Loss of MMP-8 in ductal carcinoma in situ (DCIS)-associated myoepithelial cells contributes to tumour promotion through altered adhesive and proteolytic function.
Normal myoepithelial cells (MECs) play an important tumour-suppressor role in the breast but display an altered phenotype in ductal carcinoma in situ (DCIS), gaining tumour-promoter functions. Matrix metalloproteinase-8 (MMP-8) is expressed by normal MECs but is lost in DCIS. This study investigated the function of MMP-8 in MECs and the impact of its loss in DCIS.



Primary normal and DCIS-associated MECs, and normal (N-1089) and DCIS-modified myoepithelial (β6-1089) cell lines, were used to assess MMP-8 expression and function. β6-1089 lacking MMP-8 were transfected with MMP-8 WT and catalytically inactive MMP-8 EA, and MMP-8 in N-1089 MEC was knocked down with siRNA. The effect on adhesion and migration to extracellular matrix (ECM), localisation of α6β4 integrin to hemidesmosomes (HD), TGF-β signalling and gelatinase activity was measured. The effect of altering MEC MMP-8 expression on tumour cell invasion was investigated in 2D and 3D organotypic models.



Assessment of primary cells and MEC lines confirmed expression of MMP-8 in normal MEC and its loss in DCIS-MEC. Over-expression of MMP-8 WT but not MMP-8 EA in β6-1089 cells increased adhesion to ECM proteins and reduced migration. Conversely, knock-down of MMP-8 in N-1089 reduced adhesion and increased migration. Expression of MMP-8 WT in β6-1089 led to greater localisation of α6β4 to HD and reduced retraction fibre formation, this being reversed by MMP-8 knock-down in N-1089. Over-expression of MMP-8 WT reduced TGF-β signalling and gelatinolytic activity. MMP-8 knock-down enhanced TGF-β signalling and gelatinolytic activity, which was reversed by blocking MMP-9 by knock-down or an inhibitor. MMP-8 WT but not MMP-8 EA over-expression in β6-1089 reduced breast cancer cell invasion in 2D and 3D invasion assays, while MMP-8 knock-down in N-1089 enhanced cancer cell invasion. Staining of breast cancer cases for MMP-8 revealed a statistically significant loss of MMP-8 expression in DCIS with invasion versus pure DCIS (p = 0.001).



These data indicate MMP-8 is a vital component of the myoepithelial tumour-suppressor function. It restores MEC interaction with the matrix, opposes TGF-β signalling and MMP-9 proteolysis, which contributes to inhibition of tumour cell invasion. Assessment of MMP-8 expression may help to determine risk of DCIS progression.
Translational Cancer Discovery Team, CRUK Cancer Therapeutics Unit, Institute of Cancer Research, 15 Cotswold Road, Sutton, Surrey, SM2 5NG, UK.
Muge
M
Sarper
Centre for Tumour Biology, Barts Cancer Institute, John Vane Science Centre, Charterhouse Square, Queen Mary University of London, Charterhouse Square, London, EC1M 6BQ, UK. m.allen@qmul.ac.uk.
Michael D
0000-0003-2121-8087
MD
Allen
Centre for Tumour Biology, Barts Cancer Institute, John Vane Science Centre, Charterhouse Square, Queen Mary University of London, Charterhouse Square, London, EC1M 6BQ, UK.
Jenny
J
Gomm
Centre for Tumour Biology, Barts Cancer Institute, John Vane Science Centre, Charterhouse Square, Queen Mary University of London, Charterhouse Square, London, EC1M 6BQ, UK.
Linda
L
Haywood
Cancer Research Centre, Qatar Biomedical Research Institute, Qatar Foundation, Doha, Qatar.
Julie
J
Decock
Centre for Tumour Biology, Barts Cancer Institute, John Vane Science Centre, Charterhouse Square, Queen Mary University of London, Charterhouse Square, London, EC1M 6BQ, UK.
Sally
S
Thirkettle
Centre for Tumour Biology, Barts Cancer Institute, John Vane Science Centre, Charterhouse Square, Queen Mary University of London, Charterhouse Square, London, EC1M 6BQ, UK.
Ahsen
A
Ustaoglu
Centre for Experimental Cancer Medicine, Barts Cancer Institute, John Vane Science Centre, Charterhouse Square, Queen Mary University of London, Charterhouse Square, London, EC1M 6BQ, UK.
Shah-Jalal
SJ
Sarker
Centre for Tumour Biology, Barts Cancer Institute, John Vane Science Centre, Charterhouse Square, Queen Mary University of London, Charterhouse Square, London, EC1M 6BQ, UK.
John
J
Marshall
School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich, NR4 7TJ, UK.
Dylan R
DR
Edwards
Centre for Tumour Biology, Barts Cancer Institute, John Vane Science Centre, Charterhouse Square, Queen Mary University of London, Charterhouse Square, London, EC1M 6BQ, UK. l.j.jones@qmul.ac.uk.
J Louise
JL
Jones
D014408:Biomarkers, Tumor; D018270:Carcinoma, Ductal, Breast / Q000235:genetics* / Q000378:metabolism* / Q000473:pathology; D002285:Carcinoma, Intraductal, Noninfiltrating / Q000235:genetics* / Q000378:metabolism* / Q000473:pathology; D002448:Cell Adhesion; D002461:Cell Line, Transformed; D045744:Cell Line, Tumor; D002465:Cell Movement; D002470:Cell Survival; D002471:Cell Transformation, Neoplastic / Q000235:genetics* / Q000378:metabolism*; D004847:Epithelial Cells / Q000378:metabolism*; D005260:Female; D015870:Gene Expression; D055785:Gene Knockdown Techniques; D006801:Humans; D007150:Immunohistochemistry; D039161:Integrin alpha6beta4 / Q000378:metabolism; D020784:Matrix Metalloproteinase 8 / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D019899:Paracrine Communication; D021381:Protein Transport; D059748:Proteolysis; D015398:Signal Transduction; D016212:Transforming Growth Factor beta / Q000378:metabolism
D014408:Biomarkers, Tumor; D039161:Integrin alpha6beta4; D016212:Transforming Growth Factor beta; D020784:Matrix Metalloproteinase 8; D020780:Matrix Metalloproteinase 9
Adhesion; Ductal carcinoma in situ; Hemidesmosomes; Invasion; MMP-8; Microenvironment; Myoepithelial cell; Organotypic assays