Identification of a Novel Cancer Biomarker
Normal cervical epithelial cells show a restricted GalNAcα1-3Gal localization when stained with antibody 132-3 (dark brown area) (A). In squamous cell carcinomas of the cervix, however, the staining is more intense and spread throughout (B). This staining is specific to this cancer subtype, since in other tumors, GalNAcα1-3Gal cannot be detected (C).
During cancer development, cells accumulate a variety of mutations which alter their normal components and activities. One potential change is in the carbohydrate or sugar polymers which decorate proteins predominately found on the cell surface. The accessibility of these residues makes them ideal targets for the development of diagnostics or therapeutics.
To evaluate differences in normal tissue and tumor-associated carbohydrates, Qian Li, Ph.D., a member of the CCR Laboratory of Medicinal Chemistry, decided to generate specific antibodies to two carbohydrate molecules, GalNAcα1-3Gal and GalNAcα1-6Gal. Many carbohydrates have similar structures, and some previously generated antibodies appear to react non-specifically with multiple residues, confounding any results.
Rabbits were inoculated with the carbohydrate group attached to a scaffold protein to induce an immune response. Antibodies were then collected and purified. To verify the antibodies’ specificities, 163 distinct carbohydrates, including GalNAcα1-3Gal, GalNAcα1-6Gal and similarly structured sugars, were spotted onto a slide, known as a microarray, and exposed to the antibodies. Antibody binding was observed by detection of a fluorescently labeled probe. Two high affinity antibodies, 132-3 to GalNAcα1-3Gal and 74-3 to GalNAcα1-6Gal, were identified.
Binding of these antibodies to normal tissue samples revealed good reactivity between 132-3 and the epithelium of esophagus, larynx, skin and cervix while 74-3 staining was absent and thus, used as a negative control for the remaining studies. As further proof of 132-3’s specificity, its staining pattern failed to overlap with those of antibodies for structurally similar glycans.
Cervical cancer is the leading cause of death among women 21-39 in the US. Currently, cervical tumors are diagnosed by size and depth because no biochemical markers are available. Dr. Li and colleagues investigated whether there were changes in GalNAcα1-3Gal in cervical cancer by examining 132-3 staining. In normal tissue 132-3 was localized to a particular area. However, more widespread and intense staining was observed in a subset of tumors, squamous cell carcinomas, as well as metastatic lymph nodes. Interestingly, patients with increased tumor cell staining had a statistically significant, higher five year survival rate compared with patients whose tumors were low or negative.
By creating specific antibodies, Dr. Li and colleagues demonstrated altered GalNAcα1-3Gal localization and increased expression levels, which correlated with patient survival, in a collection of cervical cancer tumors. This suggests that GalNAcα1-3Gal is a potential diagnostic marker and, with further studies, may serve as a target for therapy in cervical cancer.Summary Posted: 01/2010
Int. J. Cancer 2010 Jan 15;126(2):459-68 PubMed Link