The central aim of this sub-project is to define cellular gene expression patterns associated with the infection by different oncogenic agents. We hope to identify key steps that could be amenable to pharmacological intervention and to establish the role of individual microbial gene products in the cancers associated with infection. Specific objectives: Identification of cellular pathways and gene expression patterns activated by oncogenic infections Identification of key steps in signalling cascades that could yield putative drug targets in cancers caused by infection Validation of targets using RNAi leading to creation of screening assays for inhibitors Identifying changes in gene expression involved in cancers linked to viral or bacterial infection For each infectious agent the objective is the same – to identify key mechanisms by which it may contribute to cancer, providing targets for novel treatment approaches or diagnostic procedures. EBV is associated with cancers of both lymphoid and epithelial cells so cell genes and proteins affected by the viral infection will be identified in these cell types. The roles of viral genes, particularly EBER RNAs, the BART gene products and LMP proteins will be investigated. RNAi will be used to identify cellular genes that are required for the downstream signalling activities of the LMP-1 protein, a key mediator of the oncogenic effects of EBV in a search for new ‘druggable’ effectors of LMP-1 function. For KSHV, endothelial cells infected with recombinant KSHV deletion mutants lacking the viral genes for K1, K15, orf74/GCR, vFLIP, vcyc, vMIPs or vIL6 will be used to determine the effects of these genes on cell signalling pathways. Knock down of these viral genes by RNAi in KSHV infected cells will be carried out in parallel. Comparison of the gene expression microarray profiles will predict how targeting individual viral genes or signalling pathways triggered by them with small molecule inhibitors could affect cellular response patterns. The predictions will be tested experimentally using novel inhibitors of cellular kinases. High risk HPV types are known to cause changes in the cellular transcriptome of infected cells but it is unclear whether identified markers (eg p16INK4a) constitute bona fide predictive markers or just allow better recognition of incident disease. Cellular gene expression patterns associated with premalignant/malignant lesions and with progression to invasive cervical cancer will be identified. Such predictive markers would be a major improvement to the current diagnostic procedures applied for cervical cancer screening. Signalling in gastric epithelial cell lines infected by Helicobacter pylori will be examined using clinical isolates from patients with gastric cancer/ chronic gastritis as well as mutants in H. pylori virulence genes. Selected strains with high and low signaling responses will be further tested for induction of changes in gastric epithelial gene expression of relevance to gastric cancer. Clinical isolates of interest will be tested in the gerbil model, which closely mirrors the situation observed in H. pylori infected patients. This will be used to examine the functional importance of specific H. pylori virulence factors in the development of gastric inflammation and cancer. The sub-project will deliver Specific pathways regulated by infectious agents that cause cancer Targets within those pathways for inhibition by RNAi or pharmacological inhibitors Assays and methods for identifying inhibitors of those pathways In some cases inhibitors that will be lead compounds for drug development