Integrin α3 β1 : cancer-associated glycans and colon cancer
Prokopishyn, Nicole Lesley
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Colon cancer arises from the gradual accumulation of several genetic and biochemical changes in cells. Ultimately, these changes give cancer cells the ability to spread throughout the body or metastasize. Cancer cells display a variety of alterations to their cell surface carbohydrates. Cell surface glycoconjugates have been implicated in the adhesion, migration and invasion of cells, suggesting that changes to these structures may confer properties necessary for tumor cell metastasis. One such alteration is increased expression of β1-6 branched Asn-linked oligosaccharides on glycoproteins, which has been linked to the metastatic potential of cells. Hybridoma technology was used to generate monoclonal antibodies which detect glycoproteins bearing $\beta 1$-6 branched Asn-linked oligosaccharides which may be important in colon cancer. MAb 3A7 was selected for further study because it detected an epitope expressed at high levels in rat and human colon tumors. In addition, expression of the epitope defined by mAb 3A7 was shown to be developmentally-regulated in rat intestine. Thus, mAb 3A7 detected an oncodevelopmentally-regulated determinant in colon. As well, mAb 3A7 detects a major glycoprotein species of 140 kDa (gp140) which is differentially expressed in human colon cancer cell lines. MAb 3A7 recognizes an epitope containing blood group A (GalNAc$\alpha 1$-3Galβ-) or B (Gal$\alpha 1$-$3Gal\beta$-R) structures exclusively on type 2 chains (Gal$\beta 1$-4GlcNAc). 3A7-immunoreactive gp140 was isolated from the human colon cancer cell line, HT29, by lectin affinity and gel filtration chromatography. Partially purified gp140 was used to generate monoclonal antibodies which detect the polypeptide portion of gp140, namely mAbs 7A8, 7B11, 8C7 and 8H7. Immunological, molecular and biochemical analyses were used to demonstrate that the 3A7-immunoreactive gp140 corresponds to α3β1 integrin, a cell surface adhesion molecule which mediates cell-cell and cell-extracellular matrix interactions. Analysis of $\alpha 3\beta 1$ integrin expression in human colon carcinoma cell lines revealed that this glycoprotein is a major target for the addition of several cancer-associated carbohydrate structures, including $\beta 1$-6 branched Asn-linked oligosaccharides, poly-N-acetyllactosamine (type 2 chain repeats) and the 3A7 epitope. Significantly, the 3A7 epitope appears to be located primarily on the $\beta 1$-6 branch of Asn-linked oligosaccharides on $\alpha 3\beta 1$ integrin. Analysis of a panel of blood group A, AB and B positive human colon carcinoma cell lines revealed that expression of $\alpha 3$ integrin subunit, rather than glycosyltransferase levels, appears to regulate cell surface expression of the 3A7 epitope in colon cancer cell lines. Finally, $\alpha 3\beta 1$ integrin expressed by human colon cancer cells contributes to the adhesion and migration of cells toward extracellular matrix proteins. These data suggest that $\alpha 3\beta 1$ integrin and perhaps its glycan moiety, including the 3A7 epitope, contribute to colon cancer progression.