ABSTRACT
Infiltration of specialized immune cells regulates the growth and survival of neoplasia. Here, in a survey of public whole genome expression datasets we found that the gene for chemerin, a widely expressed endogenous chemoattractant protein, is down-regulated in melanoma as well as other human tumors. Moreover, high chemerin messenger RNA expression in tumors correlated with improved outcome in human melanoma. In experiments using the B16 transplantable mouse melanoma, tumor-expressed chemerin inhibited in vivo tumor growth without altering in vitro proliferation. Growth inhibition was associated with an altered profile of tumor-infiltrating cells with an increase in natural killer (NK) cells and a relative reduction in myeloid-derived suppressor cells and putative immune inhibitory plasmacytoid dendritic cells. Tumor inhibition required host expression of CMKLR1 (chemokine-like receptor 1), the chemoattractant receptor for chemerin, and was abrogated by NK cell depletion. Intratumoral injection of chemerin also inhibited tumor growth, suggesting the potential for therapeutic application. These results show that chemerin, whether expressed by tumor cells or within the tumor environment, can recruit host immune defenses that inhibit tumorigenesis and suggest that down-regulation of chemerin may be an important mechanism of tumor immune evasion.
Subject(s)
Chemotactic Factors/immunology , Intercellular Signaling Peptides and Proteins/immunology , Killer Cells, Natural/immunology , Melanoma, Experimental/immunology , Animals , Cell Growth Processes/immunology , Cell Line, Tumor , Chemokines/genetics , Chemokines/immunology , Chemokines/metabolism , Chemotactic Factors/genetics , Chemotactic Factors/metabolism , Dendritic Cells/immunology , Dendritic Cells/metabolism , Down-Regulation , HEK293 Cells , Humans , Intercellular Signaling Peptides and Proteins/genetics , Intercellular Signaling Peptides and Proteins/metabolism , Killer Cells, Natural/metabolism , Melanoma, Experimental/genetics , Melanoma, Experimental/metabolism , Melanoma, Experimental/pathology , Mice , Mice, Inbred C57BL , Mice, Knockout , Myeloid Cells/immunology , Myeloid Cells/metabolism , RNA, Messenger/genetics , RNA, Messenger/immunology , RNA, Messenger/metabolism , Receptors, Chemokine/genetics , Receptors, Chemokine/immunology , Receptors, Chemokine/metabolism , Receptors, G-Protein-Coupled/genetics , Receptors, G-Protein-Coupled/immunology , Receptors, G-Protein-Coupled/metabolismABSTRACT
The secreted immunoglobulin footprint of single hybridoma cells, containing ~10 fg of antibody purified in situ, has been probed for 9 properties concurrently by use of detection labels comprising 280 nm combinatorially colored fluorescent latex beads functionalized with proteins. Specificity of each individual hybridoma cell's product has thereby been assessed in a primary screen. Varying the density of antigen on beads to modulate the avidity of the interaction between bead and secreted antibody footprint allowed rank ordering by affinity in the same primary screen. As more criteria were added to the selection process, the frequency of positive cells went down; in some cases, the favorable cell was present at <1/50,000. Recovery of the cell of interest was accomplished by plating the cells in a viscous medium on top of a membrane. After collecting the antibody footprint on a capture surface beneath the membrane, the immobilized cells were transferred to an incubator while the footprints were analyzed to locate the hybridoma cells of interest. The desired cells were then cloned by picking them from the corresponding locations on the membrane.
