The purposes of the project are to investigate the biological roles of members of the chemokine family of cytokines, to use chemokine receptors to understand the relationships between the trafficking patterns and broader biological functions of subsets of effector/memory T cells, and to understand the contributions of the chemokine system to infectious and inflammatory/autoimmune disease and cancer. Chemokines and their receptors are critical for leukocyte trafficking, and our experiments are directed to understanding how manipulating the chemokine system could be used to treat diseases in which leukocytes play a critical role. In addition, some chemokine receptors are expressed by cancer cells, and these receptors can potentially be exploited for diagnosis and as targets for therapy. In FY 2014, we have continued to investigate a mouse model of skin inflammation that has features of psoriasis. The model involves injection of a cytokine, IL-23, which appears to have a role not only in psoriasis, but also in other immune-mediated diseases, such as Crohns disease. We and others had described that the chemokine receptor CCR6 is expressed by IL-23-dependent T cells that produce the cytokines IL-17 and IL-22. IL-22 and IL-17 are important in producing disease in the mouse psoriasis model, and are thought to be important in causing tissue injury in some autoimmune diseases. We have shown previously that mice lacking CCR6 are resistant to the IL-23-induced disease, but the mechanism has been unclear. In the past year, we have further characterized the origins and types of monocyte/macrophage and dendritic cell subsets that infiltrate the skin and contribute to the psoriasis-like inflammation. Monocyte/macrophages and dendritic cells are cells of the innate immune system that produce pro-inflammatory factors and affect the activities of other immune cells, such as T cells. We have identified a subset of inflammatory dendritic cells that enter the outer layers of the skin (epidermis), determined that these and other dendritic cells important for the inflammatory response are derived from infiltrating monocytes, and studied how these cells are recruited. In the last year we have concluded a project focused on the role of the cytokine IL-22 and natural killer (NK) cells in a model of bacterial pneumonia. Published data suggested that the Th17 cell-associated cytokine, IL-22, was critical for host defense against bacterial infection in the lung, and that the IL-22 was derived from Th17 cells. On the contrary, we found that NK cells were an important source of IL-22 in response to lung infection in mice with Klebsiella pneumoniae, a pathogen of increasing concern due to nosocomial infections with multi-drug resistant strains. We found that mice could control Klebsiella pneumoniae pneumonia in the absence of T cells, but that they required both IL-22 and NK cells, and that infection made lung NK cells competent to produce IL-22. Importantly, the NK cells were not the recently described NK cell-like innate lymphoid cells that have been described as IL-22 producers in the gastrointestinal tract, but NK cells with a conventional phenotype. In the last year we have continued using CCR6 expressed on helper T cells from human blood to study the mechanisms controlling gene expression that allow these cells to produce the factors, such as IL-17 and IL-22, that contribute to autoimmune/inflammatory disease. We have identified a number of proteins that regulate the expression of the genes for CCR6, IL-17 and other proteins important for the activities of Th-17-cells. These regulatory proteins affect both the ability of nave T cells to become Th17 cells and the ability of Th17 cells to maintain their activities. In the last year we have continued studies of the process whereby effector/memory T cells migrate from the blood, across the layer of endothelial cells that line the inside off the blood vessel, into a site of tissue infection or inflammation. We have characterized subsets of CD8+ (killer) T cells that are particularly efficient at migrating across the endothelium, and we have identified some of the molecular features of the surfaces of these cells, including the combination of chemokine receptors, that make them so efficient. In the last year we have identified a transcription factor that regulates a number of genes that encode proteins important for the ability of these cells to migrate across endothelium into inflammatory sites. In the last year we have continued studies focused on functions for CXCR4 and its ligand, CXCL12, in the physiology of the adrenal gland and in adrenal adenomas and malignancies. CXCR4 is an unusual chemokine receptor, in that it has roles outside of the hematopoietic system, including in the development of the nervous and cardiovascular systems and the positioning of germ cells. We found that CXCR4 is highly expressed on zona glomerulosa cells, in both mouse and human. In mouse adrenal cortex, CXCL12 mRNA and protein, respectively, are expressed near the adrenal capsule, correlating with the site of CXCR4 expression. We have also found high levels of CXCR4 expression in aldosterone-producing adenomas and hyperplasias. In the last year we have opened a phase 0 clinical protocol, 14-I-0050, entitled Imaging CXCR4 Expression in Subjects with Cancer Using 64Cu-Plerixafor. In mouse models and in patients, expression of the chemokine receptor CXCR4 on various cancers has been correlated with aggressive biological behavior, including increased rates and certain sites of metastasis, and decreased survival. Plerixafor has been identified as a specific inhibitor of CXCR4, and it is currently approved by the Food and Drug Administration as a mobilizing agent for hematopoietic stem cells. We have previously shown that plerixafor can be labeled with the positron-emitting radionuclide copper-64 (64Cu) to form 64Cu-plerixafor, which can be used to visualize CXCR4-positive tumor xenografts in mice using small-animal positron emission tomography (PET). Determining CXCR4 expression in tumors using 64Cu-plerixafor and PET/computerized tomography (CT) scanning could be useful in predicting tumor behavior and responses to current and experimental therapies, including therapies targeting CXCR4. Our protocol will enroll subjects with a range of malignancies in order to determine if 64Cu-plerixafor can be used safely in order to quantify CXCR4 expression on tumors.
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