CCN1 – A Killer of Esophageal Adenocarcinoma

Sa1087 CCN1 - A Killer of Esophageal Adenocarcinoma Jianyuan Chai, Jennifer Pham, Cristina Modak, Manith Norng Background: While the incidence of most cancers is declining, esophageal adenocarcinoma (EAC) has been the fastest growing malignancy in the world. This rise mainly attributes to gastroesophageal reflux disease in association with obesity. CCN1 is a matricellular protein activated by injury to support healing. Up-regulation of CCN1 has been documented in esophageal epithelial cells under acidic condition and in Barrett's esophagus. The aim of this study is to determine whether CCN1 is a pro or con factor in EAC. Methods: EAC cells (OE19 and OE33) were treated with recombinant CCN1 at 500ng/ml for 2 and 24 hours. Apoptosis was assessed by real time RT-PCR array and mitochondrial permeability assay. Normal esophageal epithelial cells (Het-1A) were included for comparison. Results: In EAC cells, 2-hour treatment with CCN1 highly up-regulated pro-apoptotic genes, including CASP4 (570 fold), CRADD (31 fold), FASTK (175 fold), FOXO3 (82 fold), IKBKG (188 fold) and IRF7 (188 fold), and completely shut down the expression of 18 anti-apoptotic genes including BIRC5, BCL2L1, HIPK3, MYC, JUN, NFKB1, PIK3R1, MAP2K4, XIAP, etc. Apoptosis was also confirmed by severe mitochondrial leakage. On the contrary, the same treatment mainly promoted survival in normal esophageal epithelial cells, including 137fold up-regulation of BIRC5. Conclusions: CCN1 promotes EAC cell death while protects normal esophageal epithelial integrity.

Top Panel: Western blot Bottom panel: Proliferation assay Sa1085 Gdf15 is Over-Expressed in Barrett's Esophagus (BE), Induces SMAD Activation and May Play a Crucial Role in the Transcriptional Cascade Leading to Cell Differentiation and Transformation Murat Kirca, Shane P. Duggan, Fiona M. Behan, Abdul Zaheer, Dermot P. Kelleher Introduction:- Previous micro array studies and validation with Rt-PCR in a patient cohort from our group have shown that GDF 15 is up regulated in BE. GDF15 is a member of the transforming growth factor-beta superfamily which includes TGF-beta and BMP ligands and may be involved in regulation of tissue differentiation and maintenance. TGF-beta and BMP ligands signal through cell receptors inducing Smad activation. Activated Smad 2/3 builds a complex with Smad 4. The activated Smad-complex accumulates in the nucleus and can initiate gene expression. This transcriptional cascade appears crucial in cell transformation and differentiation as seen in BE. The aim was to examine the signaling potential of GDF15 in this cascade. Methods: - Recombinant human GDF15 was used in stimulating Het1a cells (human esophageal squamous epithelial cell line). Smad activation into the nucleus was measured with immunostaining using Smad 1/2/3 antibodies and their nuclear intensity after stimulation. The intensity of activated Smad in the nucleus and quantitative amount of GDF15 utilizing Rt-PCR in Oe 33 cells (esophageal cancer cell line) was measured and then compared with levels in HET1a cells. Results: We found that stimulation of Het1a cells with GDF 15 leads to a time and dose dependant Smad activation. We established that optimal induction of Smad activation in Het1a cells is at 5 ng/ml with GDF15 for 1 h (p