What does a common arthritis medication have to do with skin cancer? More than you might think.
Researchers at Children's Hospital Boston have found a promising link between the anti-inflammation drug leflunomide, often prescribed for arthritis, and treatment for melanoma. In a recent issue of Nature, they write that leflunomide could become an affordable treatment for the deadly disease — a valuable resource given that incidence of skin cancer is on the rise around the world, and has traditionally been very expensive to treat.
So when is this possible wonder cure coming? The answer starts with genetically-modified zebrafish and drug patents.
Using zebrafish to model human melanoma
For the last five years, researchers in the laboratory of Dr. Leonard Zon, a Howard Hughes Medical Institute investigator and Director of the Stem Cell Program at Children's Hospital Boston, have been using genetically modified zebrafish to investigate melanoma, the most aggressive form of skin cancer. Dr. Richard White, a post-doctoral researcher in the Zon lab and lead author of the paper, explains the attractiveness of the zebrafish model:
The zebrafish initially began to be studied because of its strengths in understanding the genetics of developmental biology. With the realization that the fish was highly amenable to transgenic approaches, it made sense to develop models of cancer in the fish which could then be used as a platform for so-called "modifier screens," in which you look for perturbations that [alter] the propensity of the transgenic fish to develop cancer.
Not only do these fish develop cancers that are very similar to human cancer, they are "amenable to genetic and chemical screens that are incredibly difficult if not impossible in mice or other models," explains White.
Their studies incorporate zebrafish like the bottom one in the image on the left. This fish has been genetically altered to to express the human oncogene BRAF(V600E), the most common mutation in human melanoma patients. The top fish is representative of a wild type (genetically standard) zebrafish.
Melanoma is a tumor of transformed melanocytes, which are originally derived from the neural crest – a cell population unique to the vertebrate embryo that can differentiate into a variety of different cell types, including craniofacial cartilage and bone; smooth muscle; and pigment-producing melanocytes.
In their paper, Drs. White, Zon, and their colleagues show that the differentiation of neural crest progenitors is disrupted in the embryos of the genetically modified zebrafish; the neural crest progenitors fail to differentiate into normal melanocytes, and instead remain in a progenitor-like state. Further investigation led the researchers to conclude that "the majority of human melanomas reflect events that lead to the maintenance of a neural crest progenitor phenotype."
Bearing this in mind, the team reasoned that chemical suppressors of neural crest progenitors would be useful for treating melanoma. Dr. White headed up a chemical genetic screen of 2,000 compounds to identify ones that could inhibit neural crest stem cells in the zebrafish model.
When White combined the findings of his screen with the research of UEA scientists Grant Wheeler and Matt Tomlinson (who had been conducting related chemical screens with tadpoles), they discovered that inhibitors of an enzyme called dihydroorotate dehydrogenase (DHODH), including leflunomide, "led to an almost complete abrogation of neural crest development in zebrafish and to a reduction in the self-renewal of mammalian neural crest stem cells."
Treating cancer with arthritis medication
Given the effect of DHODH-inhibition on neural crest development, the researchers decided to test leflunomide's effects on the growth of various melanoma cell lines in vitro.
Leflunomide's effects, which are illustrated in the figure to the left, were dramatic; administration of A771726 (also known as Teriflunomide, the active metabolite of leflunomide) triggered a distinct dose-dependent decrease in the proliferation of several human melanoma cell lines in vitro. All three of the cell lines contain the BRAF(V600E) mutation.
While administration of leflunomide alone led to a marked decrease in melanoma growth, its effect was even more dramatic when it was administered in combination with a specific inhibitor of the BRAF(V600E) oncogene.
PLX4720, a BRAF enzyme inhibitor developed by Plexxikon, has been shown to cause programmed cell death in melanoma cell lines with the BRAF(V600E) mutation, which is present in the majority of melanomas. PLX4720's analogue, PLX4032, is currently in phase three clinical trials for the treatment of late-stage melanoma. This figure illustrates the combined effect of leflunomide and PLX4720 in vitro.
The researchers went on to examine the in vivo effects of leflunomide and PLX4720 by using xenografts of human melanoma cells transplanted into nude mice. To summarize, the combined administration of PLX4720 and leflunomide led to a dramatic suppression of tumor growth, and resulted in almost complete tumor regression in 40% of animals. The images of the xenograft transplants on the left demonstrate their findings, which are explained in all their technical glory below.
At 12 days post treatment, tumours in mice that had been treated with dimethylsulphoxide (DMSO) as a control had grown 7.4+/-1.3 fold. By contrast, tumours in PLX4720-treated mice had grown 5.7+/-0.16-fold, and those in leflunomide-treated mice had grown 4.7+/-0.12 fold. The combination of PLX4720 and lefluno- mide led to an enhanced abrogation of tumour growth, with only 2.2+/-0.9 fold growth. In 40% of animals, this combination led to almost complete tumour regression. Therefore, we have found that an inhibitor of embryonic neural crest development, leflunomide, blocks in vivo tumour growth in combination with the BRAF(V600E) inhibitor PLX4720 when used at clinically meaningful doses.
So what's next?
Unlike most other cancers, the incidence of melanoma is increasing. The fact that leflunomide is already FDA-approved to treat arthritis bodes well for its expedited approval as a much-needed melanoma treatment.
"This is a really exciting discovery – making use of an existing drug specifically to target melanoma," said Dr. Grant Wheeler, head of the research team at the UEA.
"Deaths from melanoma skin cancer are increasing and there is a desperate need for new, more effective treatments. We are very optimistic that this research will lead to novel treatments for melanoma tumours which, working alongside other therapies, will help to stop them progressing."
Dr. White seems equally optimistic about the future of the drug.
"If the BRAF inhibitor [Plexxikon's PLX4032] is approved in the near future as expected, then [green lighting] a trial using two FDA approved drugs is not that difficult," said White, who told us he and Dr. Zon are currently in the early stages of establishing a clinical trial for leflunomide in combination with a BRAF inhibitor like PLX4032.
"We personally think [leflunomide] should be tested in combination with the BRAF inhibitor since it showed a stronger effect."
In 2005, the FDA granted approval for the production of a generic version of Arava — the trade name for leflunomide — which has been marketed by Sanofi-Aventis since 1998. This means that if clinical trials of leflunomide were to be carried out successfully, there could soon be a highly-effective melanoma treatment available at affordable prices in as little as three to five years — an incredible resource given the increasing incidence of this deadly form of cancer.
Images of wild type and transgenic zebrafish generously provided by the Zon Lab
Graphs depicting effects of leflunomide and PLX4720 on melanoma via Nature, with authors' permission. Top photo via Shutterstock.
For further reading
Original scientific paper published in Nature
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