News & Analysis

Leonard Zon from the Howard Hughes Medical Institute, Boston, MA, USA, and a team of collaborators have used the zebrafish to identify two new potential targets for melanomas.

Research to identify the cause of what turns pigmented cells cancerous discovered the BRAF gene as a target for the treatment of melanomas as it drives the proliferation of melanocytes. In a previous study, Zon put BRAF into a zebrafish that had been genetically altered to lack the tumor suppressor gene p53. He and his team observed that the zebrafish’s stripes became darker and cancerous on many occasions but they also observed that BRAF mutations occurred in benign moles too.

Now, Zon and his coworkers have again used the zebrafish to investigate possible therapeutic targets of melanoma and have identified two.

The research team identified 17 genes that were amplified in melanoma samples by observing a region of human chromosome one. Members from Zon’s group created a device that carried the 17 genes into melanoma-probe zebrafish one by one and then analyzed tumors from more than 3000 fish. SETDB1, a gene that is overexpressed in ovarian and breast cancers, was found to be the only one of the 17 that accelerated melanoma and was upregulated in 70% of the tumors when investigated on human cell lines. The gene expresses an enzyme that modifys the DNA-bearing chromatin. The group hopes that this enzyme can now be used as a target for future therapies to treat melanoma.

The researchers’ second published study identified another therapeutic target by studying the embryos of zebrafish that had been genetically modified to be more susceptible to developing melanomas. A set of 127 genes was found to be misexpressed in melanomas and could be used to predict which fish embryo would develop melanoma. The genes are important in a special stem cell called the embryonic neural crest cell. This cell differentiates during development and is responsible for melanocytes and connective tissue. The genetically altered zebrafish had more of the embryonic neural crest cells. The group screened 2000 molecules in order to identify chemical entities that perturbed the development of these cells. The team’s database implied that one of the molecules could inhibit the enzyme dihydroorotate dehydrogenase (DHODH), which produces uridine, one of the four RNA nucleic bases, and is involved in transcribing DNA into RNA. The database identified leflunomide, a drug currently used to treat rheumatoid arthritis, as the potential inhibitor of DHODH.

They then tested the drug on human cell lines and zebrafish and found selectively blocked transcriptional elongation. Mice models were then used to further validate the potential use of the drug. Zon and coworkers combined leflunomide with a BRAF inhibitor, currently entering late-stage clinical trials for metastic melanoma. Alone, both the BRAF inhibitor and leflunomide were successful, but in combination they worked better, completely destroying 40% of the tumors. The researchers hope that this combination therapy may enter human clinical trials in the coming year and believe that lower doses can be used than are currently prescribed for the treatment of melanoma and rheumatoid arthritis in order to minimize the side effects.

Zon and his colleagues believe that leflunomide could have other applications given that it halts some genes that are controlled by myc, an oncogene associated with many forms of cancer. The group intends to continue its research into cancer treatments using the zebrafish as their model believing that the work carried out on SETDB1 could be applied to other forms of cancer.

Sources: Ceol CJ, Houvras Y, Jane-Valbuena J et al. The histone methyltransferase SETDB1 is recurrently amplified in melanoma and accelerates its onset. Nature 471, 513–517 (2011); White RM, Cech J, Ratanasirintrawoot S et al. DHODH modulates transcriptional elongation in the neural crest and melanoma. Nature 471, 518–522 (2011).

Researchers based at the Pfizer Global Research and Design facility in St Louis (MO, USA) and BIOCUS Life Science Inc. Wakefield (MA, USA), utilized high-throughput MS technology to measure sphingosine-1-phosphate formation in human whole blood. The purpose of the research was to develop an assay to detect compounds that inhibit sphingosine-1-phosphate signaling.

Sphingosine-1-phosphate is a bioactive sphingolipid metabolite and is an important lipid mediator. The metabolite is formed by phosphorylation of sphingosine, catalyzed by sphingosine kinase, and has been detected in organisms such as mammals, flies, yeasts, plants and worms. Sphingosine-1-phosphate has been implicated in numerous biological processes and has been previously suggested to regulate cell growth.

The researchers used MS to detect the conversion of a 17-carbon-long variant of sphingosine, C17SPH, which was artificially added to human blood, into the sphingosine-1-phosphate analogue, C17S1P. Novel procedures were developed to achieve homogenous mixing of whole blood in 384-well plates, and to extract sphingosine-1-phosphate from blood in 96- and 384 plates with minimal manipulation prior to analysis. The high-throughput assay was performed using the RapidFire^® ms system.

Source: Highkin MK, Yates MP, Nemirovskiy OV et al. High-throughput screening assay for sphingosine kinase inhibitors in whole blood using RapidFire^® Mass spectrometry. J. Biomol. Screen. 16(2) 272–277 (2011).

