Fish lead to new oncogene and potential treatment
March 23, 2011
Researchers at Children's Hospital Boston and collaborators from several other institutions have utilized zebrafish to identify two new important mechanisms that promote the growth of the deadly human skin cancer melanoma. In one of two papers featured on the cover of the March 24 issue of Nature, the scientists established SETDB1 as a new oncogene in melanoma, the first to be identified using a zebrafish model of the disease. In the second paper, the same Children's researchers utilized the melanoma-prone zebrafish to determine that the combination of an existing arthritis drug and a new drug currently under study for late-stage melanomas with mutations in the gene BRAF may hold promise in treating some types of melanoma.
Both studies relied on a zebrafish melanoma model developed by the laboratory ofLeonard Zon, MD, director of the hospital's Stem Cell Program, a Howard Hughes Medical Institute investigator, and senior author of both papers. "We developed the zebrafish melanoma model about six years ago," said Zon. "The purpose of the model was to find new genes that caused human melanoma and support the development of new drugs against a disease for which there are very few treatment options."
Melanoma is an aggressive skin cancer that begins in melanocytes, the pigment-producing cells responsible for skin color. While melanoma accounts for less than five percent of skin cancer cases, its numbers are rising, and it has a poor prognosis when diagnosed late. In the United States alone, there were about 68,000 new cases and 8,700 deaths in 2009.
Normal zebrafish (top), zebrafish with melanoma (bottom)
Mutations in the BRAF gene, a core characteristic of the zebrafish melanoma model, are present in about 50 to 60 percent of human melanomas. But BRAF mutations are also found in benign moles and are not themselves sufficient to cause cancer; other mutations must be present as well. In the study "The histone methyltransferase SETDB1 is recurrently amplified in melanoma and accelerates its onset," the researchers set out to pinpoint other candidates in a region of chromosome 1 called 1q21, which contains a stretch of 54 genes that is amplified in about 30 percent of melanoma patients. Of the 54, the gene SETDB1 stood out as the only one in this region to work with BRAF to fuel tumor development.
"Along with being a creative approach, it was truly a brute force scientific effort to home in on SETDB1," Zon noted. "We looked at each of the genes in this region one by one, and between discovery and validation ultimately assayed more than 2,100 tumors from more than 3,100 fish."
SETDB1 encodes an enzyme that helps turn other genes on or off, and is overactive in numerous other tumors (e.g., ovarian, breast, and liver cancers). Because in the model the level of a tumor's malignancy rose with the level of SETDB1 activity, the gene could be a valuable target for prognostic testing or for designing new melanoma treatments.
BRAF abnormalities also led to the recognition that zebrafish on the way to developing melanoma harbor excess numbers of immature "embryonic" cells called neural crest cells, raising the fishes' risk of later cancer formation. Zon and a second research team used the zebrafish to screen 2,000 chemicals for candidates that would suppress these excess embryonic neural crest cells. In the paper "DHODH modulates transcriptional elongation in the neural crest and melanoma," Zon and his colleagues found that a compound that interfered with dihydroorotate dehydrogenase (DHODH), an enzyme involved in neural crest cell development, showed promise.
The team turned to leflunomide, a DHODH-inhibitor approved to treat forms of arthritis. In the zebrafish model, leflunomide knocked down expression of a number of genes overexpressed in both melanomas and neural crest cells, while in rats it prevented neural crest stem cells from renewing themselves. Leflunomide also stopped the growth of cultured cells from human melanomas and caused regression of human tumors transplanted into nude mice.
According to Zon, "We realized that a combined blockade of DHODH and BRAF would cooperate to suppress melanoma growth by targeting both the fate and the growth of melanoma precursors." The researchers then tested in mice a combination of leflunomide and a BRAF inhibitor developed by Plexxikon, which is in late-stage clinical trials. Together, the combination of drugs led to an almost complete abolition of tumor growth; 40 percent of the mice experienced a near complete tumor regression.
"The combination of the two drugs was more effective than either drug alone, and allowed us to use lower doses of each," said Zon, who has begun to plan a clinical trial of the combination treatment. "It will be interesting to put them into the clinic together."
Collaborator Affiliations Affiliations for collaborators on the SETDB1 study are as follows: Children's Hospital Boston (Stem Cell Program and Department of Hematology/Oncology), Howard Hughes Medical Institute, Harvard Stem Cell Institute, Massachusetts General Hospital Cancer Center, Dana-Farber Cancer Institute (Center for Cancer Genome Discovery and Departments of Medical Oncology and Cancer Biology), The Broad Institute, Whitehead Institute for Biomedical Research, Brigham and Women's Hospital (Center for Molecular Oncologic Pathology), Universitá Paris-Diderot (UMR 7216 Epigánátique et Destin Cellulaire), and Sapienza University of Rome (A. Rossi Fanelli Biochemical Sciences Department.)
Affiliations for collaborators on the DHODH study are as follows: Children's Hospital Boston (Stem Cell Program and Department of Hematology/Oncology), Howard Hughes Medical Institute, Harvard Stem Cell Institute, Dana-Farber Cancer Institute (Departments of Medical Oncology and Biostatistics and Computational Biology), University of East Anglia, University of Michigan (Center for Stem Cell Biology), Whitehead Institute for Biomedical Research, Massachusetts Institute of Technology (Department of Biology), Harvard University, Cambridge University, Genzyme Corporation, and Brigham and Women's Hospital (Department of Pathology).
Funding Funding for the DHODH study was supported by grants from Howard Hughes Medical Institute, National Cancer Institute, Aid for Cancer Research, American Society for Clinical Oncology, National Institute of Arthritis and Musculoskeletal and Skin Diseases, and a Pfizer Industrial CASE award through the Biotechnology and Biological Sciences Research Council.
Funding for the SETDB1 study was supported by grants from Damon Runyon Cancer Research Foundation, Charles A. King Trust Foundation, and a Young Investigator Award from the American Society of Clinical Oncology, Canadian Institutes of Health Research, and National Institutes of Health.
Disclosures Leonard Zon, MD, is a founder and stockholder of Fate Therapeutics, Inc.www.fatetherapeutics.com and a scientific advisor for Stemgent.
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