Functional analysis of genetic alterations occuring in uveal melanoma

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The experiments described in this proposal aim to analyze the genetic alterations found in uveal melanoma. The goal is to examine the function of the recently discovered gene mutations in GNAQ and GNA11 both in vitro and in vivo as well as establish corresponding mouse models.1) Characterization of GNAQ and GNA11 Q209L oncogenes in melanocytesThe function of the oncogenes GNAQ and GNA11 in the development of uveal melanomas is currently poorly understood. To achieve this, the biological function of this gene is to be studied in the following fashion: 1. Determine the protein domains and signal effectors that are essential for tumor formation, 2. A detailed analysis of GNAQ/11 oncogene induce protein-phosphorylations and RNA expression, 3. Analysis of GNAQ/11 induced senescence or apoptosis, 4. Studying the metastasis profile of GNAQ/11 oncogene transformed cells, 5. Participation of other genetic factors i.e. BAP1, MYC, PTEN and p53 in GNAQ/11 oncogene induced tumors and metastasis. The goal of this first project aim is to identify functional mechanisms that could be of potential use for developing effective therapeutic strategies.2) Establishing GNAQ/11 oncogene mouse modelsIn contrast to an extensive list of existing mouse models with various oncogenes (BRAF, HRAS, KIT) for cutaneous melanomas, there are currently no existing mouse models of GNAQ/11 oncogenes. Establishing a model of uveal melanoma using these physiological oncogenes would be valuable both from a basic science as well as a translational oncology perspective. With the design and cloning of various transgenic and knock-in constructs during his postdoc in the US, the applicant has laid the groundwork for this project. These mice will allow the expression of GNAQ/11 oncogenes at various timepoints in different tissues. It is planned to: 1. Test the effect of this expression, both on tumor genesis as well as embryonic development in both melanocytic and other tissue. 2. Through different crossings, analyze in vivo the role of additional gene alterations affecting i.e. BAP1, CDKN2A, p53, MYC and PTEN in combination with GNAQ/11 oncogene expression. 3. Apply the mouse model to test a variety of different therapeutic approaches for effectively inhibiting tumor growth and metastasis.