Up-regulated p27 Kip1 Reduces Matrix Metalloproteinase-9 and Inhibits Invasion of Human Breast Cancer Cells

. Background: p27 Kip1 is a cyclin-dependent kinase inhibitor which has been reported to be associated with invasion, metastasis and angiogenesis in malignant tumors, but its mechanism of action remains unknown. Here, it was examined whether p27 Kip1 has an inhibitory effect on cancer cell invasion and correlates with matrix metalloproteinase expression (MMPs). Material and methods: The human breast cancer cell line MDA-MB-231 and MDA-MB-231 transfected p27 Kip1 MDA-MB-p27 were used for the invasion assay, Western blotting and real-time quantitative RT-PCR. Results: In the invasion assay, the invasion of MDA-MB-p27 was significantly less than that of the parent cell line. In Western blotting analyses, the protein level of MMP-9 was also reduced in MDA-MB-p27. Furthermore, the activity of MMP-9 in cell culture supernatants was lower in MDA-MB-p27 as compared with enzyme-linked immunosorbent assays. In real-time quantitative RT-PCR, the mRNA level of MMP-9 was lower in MDA-MB-p27 cells. Conclusion: Upregulation of p27 Kip1 remarkably inhibited the invasion of the breast cancer cells, in part due to the reduced expression of MMP-9. This is the first report of p27 Kip1 modulating MMP-9 and indicating that p27 Kip1 might play a key role in tumor cell invasion. RT-PCR, transcription-polymerase chain enzyme-linked immunosorbent TIMPs, metalloproteinases.

[1]  Young-Choon Lee,et al.  Quercetin exerts multiple inhibitory effects on vascular smooth muscle cells: role of ERK1/2, cell-cycle regulation, and matrix metalloproteinase-9. , 2003, Biochemical and biophysical research communications.

[2]  G. Kouraklis,et al.  Adenovirus-mediated expression of antisense MMP-9 in glioma cells inhibits tumor growth and invasion , 2002, Oncogene.

[3]  I. Jacobs,et al.  A model for co-expression pattern analysis of genes implicated in angiogenesis and tumour cell invasion in cervical cancer , 2002, British Journal of Cancer.

[4]  E. Feldman,et al.  N-Myc and Bcl-2 coexpression induces MMP-2 secretion and activation in human neuroblastoma cells , 2002, Oncogene.

[5]  A. Diez-Juan,et al.  Overexpression of p27Kip1 by doxycycline‐regulated adenoviral vectors inhibits endothelial cell proliferation and migration and impairs angiogenesis , 2001, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[6]  M. Stack,et al.  Phosphatidylinositol 3-kinase activity in epidermal growth factor-stimulated matrix metalloproteinase-9 production and cell surface association. , 2001, Cancer research.

[7]  S. Dedhar,et al.  The integrin linked kinase (ILK) induces an invasive phenotype via AP-1 transcription factor-dependent upregulation of matrix metalloproteinase 9 (MMP-9) , 2000, Oncogene.

[8]  A. Ricca,et al.  bcl‐2 over‐expression enhances NF‐κB activity and induces mmp‐9 transcription in human MCF7ADR breast‐cancer cells , 2000, International journal of cancer.

[9]  R. Medema,et al.  AFX-like Forkhead transcription factors mediate cell-cycle regulation by Ras and PKB through p27kip1 , 2000, Nature.

[10]  C. Asselin,et al.  MAP kinase cascade is required for p27 downregulation and S phase entry in fibroblasts and epithelial cells. , 1999, American journal of physiology. Cell physiology.

[11]  Michele Pagano,et al.  SKP2 is required for ubiquitin-mediated degradation of the CDK inhibitor p27 , 1999, Nature Cell Biology.

[12]  H. Wu,et al.  PTEN modulates cell cycle progression and cell survival by regulating phosphatidylinositol 3,4,5,-trisphosphate and Akt/protein kinase B signaling pathway. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[13]  A. Gulino,et al.  Transcriptional up-regulation of matrix metalloproteinase-9 expression during spontaneous epithelial to neuroblast phenotype conversion by SK-N-SH neuroblastoma cells, involved in enhanced invasivity, depends upon GT-box and nuclear factor kappaB elements. , 1999, Cell growth & differentiation : the molecular biology journal of the American Association for Cancer Research.

