Melanoma account for <10,000 deaths annually in the United States alone and current therapies for advanced-stage melanoma do not provide durable clinical benefit. Therefore, improved molecular understanding of melanoma initiation and progression is essential for developing effective therapies against melanoma. Metabolic deregulation due to the alterations in metabolic pathways is one of the key hallmarks of cancer cells. However, the role of metabolic alterations in melanoma has neither been comprehensively analyzed nor fully understood. The central hypothesis is that metabolic alterations are crucial regulators of melanoma initiation, progression and therapy response. Therefore, to identify metabolic drivers of melanoma, we developed an integrative genomics approach by combining patient-derived melanoma sample gene expression profiling with the functional genomics approach of an in vivo RNAi screening and identified N-Acylsphingosine amidohydrolase (ASAH1) as a gene necessary for melanoma growth and metastasis. The overall objective is to determine the role of ASAH1 in melanoma initiation and progression, ascertain its mechanisms-of-action and determine its role in oncogenic BRAF inhibitor vemurafenib resistance. Specifically, in Aim 1, we will determine the role of ASAH1 in melanoma initiation and progression and ascertain whether ASAH1 is necessary for oncogenic BRAF- and NRAS-induced transformation and tumor progression. To do so, we will use cell culture-based models of immortalized human melanocytes, established and patient-derived short-term melanoma cultures as well as mouse models of melanomagenesis and melanoma metastasis.
In Aim 2, we will determine the mechanism(s) of ASAH1 action. First, we will perform structure-function analyses to determine whether the lipid hydrolase activity of ASAH1 is necessary for its ability to promote melanoma initiation and progression. Additionally, guided by our preliminary results, we will test whether ASAH1 promotes melanoma initiation and progression by reducing cellular ceramide level and/or by activating MAP kinase pathway. The experiments will include biochemical, genetic, cell culture and in vivo mouse model-based approaches.
In Aim 3, we will determine the role of ASAH1 in oncogenic BRAF inhibitor vemurafenib resistance. We will test whether inhibition of ASAH1 can restore vemurafenib sensitivity in BRAF mutant melanoma cells and determine if ASAH1 induced vemurafenib resistance depend upon its ability to regulate cellular ceramide level and/or MAP kinase pathway. Finally, using melanoma patients-derived short-term melanoma culture and primary sample of pre- and post-vemurafenib treatment we will establish the clinical significance of our results. Collectively, the results of ur experiments will uncover novel driver genes and pathways of melanoma initiation and progression, improve the molecular understanding of melanoma and reveal new therapeutic targets and mechanisms of resistance to melanoma therapies.

Public Health Relevance

Melanoma account for over 85% of skin cancer-related deaths and due to the lack of effective therapies only 15% advanced stage melanoma patients can expect to survive beyond 5 years. Using an integrative genomic approach, we have identified metabolic vulnerabilities of melanoma cells. The results of our experiments will reveal novel genes and pathways that drive melanoma initiation and progression, improve our molecular understanding of melanoma and ascertain the utility of targeting specific metabolic alterations for melanoma therapy and their role in determining resistance to targeted therapies in melanoma.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Exploratory/Developmental Grants (R21)
Project #
5R21CA191364-02
Application #
8985670
Study Section
Special Emphasis Panel (ZCA1)
Program Officer
Yassin, Rihab R
Project Start
2014-12-15
Project End
2016-11-30
Budget Start
2015-12-01
Budget End
2016-11-30
Support Year
2
Fiscal Year
2016
Total Cost
Indirect Cost
Name
Yale University
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
043207562
City
New Haven
State
CT
Country
United States
Zip Code
Gupta, R; Yang, Q; Dogra, S K et al. (2017) Serine hydroxymethyl transferase 1 stimulates pro-oncogenic cytokine expression through sialic acid to promote ovarian cancer tumor growth and progression. Oncogene 36:4014-4024
Forloni, Matteo; Ho, Thuy; Sun, Lisha et al. (2017) Large-Scale RNA Interference Screening to Identify Transcriptional Regulators of a Tumor Suppressor Gene. Methods Mol Biol 1507:261-268
Nagarajan, Arvindhan; Dogra, Shaillay Kumar; Sun, Lisha et al. (2017) Paraoxonase 2 Facilitates Pancreatic Cancer Growth and Metastasis by Stimulating GLUT1-Mediated Glucose Transport. Mol Cell 67:685-701.e6
Gupta, Romi; Wajapeyee, Narendra (2017) Transcriptional Analysis-Based Integrative Genomics Approach to Identify Tumor-Promoting Metabolic Genes. Methods Mol Biol 1507:269-276
Forloni, Matteo; Gupta, Romi; Nagarajan, Arvindhan et al. (2016) Oncogenic EGFR Represses the TET1 DNA Demethylase to Induce Silencing of Tumor Suppressors in Cancer Cells. Cell Rep 16:457-471
Nagarajan, Arvindhan; Malvi, Parmanand; Wajapeyee, Narendra (2016) Oncogene-directed alterations in cancer cell metabolism. Trends Cancer 2:365-377
Gupta, Romi; Forloni, Matteo; Bisserier, Malik et al. (2016) Interferon alpha-inducible protein 6 regulates NRASQ61K-induced melanomagenesis and growth. Elife 5:
Nagarajan, Arvindhan; Petersen, Max C; Nasiri, Ali R et al. (2016) MARCH1 regulates insulin sensitivity by controlling cell surface insulin receptor levels. Nat Commun 7:12639