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Functional analysis of altered Tenascin isoform expression in breast cancer
Breast Cancer Research volume 8, Article number: P20 (2006)
Cellular interactions with the extracellular matrix (ECM) control many aspects of cell function. The complex ECM protein Tenascin-C (TN), which exists as multiple isoforms, is upregulated in breast cancer. We previously have identified a change in the TN isoform profile in breast cancer, with detection of two additional isoforms — TN16 and TN14/16 — not seen in normal breast . The purpose of this study was to investigate directly the effects of these tumour-associated TNC isoforms on breast cancer cell behaviour.
A PCR-ligation approach was used to generate specific TNC isoform sequences which were Flag tagged and inserted into a pCMV vector. Transient transfection into breast cancer cell lines or primary normal fibroblasts was confirmed by RT-PCR, western blotting and immunohistochemistry. The effect of different TNC isoforms on breast cancer cell invasion, proliferation and gene expression was analysed.
Expression of TN16 and TN14/16 in breast cancer cells (MCF-7, T47D, MDAMB231) resulted in significantly enhanced tumour invasion compared with adult-type truncated TN, large TN and vector-only controls. A similar increase in tumour cell proliferation was detected. Coculture of tumour cells with primary breast fibroblasts overexpressing TN16 or TN14/16 or conditioned medium from these fibroblasts also led to enhanced tumour cell invasion. Expression of TN resulted in upregulation of MMP-1; however, this was equivalent for all TN isoforms. The invasion-promoting effect of TN16 and TN14/16 was dependent on direct interaction between tumour cells and was blocked by incorporation of anti-TN blocking antibodies. Furthermore, TN appears to be essential for tumour cell invasion, since with all isoforms invasion was minimal in the presence of anti-TN antibodies.
This study has demonstrated that the tumour-associated TN isoforms TN16 and TN14/16 significantly enhance breast cancer cell invasion and that blocking TN inhibits invasion. We aim to further investigate the invasion-promoting activity of these isoforms and to explore their therapeutic potential in more sophisticated tumour models.
Adams M, Jones JL, Walker RA, Pringle JH, Bell SC: Altered Tenascin-C isoform expression in invasive and pre-invasive breast cancer. Cancer Res. 2002, 62: 3289-3297.
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Alcock, R., Pringle, J., Shaw, J. et al. Functional analysis of altered Tenascin isoform expression in breast cancer. Breast Cancer Res 8 (Suppl 2), P20 (2006). https://doi.org/10.1186/bcr1575