[14]  Q. Li,et al.  Recombinant adenovirus expressing Von Hippel-Lindau-mediated cell cycle arrest is associated with the induction of cyclin-dependent kinase inhibitor p27Kip1. , 1998, Biochemical and biophysical research communications.

[15]  S. Srivastava,et al.  Effects of adenovirus-mediated p16INK4A expression on cell cycle arrest are determined by endogenous p16 and Rb status in human cancer cells , 1998, Oncogene.

[16]  P. Seth,et al.  Promoting apoptosis: a novel activity associated with the cyclin-dependent kinase inhibitor p27. , 1997, Cancer research.

[17]  S. R. Datta,et al.  Akt Phosphorylation of BAD Couples Survival Signals to the Cell-Intrinsic Death Machinery , 1997, Cell.

[18]  L. Matrisian,et al.  Changing views of the role of matrix metalloproteinases in metastasis. , 1997, Journal of the National Cancer Institute.

[19]  Esposito,et al.  Prognostic role of the cyclin-dependent kinase inhibitor p27 in non-small cell lung cancer. , 1997, Cancer research.

[20]  Kenneth Cowan,et al.  A recombinant adenovirus expressing p27Kip1 induces cell cycle arrest and loss of cyclin-Cdk activity in human breast cancer cells , 1997, Oncogene.

[21]  M. Loda,et al.  Increased proteasome-dependent degradation of the cyclin-dependent kinase inhibitor p27 in aggressive colorectal carcinomas , 1997, Nature Medicine.

[22]  Herman Yeger,et al.  Decreased levels of the cell-cycle inhibitor p27Kip1 protein: Prognostic implications in primary breast cancer , 1997, Nature Medicine.

[23]  James M. Roberts,et al.  Expression of cell-cycle regulators p27Kip1 and cyclin E, alone and in combination, correlate with survival in young breast cancer patients , 1997, Nature Medicine.

[24]  C. Sherr.  Cancer Cell Cycles , 1996, Science.

[25]  Y. Okada,et al.  Production of Matrix Metalloproteinases and Tissue Inhibitors of Metalloproteinases in Human Breast Carcinomas , 1996, Japanese journal of cancer research : Gann.

[26]  P. Seth,et al.  Effects of a recombinant adenovirus expressing WAF1/Cip1 on cell growth, cell cycle, and apoptosis. , 1995, Cell growth & differentiation : the molecular biology journal of the American Association for Cancer Research.

[27]  T. Hunter,et al.  Cyclins and cancer II: Cyclin D and CDK inhibitors come of age , 1994, Cell.

[28]  C. Sherr.  G1 phase progression: Cycling on cue , 1994, Cell.

[29]  James M. Roberts,et al.  Cloning of p27 Kip1 , a cyclin-dependent kinase inhibitor and a potential mediator of extracellular antimitogenic signals , 1994, Cell.

[30]  R. Davis,et al.  The mitogen-activated protein kinase signal transduction pathway. , 1993, The Journal of biological chemistry.

[31]  H. Sato,et al.  Regulatory mechanism of 92 kDa type IV collagenase gene expression which is associated with invasiveness of tumor cells. , 1993, Oncogene.

[32]  W. Stetler-Stevenson.  Type IV collagenases in tumor invasion and metastasis , 1990, Cancer and Metastasis Reviews.

[33]  D. Clayton,et al.  Tumour basement membrane laminin in adenocarcinoma of rectum: An immunohistochemical study of biological and clinical significance , 1986, International journal of cancer.

[34]  J. Lissitzky,et al.  Immunohistochemical detection of laminin in 98 human breast carcinomas: a light and electron microscopic study. , 1986, Human pathology.

[35]  C. Denkert,et al.  An inhibitor of stress-activated MAP-kinases reduces invasion and MMP-2 expression of malignant melanoma cells , 2004, Clinical & Experimental Metastasis.

[36]  V. Gouyer,et al.  Specific expression of matrix metalloproteinases 1, 3, 9 and 13 associated with invasiveness of breast cancer cells in vitro , 2004, Clinical & Experimental Metastasis.

[37]  S. Marx,et al.  Role for p 27 Kip 1 in Vascular Smooth Muscle Cell Migration , 2001 .

[38]  P. Watson,et al.  A matrix metalloproteinase inhibitor, batimastat, retards the development of osteolytic bone metastases by MDA-MB-231 human breast cancer cells in Balb C nu/nu mice. , 2001, European journal of cancer.