High-affinity antibodies from hen's-egg yolks against human mannose-6-phosphate/insulin-like growth-factor-II receptor (M6P/IGFII-R): Characterization and potential use in clinical cancer studies

The mannose-6-phosphate/insulin-like growth-factor-11 receptor (M6P/IGFII-R ) involved in trafficking of newly synthesized lysosomal enzymes, degradati on of IGFII and activation of TGF beta 1, was suggested as being coded by a tumor-suppressor gene. No specific antibodies are currently available for clinical studies. Since M6P/IGFII-R is a highly conserved protein in mammal s, we immunized chicken with human M6P/IGFII-R. Up to 200 mg of specific Ig Y from weekly pooled egg yolk was extracted by the polyethylene glycol proc edure. Chicken IgY antibodies specifically recognized the human and bovine 270-kDa M6P/IGFII-R but not the 46-kDa M6P-R, as documented by immunoprecip itation and immunoblotting. Using biosensor analysis, IgY antibodies were s hown to bind M6P/IGFII-R with high affinity (K-D = 7.5 x 10(-9)M). A solid- phase competitive ELISA using bovine M6P/IGFII-R coated on 96-well micropla tes, allowed us to titrate the M6P/IGFII-R in human sera at a sensitivity o f 300 ng/ml, The M6P/IGFII-R was stained by immunoperoxidase in breast- and ovarian-cancer cell lines (T47D, MDA-MB231 , MCF7 and BG1) and in frozen b reast-cancer tissues, showing predominant localization in the trans-Golgi n etwork. Staining specificity was shown with irrelevant IgY and by extinctio n with antigen excess. Quantitative immunohistochemical analysis of frozen sections from 40 invasive breast carcinomas indicated varying levels (from 5 to 400 units) of the M6P/IGFII-R protein which were not correlated with t umor size, histological grade and estrogen receptor or progesterone recepto r. There was a trend (p = 0.08) between lymph-node invasiveness and low rec eptor lever. Moreover, the M6P/IGFII-R lever was significantly tower in can cer cells than in normal cells in 10 out of the 21 tumors in which the peri tumoral normal glands could be quantified in parallel. These 2 last results agree with the hypothesis of a tumor-suppressor gene for this receptor and suggest more basic and clinical studies to prove it. Int. J. Cancer 80:896 -902, 1999. (C) 1999 Wiley-Liss, Inc.