Project 5. Development of NF-kB Directed Therapy

The level of nuclear factor kappa-B (NF-kB) has been shown to be elevated in ER- human breast cancers, as compared to ER+ tumors. This is correlated with the increased level of epidermal growth factor family receptors (EGF-R) in ER- cells. Results from the Pardee lab have demonstrated that activation of NF-kB is a downstream consequence of EGF signaling. A pathway has been proposed for the EGF-EGFR mediated cell proliferation signal that involves activation of PI3 kinase, protein kinase C and NF-kB with over expression of the down stream cell cycle regulatory protein cyclin D1 and Rb phosphorylation. These results, along with its anti-apoptotic action, strongly suggest the involvement of activated NF-kB in ER- breast cancers. This role has been examined in a mouse tumor model generated with an ER- mouse mammary epithelial carcinoma cell line CSMLO. Blocking of the activation of NF-kB by a protein kinase C inhibitor and by the expression of dominant negative mutant of a subunit of the Ikk-complex caused regression of tumors and is associated with the loss of tumorigenic potential of CSMLO cells.

We will be to test the hypothesis that blocking the activation of the nuclear factor kappa B (NF-kB) will preferentially block the growth of the class of ER- breast cancers that over-express EGF-R family receptors.

1. Screen potential small molecule inhibitors of NF-kB in ER- breast cancer cells with activated EGF signaling pathways
   
   
2. Determine whether influencing the interactions of different components of the Ikk-complex has a significant effect on growth and proliferation of the ER- breast cancer cells

Sankar Ghosh, Ph.D.-Dr. Ghosh is Professor of Immunobiology and Molecular Biophysics & Biochemistry, Yale Medical School and an Associate Investigator, Howard Hughes Medical Institute. He cloned NF-kB and remains at the forefront of the field. He will serve as a co-investigator on Project 5. Email: sankar.ghosh@yale.edu

Arthur B. Pardee, Ph.D.-Dr. Pardee is Emeritus Professor of Biological Chemistry and Molecular Pharmacology, Dana-Farber Cancer Institute and Harvard Medical School. He is a member of the National Academy of Sciences and one of the fathers of molecular biology. Dr. Pardee has made numerous contributions to cancer biology and the current major focus of his research is the development of novel cancer therapeutics. He will serve as a co-investigator on Project 5. Email: Arthur_Pardee@dfci.harvard.edu

DATA

Project Update 2003

The regulatory role of NF-kB on cell proliferation and tumorigenesis was investigated in ER- human breast cancer cells (MDA-MB 231 and MDA-MB-435) and in a mouse mammary epithelial adenocarcinoma cell line (CSMLO). This study was extended to a mouse tumor model by implanting ER- CSMLO cells in syngeneic female A-J mice.

Activation of the NF-kB-IkB complex by EGF signaling pathways in these cells was blocked by the protein kinase C (alpha and beta) inhibitor Go6976 (Nonglycosidic indolcarbazole). The stable expression of dominant-negative mutants of the subunits of IkB kinase (Ikk), more specifically of Ikk-b also blocked NF-kB activation and its down stream events.

Both of these experimental manipulations inhibited CSMLO-induced tumor growth in the mice, and Go6976 caused regression of these tumors. In addition stable expression of dominant negative Ikk-b blocked phorbol myristate acetate (PMA) induced activation of NF-kB and over expression of cyclin D1 and led to reduced tumorigenic potential of CSMLO cells in A-J mice.

These results with both in vivo and in vitro experimental systems strongly suggest the involvement of NF-kB in ER- mammary epithelial cell-mediated tumorigenesis, and also demonstrate that this transcription factor is a potential target for therapy of this class of ER- breast cancers.

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