title: Tumour-associated tenascin-C isoforms promote breast cancer cell invasion and growth by matrix metalloproteinase-dependent and independent mechanisms.
abstract: BACKGROUND
  The stromal microenvironment has a profound influence on tumour cell behaviour. In tumours, the extracellular matrix (ECM) composition differs from normal tissue and allows novel interactions to influence tumour cell function. The ECM protein tenascin-C (TNC) is frequently up-regulated in breast cancer and we have previously identified two novel isoforms - one containing exon 16 (TNC-16) and one containing exons 14 plus 16 (TNC-14/16).
  
  
  METHODS
  The present study has analysed the functional significance of this altered TNC isoform profile in breast cancer. TNC-16 and TNC-14/16 splice variants were generated using PCR-ligation and over-expressed in breast cancer cells (MCF-7, T47D, MDA-MD-231, MDA-MB-468, GI101) and human fibroblasts. The effects of these variants on tumour cell invasion and proliferation were measured and compared with the effects of the large (TNC-L) and fully spliced small (TNC-S) isoforms.
  
  
  RESULTS
  TNC-16 and TNC-14/16 significantly enhanced tumour cell proliferation (P < 0.05) and invasion, both directly (P < 0.01) and as a response to transfected fibroblast expression (P < 0.05) with this effect being dependent on tumour cell interaction with TNC, because TNC-blocking antibodies abrogated these responses. An analysis of 19 matrix metalloproteinases (MMPs) and tissue inhibitor of matrix metalloproteinases 1 to 4 (TIMP 1 to 4) revealed that TNC up-regulated expression of MMP-13 and TIMP-3 two to four fold relative to vector, and invasion was reduced in the presence of MMP inhibitor GM6001. However, this effect was not isoform-specific but was elicited equally by all TNC isoforms.
  
  
  CONCLUSIONS
  These results demonstrate a dual requirement for TNC and MMP in enhancing breast cancer cell invasion, and identify a significant role for the tumour-associated TNC-16 and TNC-14/16 in promoting tumour invasion, although these isoform-specific effects appear to be mediated through MMP-independent mechanisms.
authors.affiliation: Department of Cancer Studies and Molecular Medicine, Infirmary Close, University of Leicester, Robert Kilpatrick Clinical Sciences Building, Leicester Royal Infirmary, Leicester, UK. raa9@le.ac.uk
authors.forename: Rachael A
authors.initials: RA
authors.lastname: Hancox
authors.forename: Michael D
authors.initials: MD
authors.lastname: Allen
authors.forename: Deborah L
authors.initials: DL
authors.lastname: Holliday
authors.forename: Dylan R
authors.initials: DR
authors.lastname: Edwards
authors.forename: Caroline J
authors.initials: CJ
authors.lastname: Pennington
authors.forename: David S
authors.initials: DS
authors.lastname: Guttery
authors.forename: Jacqueline A
authors.initials: JA
authors.lastname: Shaw
authors.forename: Rosemary A
authors.initials: RA
authors.lastname: Walker
authors.forename: J Howard
authors.initials: JH
authors.lastname: Pringle
authors.forename: J Louise
authors.initials: JL
authors.lastname: Jones
mesh_terms: D017398:Alternative Splicing; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000473:pathology*; D002448:Cell Adhesion; D002465:Cell Movement; D049109:Cell Proliferation; D005260:Female; D006801:Humans; D020782:Matrix Metalloproteinases / Q000235:genetics / Q000378:metabolism*; D009361:Neoplasm Invasiveness; D020033:Protein Isoforms; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019063:Tenascin / Q000502:physiology*; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000235:genetics / Q000378:metabolism; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000235:genetics / Q000378:metabolism; D019717:Tissue Inhibitor of Metalloproteinase-3 / Q000235:genetics / Q000378:metabolism; D019714:Tissue Inhibitor of Metalloproteinases / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation; D000097792:Tissue Inhibitor of Metalloproteinase-4
chemical_list: D020782:Matrix Metalloproteinases; D020033:Protein Isoforms; D012333:RNA, Messenger; D019063:Tenascin; D019715:Tissue Inhibitor of Metalloproteinase-1; D019716:Tissue Inhibitor of Metalloproteinase-2; D019717:Tissue Inhibitor of Metalloproteinase-3; D019714:Tissue Inhibitor of Metalloproteinases; D000097792:Tissue Inhibitor of Metalloproteinase-4; C088390:TIMP3 protein, human