[Frontiers in Bioscience 14, 2170-2180, January 1, 2009]

2-Methoxyestradiol mediated signaling network in pancreatic cancer

Aruna Basu, Subrata Haldar

Center for Biomedical Sciences, Department of Pharmacology, Case Comprehensive Cancer Center, MetroHealth Campus, Case Western Reserve University, Cleveland, OH 44109, USA

TABLE OF CONTENTS

1. Abstract
2. Introduction
3. Materials and Methods
3.1. Cell lines
3.2. Sample preparation for powerblot analysis
3.3. PowerBlot Western array analysis
3.4. Conventional Western blot analysis
3.5. Cell based ELISA
3.6. NF-kB transcription factor assay
3.7. Apoptosis assay
4. Results
4.1. Differential expression of proteins regulated by 2-Methoxyestradiol (2-ME) in pancreatic carcinoma cells
4.2. 2-ME and Transforming growth factor b (TGFb)-Smad signaling in pancreatic cancer
4.3. Decreased expression of IRS-1 following 2-ME exposure
4.4. Modulation of cell cycle regulatory proteins by 2-ME
4.5. Effect of 2-ME on Hypoxia Inducible Factor /Vascular Endothelial Growth Factor (VEGF) signaling
4.6. 2-ME can down regulate Glucocorticoid receptor (GR)
4.7. 2-ME increases nuclear factor-kB (NF-kB) DNA binding activity by diminishing total IkBa level
5. Discussion
6. Acknowledgments
7. References

1. ABSTRACT

2-Methoxyestradiol (2-ME), an endogenous metabolite of 17 beta-estradiol, is known to be a potent inhibitor of neovascularization. Our previous studies have shown that 2-ME can suppress growth of pancreatic tumor cells in vitro and in vivo by the induction of apoptosis (Cancer Res 66: 4309-18, 2006). In order to better comprehend the signaling modulators of 2-ME in pancreatic cancer, we employed a PowerBlot Western Array screening system. Our proteomic profiling has provided framework to define the novel mechanisms of actions of 2-ME in pancreatic cancer. Interestingly, this high-throughput analysis identified proteins such as Rac1, Gelsolin, Glucocorticoid receptor (GR), Smad 2/3, Smad 4, IRS-1, which were not previously reported with 2-ME response. Interestingly, 2-ME modulated down regulation of GR level is accompanied by NF-k B activation in 2-ME responsive but not in resistant pancreatic cancer cells. In view of this observation, possible reciprocal relationship between GR and NF-kappaB activation might be an important regulatory factor in 2-ME mediated demise of a subpopulation of pancreatic cancer cells.