dose of 100 microg; prime interval is six weeks.