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B. R. Ksander, J. J. Bosch, N. Y. Frank, S. Ostrand-Rosenberg, T. G. Murray, M. H. Frank, P. E. Kolovou; T Cells Targeting Cancer Stem Cells in Uveal Melanoma.. Invest. Ophthalmol. Vis. Sci. 2010;51(13):842.
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At least one reason for the controversy surrounding the cancer stem cell theory is the lack of specific markers that allow purification of this tumor cell subpopulation. Recently, an ABCB5+ subpopulation of skin melanoma cells was shown to contain cancer stem cells with enhanced tumorigenicity and metastatic potential. ABCB5 is an ATP-binding cassette transporter that acts as an efflux pump. The current studies were performed to determine if uveal melanoma cells (primary and metastatic) possess an ABCB5+ subpopulation. We hypothesize that activation of T cells specific for this cancer stem cell subpopulation will achieve destruction of metastatic tumors. In order to test this hypothesis, we transfected ABCB5+ uveal melanoma cells with a series of genes that allows the tumor cells to express endogenous tumor antigens and activate specific T cells.
Primary and metastatic uveal melanoma cell lines (Mel 202 and 270; OMM1.3) were examined for the ABCB5 stem cell marker by flow cytometry. ABCB5 positive and negative tumor cells were transfected with vectors containing the cDNA for: (i) MHC Class II (DRA*0101 and DRB1*0101), and (ii) the co-stimulatory molecule CD80. Three-color flow cytometry and cell sorting were used to identify ABCB5 positive and negative viable tumor cells that expressed the transgenes.
All uveal melanoma cell lines (primary and metastatic) possessed a small subpopulation of ABCB5+ cells (ranging from 2-9%). ABCB5+ and ABCB5- tumor cells were transfected with MHC Class II and CD80, as demonstrated by three-color flow cytometry. Cell sorting was used to purify and separate ABCB5 positive and negative tumor cells for restimulation of PBL in vitro.
A uveal melanoma ABCB5+ cancer stem cell subpopulation was genetically modified to express CD80 and Class II, in order to transform the tumor cells into antigen presenting cells that stimulate CD4+ T helper cells. We predict these tumors will activate specific T cells that effectively eliminate and prevent progressive tumor growth.
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