RNA-interference-based genomic screeing for regulators of radiation sensitivity
In clinical radiation therapy, the cancer cells are killed due to DNA damage induced by ionizing radiation. In order to find new ways to predict responses to radiation therapy, and to develop new strategies to sensitize resistant tumors, it is important to understand the molecular mechanisms that determine cellular radio-sensitivity. In this project we are applying RNA-interference-based genomic screening to search for new regulators of radiation sensitivity. We are applying siRNA libraries to deplete, one by one, every single protein in human cells, and measuring the effects on cellular radiation responses. This project is a collaboration with researchers at BRIC Biocenter in Copenhagen. Essential for the siRNA screening is a fully equipped automatic robot workstation available at BRIC for rapid handling and imaging of large scale experiments. During 2007 we have explored the experimental conditions for such screens and particularly succeeded to develop a robust experimental setup for a screen related to the G2 checkpoint and radio-sensitivity. By using Chk1 siRNA as a positive control in human osteosarcoma U2-OS cells, we have achieved very good differences between negative and positive control in our test screens. By using this setup we have recently got the first results from a screen with a kinase siRNA library (purchased from Ambion). We are currently in the process of verifying the positive hits from this screen, as well as broadening the screen by applying several other siRNA libraries in addition to the kinase library. This project continues in 2008-2010 with support from Helse Sør-Øst. Following the identification of positive hits in the initial screening, we will select some of the most interesting genes and work on the molecular mechanisms involved in regulating radiation responses. Our long term goal is to explore potential clinical applications of our findings.