Identification of Cytokines Regulated by Phosphatidylinositol-3 Kinase in Ovarian Cancer.

Michelle Moscova, Deborah J. Marsh, Robert C. Baxter

Research output: Book chapter/Published conference paperConference paperpeer-review

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Abstract

Activation of phosphatidylinositol-3 kinase (PI3K) is a frequent event in ovarian cancer, leading to increased cell proliferation and inhibition of apoptosis. Although a number of proteins are reported to be involved in PI3K signalling, many targets of this pathway remain unknown. We have previously used proteomic tools to identify several cytokines, including chemokines IL-8 and CXCL1, regulated by PI3K in ovarian cancer. This study aims to identify further novel cytokine targets of PI3K signalling which may point to candidate markers for this pathway and lead to novel therapeutic targets for ovarian cancer. We measured the expression of 120 cytokines in conditioned media and cell lysates from OV167 ovarian adenocarcinoma cells using RayBiocytokine antibody arrays (RayBiotech, Norcross, GA, USA). Cells were subjected to the following treatments for 48h: (1) untreated control; (2) 50 mg/ml EGF; (3) 10 M PI3Kinhibitor LY294002; (4) EGF + LY294002. Conditioned media or cell lysates were incubated with the blocked arrays at 4C overnight and arrays were processed according to the manufacturer's instructions. The chemiluminescent signal was detected usingthe Fuji LAS3000 chemiluminescence reader and quantified by Image Gauge software. Proteins induced at least 2-fold by EGF and inhibited by LY294002 were considered regulated by PI3K signalling. To validate this approach, the concentration of CXCL1was also measured by in-house RIA and compared to the chemiluminescent signal intensity detected on the cytokine arrays. The array signal intensity for CXCL1 showed good correlation to RIA concentrations (R2 = 0.94), confirming the quantitative nature of the arrays. Several cytokines up-regulated by EGF and inhibited by LY294002were identified. These cytokines included IL-6, IL-8 and CXCL1, previously associated with ovarian cancer, as well as a number of other cytokines, not previously reported to beinvolved in PI3K signalling in ovarian cancer. Validation of these results is being performed by immunoassays. This study illustrates the use of antibody arrays in the identification of potential pathway-specific therapeutic targets using a cell model of ovarian cancer. This study was supported by an Australian Postgraduate Award and the Cancer Institute NSW.
Original languageEnglish
Title of host publicationMolecular Targets and Cancer Therapeutics
Place of PublicationPhiladelphia PA
PublisherAmerican Association for Cancer Research
Pagesost-A47/pg67
Publication statusPublished - 2005
EventAACR-NCI-EORTC International Conference . Molecular Targets and Cancer Therapeutics - Philadelphia PA, New Zealand
Duration: 14 Nov 200518 Nov 2005

Conference

ConferenceAACR-NCI-EORTC International Conference . Molecular Targets and Cancer Therapeutics
CountryNew Zealand
Period14/11/0518/11/05

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