title: Proteomic and postproteomic characterization of keratan sulfate-glycanated isoforms of thyroglobulin and transferrin uniquely elaborated by papillary thyroid carcinomas.
abstract: Previous studies have suggested that surface components of papillary thyroid carcinoma (PTC) cells may be aberrantly glycanated, but the precise nature of these molecules has not been unveiled nor documented to be of clinical relevance. A monoclonal antibody was raised against a unique keratan sulfate (KS) determinant and used to differentially screen benign and malignant thyroid tissue for the expression of components carrying these moieties. In a total of 349 cases of benign and malignant thyroid lesions, 100% of the 115 PTC cases examined (including various histological subtypes) were found to contain KS-bearing molecules, whereas these were virtually absent from benign tissues and other thyroid tumors, with the exception of 21% of the follicular carcinoma cases analyzed. A composite immunoaffinity chromatography, immunochemistry, and mass spectrometric approach revealed that the PTC-specific KS-bearing macromolecules were unique glycoforms of thyroglobulin and transferrin. Combined, reciprocal immunoprecipitation and Western blotting further indicated that the former glycoform predominated and that most of the transferrin produced by PTC was glycanated with KS moieties. Fluorescent keratanase II-based fingerprinting of the KS moieties bound to these isoforms further demonstrated several PTC-specific peculiarities: 1) that a considerable portion of the moieties was covalently attached via a novel core protein linkage structure; 2) they had an unusual extended average length; 3) an unusual relative ratio of highly sulfated disaccharides terminating with alpha (2-3)-linked N-acetylneuraminic acid capping residues; and 4) a novel unidentified oligosaccharide moiety at the nonreducing terminus. Comparative analysis of the relative distribution of transferrin in benign versus PTC tissues highlighted a marked malignancy-associated abundance of the molecule, with a >75% frequency in expression in PTC. These findings demonstrate that PTC cells synthesize unique post-translationally modified thyroglobulin and transferrin variants in situ that may be directly exploitable for diagnosis, through histological and noninvasive cytological procedures; for devising novel strategies for antibody-guided imaging of this tumor in vivo; and for postsurgery follow-up of PTC patients.
authors.affiliation: Department F.G. Ingrassia, Section of Anatomic Pathology, University of Catania, Catania, Italy.
authors.forename: Gaetano
authors.initials: G
authors.lastname: Magro
authors.forename: Daniela
authors.initials: D
authors.lastname: Perissinotto
authors.forename: Monica
authors.initials: M
authors.lastname: Schiappacassi
authors.forename: Steffen
authors.initials: S
authors.lastname: Goletz
authors.forename: Albrecht
authors.initials: A
authors.lastname: Otto
authors.forename: Eva-Christina
authors.initials: EC
authors.lastname: Müller
authors.forename: Michele
authors.initials: M
authors.lastname: Bisceglia
authors.forename: Gavin
authors.initials: G
authors.lastname: Brown
authors.forename: Timothy
authors.initials: T
authors.lastname: Ellis
authors.forename: Sebastiano
authors.initials: S
authors.lastname: Grasso
authors.forename: Alfonso
authors.initials: A
authors.lastname: Colombatti
authors.forename: Roberto
authors.initials: R
authors.lastname: Perris
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000378:metabolism; D002291:Carcinoma, Papillary / Q000378:metabolism* / Q000473:pathology; D000939:Epitopes; D005260:Female; D006801:Humans; D007632:Keratan Sulfate / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D020033:Protein Isoforms / Q000737:chemistry / Q000378:metabolism; D020543:Proteome; D021241:Spectrometry, Mass, Electrospray Ionization; D013954:Thyroglobulin / Q000737:chemistry / Q000378:metabolism*; D013961:Thyroid Gland / Q000166:cytology / Q000378:metabolism; D013964:Thyroid Neoplasms / Q000378:metabolism* / Q000473:pathology; D014168:Transferrin / Q000737:chemistry / Q000378:metabolism*
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D020033:Protein Isoforms; D020543:Proteome; D014168:Transferrin; D013954:Thyroglobulin; D007632:Keratan Sulfate