|Budget Amount *help
¥2,100,000 (Direct Cost : ¥2,100,000)
Fiscal Year 1997 : ¥800,000 (Direct Cost : ¥800,000)
Fiscal Year 1996 : ¥1,300,000 (Direct Cost : ¥1,300,000)
A murine monoclonal antibody MAbG250 is known to react with the majority of renal cell carcinomas (RCC) whereas reactivity with normal tissues is restricted to gastric mucosal cells and large blle duct cells. Recently, the G250 antigen has been identified (Oosterwijk et al.1995). Sequence analysis and database searching revealed that G250 antigen is identical to MN,a human tumor-associated antigen identified in cervical carcinoma (Pastorek et al, 1994). This antigen (G250, MN) is a transmembrane glycoprotein of 54 kDa and detectable in several types of malignancies but not in corresponding normal tissues. In the present study, we investigate MN/G250 expression with immunohistochemistry and RT-PCR in urologic cancers. MATERIAL & METHODS : Frozen specimens of bladder carcinoma (BT)(n=61), prostate carcinoma (PCa)(n=46), RCC (n=147) and germ cell tumor (GCT)(n=19)were studied for MN/G250 antigen peptide and gene expression. In addition, 17 RCC cell lines and 8 bladder carcinoma cell lines
, and 3 PCa cell lines were studied. Cell lines were tested for MN/G250 expression by MHA and RT-PCR analysis with primers derived from cDNA sequence of MN/G250 antigen. RESULTS : In RCC,128/147 (87%) tissue specimens showed strong and homogeneous MN/G250 expression, in accordance with previous studies. Tumor grade and cell type appeared to correlate inversely with MN/G250 expression, i.e., high grade and sarcomatoid type showed significantly low expression as compared to the others. 6 of 17 (35%) RCC cell lines were G250 positive. In bladder carcinoma, 20/61(33%)primary tumors showed MN/G250 expression heterogeneously and 1/8(13%)cell lines were G250 positive. In contrast, no G250 expression was observed in PCa and GCT specimens and the other normal normal tissues.
Based, on these results describe above, we cariied out therapeutic experiments in mice with RCC xenograft. Briefly, small pieces of NUR-2 RCC were transplanted into the right flank of male nu/nu BALB/c mice. A couple of weeks after transplantation, mice were randomized and divided into each groups (n=6-7). Mice carrying NUR-2 human RCC xenografts (MN/G250^+,20mm^3) were treated by peri-tumor injection of MAbG250 and/or cytokines (m-IFN/IL-2/MCSF) or 0.2 ml of Ab3 sera with/without MCSF.Mice were treated 5 times a week for 6 weeks. Control mice were treated with 0.1 ml medium alone (RPMl 1640) or sera from MOPC21 immunized mice (Ab3-MOPC). The tumor-take rates and tumor growth were determined every week. RESULTS : All mice showed 100% tumor take after 4 weeks. However, the tumor volume in IFN or IL-2 therapy as well as MAbG250 treated animals were significantly lower as compared to control animals (p<0.01). Treatment with MCSF resulted in tumor growth inhibition but not significant. Combination of MAbG250 with cytokines resulted in increased anti-tumor effects as compared to MAbG250 or cytokine monotherapy. IL-2/IFN/MAbG250 therapy showed significant tumor growth Inhibition as compared to MAbG250 or cytokine monotherapy (p<0.05), however the other combination therapy were not significant. Moreover, Ab3-based (Ab2-induced) immunotherapy resulted in tremendous tumor growth inhibition as compared to MAbG250 or the other cytokine combination therapies (p<0.001).
CONCLUSIONS : Our findings suggest that MN/G250 antigen may be potential therapeutic target for RCC immunotherapy. Less