Genetic approaches to next-generation breast cancer therapy

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RNA interference (RNAi) is conserved through evolution as a mechanism to regulated gene expression. This process can be manipulated experimentaly to repress the expression of any specific gene. My postdoctoral research has focused on two goals: the design and construct an RNAi library targeting the whole human and mouse genome and the development of a genetic-barcode strategy to facilitate genome-wide RNAi screens. Both aims have been accomplished. We have constructed more than 200,000 constructs that target almost the entire human and mouse genomes and we have developed and validated a microarray barcode strategy that allows the loss of function study of thousands of genes at a time. Although important regulators of breast cancer are known, these findings have not decreased remarkably the mortality of this disease. Our RNAi library represents an unique oportunity for comprehensive and systematic large scale functional studies to uncover relevant genes to tumorigenesis. In this grant I propose to use our RNAi library and the technology we have developed in the long-term goal to screen the entire genome to identify and characterize a) Novel Putative Tumor Suppressors and b) Synthetic Lethal Interactions with ErbB2 Activation. I will approach these main objectives through the following aims: a1) Genome-wide RNAi screens to identify genes that promote resistance to apoptosis and uncontrolled proliferation in normal human breast epithelial cells in vitro. a2) Screen candidates selected in the aim a1 in an orthotopic mouse model to identify genes that promote breast cancer in vivo. a3) Employ cellular, molecular and biochemical approaches to elucidate the functions and physiological relevance of candidates identified in the aim a2. b1) Genome-wide RNAi screen to identify candidate genes that show synthetic lethal interaction with activated ErbB2 in vitro. b2) Confirm the synthetic lethal interaction between ErbB2 activation and the candidate genes from the aim b1 in vivo. b3) Employ cellular, molecular and biochemical approaches to gain inside into the biology of the genetic interaction confirmed in b2. The completion of the research I am proposing will increase our understanding of tumorigenesis and will reveal new potential therapeutic targets. My career goal is to develop a streamlined approach for integrative functional genomics studies in breast cancer. The basic idea is to organize a systematic pipeline that begins with genome-wide RNA functional studies in vitro and after in vivo validation in mouse models it concludes with the molecular and biochemical characterization of newly identify genes. The arrangement of this research core will provide me with a consistent and homogeneous assay platform to investigate different aspects of breast cancer.