Mechanism of action and therapeutic utility of immunosuppressive oligonucleotide

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Synthetic oligonucleotides (ODN) expressing multiple TTAGGG motifs patterned after the repetitive sequences present at high frequency in mammalian teleomeres down-regulate the inflammatory immune responses elicited by a broad range of TLR ligands and the adaptive immune cell responses induced by polyclonal activators and antigens. These suppressive ODN are useful in the treatment of diseases characterized by over-exuberant immune responses, including septic shock and autoimmunity. Results from my group show that systemically administered Sup ODN alter the host immune milieu and can be harnessed to reduce susceptibility to inflammation- induced cancer. The initial focus of this research examined the effect of Sup ODN in the DMBA/TPA model of skin carcinogenesis. We found that Sup ODN significantly reduced both the number of mice that develop DMBA/TPA dependent papillomas and the number of papillomas/animal in this murine model of inflammation-associated tumorigenesis. To confirm and extend that finding, we initiated studies in a completely different model system of inflammation-promoted tumorigenesis. First, we demonstrated that Sup ODN were effective in preventing/treating the life-threatening pulmonary inflammation caused by silicosis (a disease that affects many miners in the US and abroad). Epidemiologic studies of such miners suggest that the inflammation induced by silica particles (as well as asbestos and coal dust) increases their susceptibility to lung cancer induced by exposure to cigarette smoke. This led us to develop a novel model in which exposure to silica dust plus NNK (a major carcinogen present in cigarette smoke) increases the risk of lung cancer in mice. We've now shown that Sup ODN can be used to significantly reduce silicotic inflammation, and that this returns susceptibility to NNK-induced lung cancer to background levels. To clarify the mechanism by which Sup ODN inhibit tumor development, various measures of inflammation were examined. In both models studied, leukocyte infiltration and the production of pro-inflammatory cytokines and chemokines were significantly reduced whereas control ODN had no significant effect. We are extending these studies to include other agents that cause pulmonary inflammation and disease. We are also using microarray technology to identify the genes and regulatory networks triggered by suppressive ODN. These microarray studies indicate that very large numbers of genes are rapidly down-regulated following the administration of suppressive ODN. Most recently we extended this research to evaluating the effect of Sup ODN in murine models of chronic obstructive pulmonary disease (COPD) and ischemic stroke. In both of those diseases, inflammation increases the severity and extends the duration of pathology. Our findings demonstrate that treatment with Sup ODN effectively reduced inflammation and significantly reduce the severity of both these disease states. We hypothesize that chronic treatment with Sup ODN will be needed to reduce host susceptibility to cancer. Unfortunately, ODN therapy is typically administered parenterally making it difficult to envision long-term human studies/therapy. We've therefore initiated a study to determine if ODN could be delivered orally and retain their activity. Multiple studies of local (GI) and systemic immunity were performed. Results indicate that orally delivered ODN have their primary effect on immune cells in the gut and very little effect systemically. Moreover, encapsulating the ODN to prevent degradation in the stomach has no significant effect on their uptake or activity. The long term goal of this project is to determine whether changing the immune milieu of the host by repetitive administration of Sup ODN alter susceptibility to inflammatory diseases, including cancer. Our recent collaborative work documented the role played by cGAS in mediating the immune suppression critical to that outcome. We also expanded the scope ofthese studies to examine the effect of treating mice with the TLR7/8 agonists R848. Results show that R848, administered either parenterally or orally, improves the immune response of mice such that their susceptibility to infection is significantly reduced.