Lung cancer is the leading cause of malignancy-related mortality in the United States, and is predicted to increase over the remainder of this decade. Clearly the new strategies for therapy are indicated. While carcinogenesis is complex, tumor growth beyond 2 mm in diameter is dependent upon angiogenesis. Recently, interleukin-8 (IL-8), a member of the C-X-C chemokine family, has been found to be an angiogenic factor. In contrast, platelet factor 4 (PF4), another C-X-C chemokine, has been shown to have angiostatic properties. Interestingly, the major difference between IL-8 and PF4 is the presence of the N-terminal ELR (Glu-Leu-Arg) motif that precedes the first cysteine amino acid residue of IL-8, and is important in ligand/receptor interactions. We hypothesize that angiogenesis associated with tumor growth is dependent upon members of the C-X-C chemokine family acting as either angiogenic or angiostatic factors. This paradigm predicts that the biological balance in the expression of these C-X-C chemokines dictates whether the neoplasm grows and develops metastatic potential or remains and regresses. In this proposal, we will focus on the role of angiogenic and angiostatic C-X-C chemokines and establish whether a biological imbalance in their expression favors tumor-derived angiogenic activity. Our experimental strategy will address the following questions: l) Do angiogenic and angiostatic C-X-C chemokines contribute to the neovascularization of non- small cell lung cancer (NSCLC)? 2) Do subclones of NSCLC cells that constitutively express angiogenic C-X-C chemokines have a selective advantage for growth and metastasis in vivo? 3) Do transfected NSCLC cells with angiostatic C-X-C chemokine genes have a selective disadvantage for growth and metastasis in vivo? 4) Do amino acid alterations of the ELR motif effect the function of angiogenic C-X-C chemokines, and do NSCLC cells transfected with IL-8 mutant genes have a selective disadvantage for growth and metastasis in vivo? 5) Do specific cytokine networks exist during tumorigenesis that shifts the biological balance in favor of angiogenic C-X-C chemokines? Freshly isolated human NSCLC and NSCLC cell lines will serve as our neoplasms. Human NSCLC/SCID mouse chimeras will serve as the in vivo experimental model. Techniques employed in this application will include: Northern blot (or RT PCR) and in situ hybridization analysis; cellular transection and subcloning; immunohistochemistry and Aliases to determine cytokine production; bioassays for angiogenic activity; in vivo passive immunization with neutralizing C-X-C chemokine antibodies. The elucidation of the biology of C-X-C chemokines in the context of neovascularization of NSCLC will permit novel targeted therapy aimed specifically at attenuating tumor growth and metastasis.
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