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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
Author(s) -
Lemamy GuyJoseph,
Roger Pascal,
Mani JeanClaude,
Robert Michèle,
Rochefort Henri,
Brouillet JeanPaul
Publication year - 1999
Publication title -
international journal of cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.475
H-Index - 234
eISSN - 1097-0215
pISSN - 0020-7136
DOI - 10.1002/(sici)1097-0215(19990315)80:6<896::aid-ijc16>3.0.co;2-j
Subject(s) - mannose 6 phosphate , antibody , insulin like growth factor 2 receptor , microbiology and biotechnology , immunohistochemistry , growth factor , chemistry , receptor , biology , immunology , biochemistry , insulin like growth factor 1 receptor
The mannose‐6‐phosphate/insulin‐like growth‐factor‐II receptor (M6P/IGFII‐R) involved in trafficking of newly synthesized lysosomal enzymes, degradation of IGFII and activation of TGFβ1, 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 mammals, we immunized chicken with human M6P/IGFII‐R. Up to 200 mg of specific IgY from weekly pooled egg yolk was extracted by the polyethylene glycol procedure. Chicken IgY antibodies specifically recognized the human and bovine 270‐kDa M6P/IGFII‐R but not the 46‐kDa M6P‐R, as documented by immunoprecipitation and immunobloting. Using biosensor analysis, IgY antibodies were shown to bind M6P/IGFII‐R with high affinity (K D = 7.5 × 10 −9 M). A solid‐phase competitive ELISA using bovine M6P/IGFII‐R coated on 96‐well microplates, allowed us to titrate the M6P/IGFII‐R in human sera at a sensitivity of 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 breast‐cancer tissues, showing predominant localization in the trans‐Golgi network. Staining specificity was shown with irrelevant IgY and by extinction 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 tumor size, histological grade and estrogen receptor or progesterone receptor. There was a trend ( p = 0.08) between lymph‐node invasiveness and low receptor level. Moreover, the M6P/IGFII‐R level was significantly lower in cancer cells than in normal cells in 10 out of the 21 tumors in which the peritumoral 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. © 1999 Wiley‐Liss, Inc.