The Molecular Genetics Section (MGS) seeks to elucidate the complex molecular/genetic program governing tumor genesis and progression through the development of genetically engineered mouse models of human cancer. A major goal is to identify candidate targets or pathways for both mechanistic enlightenment and future therapeutic utility. Our efforts in this regard are focused on two tumor types, cutaneous malignant melanoma (CMM) and the pediatric malignancy rhabdomyosarcoma (RMS). Exposure to UV radiation is a causal agent in the vast majority of CMM. Retrospective epidemiological data suggest that CMM is provoked by intense intermittent exposure to UV, particularly during childhood. We tested this hypothesis in mice in which expression of a transgene encoding the c-Met ligand, hepatocyte growth factor/scatter factor (HGF/SF), induced sporadic melanocytic tumors in aged animals. We discovered that a single neonatal dose of erythemal UV radiation was necessary and sufficient to induce cutaneous melanoma reminiscent of human CMM with high penetrance and relatively short latency (Noonan et al., Nature 413: 271-2, 2001). A critical role for the INK4a/ARF locus, widely regarded as a key melanoma suppressor in human patients, in our HGF/SF transgenic mouse model was confirmed by demonstrating that UV-induced melanoma was significantly accelerated on a genetic background deficient in Ink4a/Arf (Recio et al., Cancer Res. 62: 6724-30, 2002). These results strongly suggest that sunburn is a significant risk factor in kindreds harboring germline mutations in INK4a/ARF (Merlino and Noonan, Trends Mol. Med. 9: 102-8, 2003). The childhood malignancy RMS, accounting for 5 to 10% of all pediatric neoplasms and for more than 50% of pediatric soft tissue sarcomas, is thought to arise from imbalances in skeletal muscle cell proliferation and differentiation. However, molecular pathways associated with RMS remain largely unknown, due in part to the lack of an RMS-prone mouse model. In the course of studying genetic interactions between c-MET and the INK4a/ARF locus, we discovered that virtually all HGF/SF transgenic, Ink4a/Arf-deficient mutant mice rapidly succumbed to highly invasive RMS (Sharp et al., Nature Med. 8: 1276-80, 2002). Comparable molecular lesions in c-MET, pRB and p53 pathways have been individually described for human RMS. These data provide genetic evidence that c-MET and INK4a/ARF pathways represent critical and synergistic targets in RMS pathogenesis, and suggest a rational therapeutic combination to combat this pediatric cancer. We have now generated numerous highly and poorly metastatic cell lines from arising RMS tumors, which are proving to be a valuable research tool for studying mechanisms associated with metastatic dissemination. Members of the mitogen-activated protein kinase (MAPK) superfamily, including p38 kinase and SAPK/JNK, play a central role in mediating cellular response to environmental stress, growth factors and cytokines. Binding of HGF/SF to c-MET stimulates multiple signal transduction pathways, leading to the activation of numerous transcription factors. We have determined that HGF/SF can induce cyclin D1 expression in mouse melanoma cells, and that this up-regulation is mediated in part by activating transcription factor-2 (ATF-2) (Recio et al., Oncogene 21: 1000-8, 2002). Moreover, our studies showed that ATF-2 becomes activated by HGF/SF through p38 MAPK and SAPK/JNK, and that the p38-ATF-2 pathway can help mediate proliferation signals in tumor cells through transcriptional activation of key cell cycle regulators. Variants of the CD44 family of surface adhesion molecules, including CD44v6, have been implicated in cancer progression and metastasis. CD44 isoforms bearing heparin sulfate chains can bind to HGF/SF and facilitate its presentation to c-MET. We have discovered that HGF/SF-Met binding upregulates expression of CD44v6 in murine melanoma cells, serving to compensate for loss by internalization (Recio et al., Cancer Res. 63: 1576-82, 2003). c-Met-mediated CD44v6 up-regulation was achieved through transcriptional activation of the immediate early gene egr-1. HGF/SF induced egr-1 activation via the Ras>Erk1/2 pathway, but not through either PI3K or PKC. We identified an Egr-1 binding site in the mouse CD44 gene promoter that accounts for its responsiveness to HGF/SF in melanoma cells. The compensatory up-regulation of both c-Met and CD44v6 in response to HGF/SF has important implications with respect to strategies employed by cancer cells to sustain stimulation of growth- and metastasis-promoting pathways associated with tumor progression.
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