Margaret Hamburg, the US FDA Commissioner, has warned Congress that cutting the FDA’s funding for the rest of fiscal 2011 would have a “devastating” effect. Addressing the House Appropriations Agriculture, Rural Development, FDA and Related Agencies Subcommittee in March 2011, Hamburg said that, “(the impact) will be significant. We will be unable to do inspections domestically and internationally at the level we need to be doing.”

Congress plans to cut over US$200 million from the FDA’s funding, an amount that Hamburg claims equates to the budget of one of agency’s centers. The House approved the cuts in February 2011 when it voted 235–189 on H.R. 1, the continuing resolution that would fund the FDA through fiscal 2011 at $3.31 billion, a reduction of $241 million from the FDA’s 2010 appropriations.

Hamburg’s warning comes in a month of uncertainty for the FDA. On 17 March 2011, the Senate voted to approve another temporary funding measure to keep the government agency running until April 8th 2011.

Source: FDA Newsletter – www.fdanews.com/newsletter/article?articleId=134983&issueId=14544

A UK task force assembled to minimize the adverse effects of the closure of Pfizer’s site in Sandwich, UK, is in discussions with CROs that may step in to hire several hundred of the 2400 workers being laid off.

Pfizer hopes the CROs will keep the research unit in operation, with the company having already signed confidentiality agreements with ten firms that have indicated an interest in continuing research at the site. The task force is thought to be seeking to identity tax and planning breaks for companies moving into the research hub.

“Our report focuses on the opportunities presented by the world-class skills and facilities built by Pfizer over the last 60 years”, commented one of the task force leaders. “Combined, they offer a unique asset on which to build a new model of high-tech growth and employment, which would be of importance not only to Kent, but to the country”.

The Sandwich Economic Development Task Force was set up in February with backing from the UK government’s Department for Business, Innovation and Skills.

Pfizer’s plans to use more CROs to take on development work follows similar moves taken by other companies, notably Eli Lilly, sanofi-aventis, GlaxoSmithKline.

Source: Pfizer talking with CROs to fill R&D hub: www.centerwatch.com/news-online/headline-details.aspx?HeadlineID=1206

Researchers from the University of Notre-Dame (USA) and Cornell University (USA) have published their findings into the potential of a new treatment of Niemann–Pick Type C (NPC), a disease dubbed ‘childhood Alzheimer’s’ by the NIH.

Niemann–Pick Type C is only observed in one in 150,000 children and is an inherited cholesterol-metabolism disorder. This genetic flaw stops cells from using lipids, instead holding them inside the cell. Brain cells are particularly affected and this often leads to children with NPC dying by the time they enter their teenage years.

The research group, led by Anton Meinhart, investigated the use of histone deacetylase inhibitors in the treatment of the disorder and found that they had a profound effect on the genetic flaw on the NPC1 and NPC2 genes. The team observed that the NPC cells became completely indistinguishable from normal cells following the administration of a histone deacetylase inhibitor. This class of drugs has been associated with both rare diseases, such as NPC, as well as leukemia where they increase bone marrow cell numbers.

The group hopes that the results can be confirmed in vivo and believe that, with histone deacetylase inhibitors already in clinical trials, this will accelerate the development of treatments for NPC.

Source: Pipalia NH, Cosner CC, Huang A et al. Histone deacetylase inhibitor treatment dramatically reduces cholesterol accumulation in Niemann–Pick type C1 mutant human fibroblasts. PNAS doi:10.1073/pnas.1014890108 (2011) (Epub ahead of print).

Researchers form the Max Planck Institute for Medical Research, Heidelberg, Germany, have revealed the mechanism by which zeta toxins carry out cell death. The zeta toxins are a family of proteins that are present when pathogens undedergo stress and ‘commit suicide’.

Toxin–antitoxin (TA) systems are harbored by most bacteria. This system allows a bacterial toxin to exist under normal conditions being prevented from becoming active by its counterpart antitoxin. When the bacterium undergoes stress the antitoxin is degraded and allows the toxin to attack its host from within. The research group studied the TA system of Pneumococcal epsilon zeta Antitoxin Toxin (PezAT) and its counterpart the Pneumococcal epsilon zeta Toxin (PezT) in Escherichia coli. This system is found in Streptococcus pneumoniae, which is responsible for pneumonia and meningitis. E. coli cells that had PezT activated showed similar traits to those treated using penicllin. The middle of the division stage was halted first and the intersection zone between the two cell bodies burst and the cell died. This was because PezT transforms UDP-N-acetylglucosamine (UNAG), an essential sugar building unit, into UNAG-3P, a toxic molecule that inhibits growth of the cell wall.

UNAG-3P is now being considered as a starting point in the development of novel antibiotic drugs. The group hopes that better understanding of the mechanism will lead to development of mimics of the system as a strategy for combating bacterial resistance.

Source: Mutschler H, Gebhardt M, Shoeman RL, Meinhart A. A novel mechanism of programmed cell death in bacteria by Toxin–Antitoxin systems corrupts peptidoglycan synthesis. PLoS Biol doi:10.1371/journal.pbio.1001033 (2011) (Epub ahead of print).