URPP Fellow: Sandra Varum
The main goal of our first term URPP project was to identify novel neural crest stem cell (NCSC) regulators and to determine whether some of these regulators play a role in melanoma disease formation and progression. To address this question we performed a microarray analysis of NCSCs and NC cells primed for differentiation. Among several factors, we identified the transcription factor Yin Yang (Yy1) as a potential modulator of NC biology. Conditional ablation of Yy1 in the NC lineage resulted in agenesis or hypoplasia of various NC derivatives, suggesting that Yy1 is required for NCSC development. Furthermore, conditional ablation of Yy1 in the melanoma mouse model TyR:: N-Ras Q61K Ink4a-/- revealed that depletion of Yy1 impaired skin melanoma formation but increased invasion of single melanoma cells. These results are in agreement with the phenotype-switching model that proposes that in order to give rise to metastasis, melanoma cells switch from a proliferative to an invasive phenotype and vice-versa by responding to environmental cues (Hoek et al., 2008). Our preliminary results suggest that Yy1 regulates melanoma proliferation and invasion by modulating cellular metabolism. Strikingly, metabolic changes have recently also been implicated in targeted melanoma therapies as well as in cancer immune responses. In particular, BRAF mutations in melanoma render melanoma cells dependent on the glycolytic pathway to obtain energy and building blocks required for the high proliferative rates (Haq et al., 2013). Moreover, the tumor microenvironment can significantly impact the recruitment of immune cells to the tumor site. For instance, in pancreatic cancer, enhanced secretion of lactate by tumor cells results in impaired host immune response to tumor cells (Husain et al., 2013). Intriguingly, melanoma patients with high lactate dehydrogenase activity (LDH) have poor clinical outcome. The main goal for our next URPP term is to better understand the metabolic requirements of melanoma formation and progression, to determine whether resistance to targeted therapies involves metabolic switches, and to address whether changes in the tumor’s metabolome influence type and extent of infiltrating immune cells.
- Evaluate the effects of Yy1 knockdown and BRAF/MEK inhibitors in melanoma cell proliferation and invasion in vivo.
A vast body of research in resistance associated to BRAF-targeted therapy implies that BRAF inhibitors might promote the dissemination of melanoma cells to new locations. In this aim we will evaluate the effects of BRAF/MEK inhibitors in melanoma cell dissemination by xenotransplantation experiments, in which animals are subjected to a BRAF/MEK inhibitor regime. These studies will be done in parallel to experiments involving conditional and inducible Yy1 genetic manipulation in vivo.
- Identify metabolic changes induced by Yy1 loss of function and BRAF/MEK inhibitors.
This aim will be achieved by mass spectrometry-based metabolomics and will be performed in collaboration with Prof. Nicola Zamboni at the ETH Zurich. We plan to perform a comprehensive and quantitative analysis of a wide array of metabolites in both human melanoma cell lines either subjected to Yy1 knockdown or treated with BRAF/MEK inhibitors. In addition, we will use patient biopsies isolated prior and shortly after initiation of targeted therapy.
- Determine the effect of metabolic inhibitors targeting different metabolic pathways in melanoma proliferation and invasion both in vitro and in vivo.
To achieve this aim we will test several metabolic inhibitors targeting various pathways including glycolysis, TCA cycle and mitochondrial oxidative phosphorylation in human melanoma cell lines. As a main readout we will focus on melanoma proliferation, invasion and apoptosis. In a later phase of the project we will test some of the inhibitors in vivo both in immunocompromized mice as well as in the melanoma mouse model TyR:: N-Ras Q61K Ink4a-/-.
- Analyze whether changes in tumor microenvironment associated with abundance of different metabolites impair recruitment of immune cells to the tumor site.
This objective will be performed in collaboration with the new assistant professor to be hired in the tumor microenvironment field. We plan to determine whether immune surveillance is altered upon ablation of metabolism-related genes in the melanoma mouse model TyR:: N-Ras Q61K Ink4a-/-.
- Identify whether Yy1 regulates proliferation and or invasion in other cancer types.
To address this aim we will recur to the newly established human cancer cell lines deposited in the URPP tumor cell biobank in the context of the present URPP. We will concentrate on squamous cell carcinoma, prostate, and colon cancer and initially focus on the effect of Yy1 knockdown on proliferation, EMT signatures, and metabolic changes.