Research Project
10239044
Project Summary ? Core C The Cell Biology Core provides major resources to the program project investigators, including normal human cells, an extensive library of characterized melanoma cell lines and patient-derived xenografts (PDX) (Objective 1). The melanoma cell line library encompasses >400 cell lines, at least 200 with known genetic status. To exclude cross-contamination, all experimental cell lines are fingerprinted. Our normal cell panel includes: melanocytes, keratinocytes, fibroblasts, endothelial cells, and skin-derived stem cells. The PDX panel encompasses now >500 specimens. They have been characterized for metastasis formation and ~250 have been analyzed for mutations in 108 cancer-related genes. Approximate 50 have been subjected to Whole Exome Sequencing and RNAseq. >350 PDX have been analyzed for expression of >300 proteins. We expect that all PDX will be analyzed during the course of the grant at a genome wide level for abnormalities at the DNA, RNA and protein levels. Other resources include a lentiviral vectors for growth factors, adhesion molecules, and oncogenes and monoclonal antibodies to melanoma-associated biomarkers. Three-dimensional cultures of normal human skin (skin equivalents or organotypic cultures of skin) with a dermis of fibroblasts embedded in collagen and an epidermis of melanocytes/melanoma cells and keratinocytes, are also continuously available to the program investigators (Objective 2). This three-dimensional model is superior to standard cell culture techniques because it mimics the in vivo microenvironment for the melanocytes/melanoma cells. For screening studies the Core is providing a robotics-assisted spheroid culture model that allows investigations on melanoma cells displaying in vivo relevant architecture, cell-to-cell contacts and invasion, in the presence of a collagen support matrix. Spheres are enriched in cancer stem-like cells and can be used instead of spheroids. The Core additionally provides subcutaneous melanoma growth models for standard experiments, including patient-derived xenografts that are exclusively maintained in vivo and using tissues provided by the Pathology Core B. A variety of metastasis models are available as are humanized mice to study the interactions of immune cells with tumor cells.
