Research findings signpost way to cancer drug targets
28th February 2006
By Staff Writer
Researchers supported by a grant from the US National Cancer Institute have made findings that hold promise for identification of new drug targets to treat a group of highly lethal cancers known as hedgehog cancers.
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The findings of Dr Charles Emerson, a senior scientist and director of the not-for-profit Boston Biomedical Research Institute (BBRI) and Dr Natalia Riobo, a postdoctoral fellow at the University of Pennsylvania School of Medicine, were published in the journal Cancer Research.

The research focuses on understanding a type of cell communication known as hedgehog signaling. Hedgehog signaling has an essential role in the control of stem cell growth and in the promotion of tumor growth.

Excessive activation of the hedgehog signaling pathway has been observed to be responsible for tumor growth in the most lethal of human cancers, including pancreatic, lung, skin, muscle and digestive tract cancer - all of which remain largely untreatable.

The researchers' findings show that two kinases, known as PKC-delta and MEK-1, cooperate with hedgehog signaling to activate GLI transcription factors to regulate key genes that control stem cell and tumor growth.

"Understanding the key components of the hedgehog signaling pathway, and how mutations in this pathway can lead to the growth of cancer cells, highlights the power of basic science and disease model research to reveal new therapeutic approaches for human disease, which is a hallmark of BBRI research," says BBRI's director Dr Charles Emerson. "Our preliminary findings offer tremendous promise for development of preventative and curative treatments for these devastating cancers."
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Helsingin yliopisto (University of Helsinki) 07.02.2006
Cancer researchers found a new mechanism potentially explaining evolution of signalling pathways


Cancer researchers at the University of Helsinki, Finland, in trying to find a novel tumor suppressor gene, instead found an important evolutionary change that occurred in a key developmental signalling pathway. The finding suggests a potential mechanism for evolution of complex intercellular signalling pathways. The results are published in today’s issue of the journal Developmental Cell.

A relatively small number of evolutionary conserved genes are responsible for controlling the development of the diverse range of animal species. Most of these genes have been originally identified in fruit fly, based on the analysis of mutations that alter the body pattern of a developing embryo.

During embryonic development, cells regulate the growth and differentiation of each other by secreting extracellular signalling molecules (growth factors or morphogens), which bind to receptors present on the surface of other cells. The receptors in turn activate intracellular signalling pathway composed of proteins that relay the signal to the nucleus, activating specialized proteins called transcription factors. The transcription factors then affect expression of genes that induce cell growth and differentiation.

The signal transduction molecules and mechanisms of major developmental signalling pathways are thought to be evolutionary conserved between invertebrates and vertebrates in such a way that if a signalling pathway is present in a given organism, it includes all the major classes of components found in humans. Because of the lack of intermediate forms, the evolution of these complex signalling pathways is not understood in detail, and the emergence of signalling pathways with multiple specific and essential components has even been used as an argument against evolution.

Because multiple components of the Hedgehog (Hh) signalling pathway are defective in human cancers, Markku Varjosalo in Professor Jussi Taipale’s laboratory (the University of Helsinki and National Public Health Institute of Finland) cloned the gene for mammalian homolog of a key regulator of fruit fly Hh signalling pathway, Costal-2. However, further analysis of the function of the mammalian gene revealed that it did not function as a Hh pathway regulator, let alone as the tumor suppressor gene the researchers had hoped for. Instead, together with a group led by Prof. Rune Toftgård and Dr. Stephan Teglund from Karolinska Institutet, the researchers found that another gene (Suppressor of Fused), which has a minor role in Hh signalling in fruit fly is critical for Hh pathway regulation in mammals.

The finding is the first clear demonstration of a major difference in the function of conserved signalling pathways between species. The results also show that multi-component pathways evolve, in part, by the insertion of novel proteins between existing pathway components. This insertion mechanism can potentially explain a challenging aspect of evolutionary biology regarding the emergence of signalling pathways with multiple specific components.

Loss of Hh pathway activity can cause a variety of human birth defects, such as polydactyly, cyclopia and holoprosencephaly. On the other hand, inappropriate activation of the Hh signaling pathway can cause multiple types of human cancer, including the most common type of cancer in Caucasians (basal cell carcinoma) and the most common brain tumor in children (medulloblastoma). Therefore, the study of the Hh signalling pathway is important for the understanding of and developing cures for human disease.

The work was supported by the Academy of Finland, Biocentrum Helsinki, The University of Helsinki, the Sigrid Juselius Foundation, and Finnish Cancer Organizations.
Article

Discovery by Research Group at Boston Biomedical Research Institute Opens Doors to New Treatments for the most Lethal Cancers

WATERTOWN, Mass. - February 27, 2006 - Cancer Research, a top tier journal that publishes biologically significant findings with relevance to cancer, recently published the findings of Dr. Charles P. Emerson, Jr., a Senior Scientist and Director of the not-for-profit Boston Biomedical Research Institute and Dr. Natalia Riobo, a postdoctoral fellow at the University of Pennsylvania School of Medicine. The discovery holds promise for identification of new drug targets to treat a group of the most lethal cancers known as Hedgehog cancers.

Drs. Emerson and Riobo’s research focuses on understanding a type of cell communication know as Hedgehog signaling. Hedgehog signaling has an essential role in the control of stem cell growth and in the promotion of tumor growth. Excessive activation of the Hedgehog signaling pathway has been observed to be responsible for tumor growth in the most lethal of human cancers, including pancreatic, lung, skin, muscle and digestive tract cancer – all of which remain largely untreatable. These researchers now show that two kinases, known as PKC-deltaand MEK-1, cooperate with Hedgehog signaling to activate GLI transcription factors to regulate key genes that control stem cell and tumor growth.

“Understanding the key components of the Hedgehog signaling pathway, and how mutations in this pathway can lead to the growth of cancer cells, highlights the power of basic science and disease model research to reveal new therapeutic approaches for human disease, which is a hallmark of BBRI research,” says BBRI’s Director Dr. Charles Emerson. “Our preliminary findings offer tremendous promise for development of preventative and curative treatments for these devastating cancers.” The work is being supported by a grant from the National Cancer Institute.

Boston Biomedical Research Institute is a not-for-profit institution dedicated to the understanding, treatment and prevention of specific human diseases including cancer, cardiovascular disease, Alzheimer’s disease, muscular dystrophy, diabetes and conditions such as obesity and reproductive health problems. For more information visit www.bbri.org.

Media Contact: Terence McGowan, 617-658-7707, mcgowan@bbri.org.

Study Sheds Light on Signaling Mechanism in Stem Cells, Cancer

By: UCSF on Mar 21 2006 22:42:41

Cancery therapeutic strategy
UCSF scientists have illuminated a key step in a signaling pathway that helps orchestrate embryonic development. The finding, they say, could lead to insights into the development of stem cells, as well as birth defects and cancers, and thus fuel therapeutic strategies.

The study, reported in Nature (Oct. 13, 2005), focuses on the Hedgehog family of signaling molecules, which play a central role in directing development of the early embryo's growth and spatial plan, as well as its later organ and limb development. Defects in Hedgehog signaling are a significant cause of some birth defects and cancers.

Secreted from one cell, a Hedgehog signal shoots to the surface receptor of a second cell, and then, in a rapid-fire succession of biochemical reactions, relays a message into the cell's nucleus. There, it issues an instruction, prompting the cell to divide, or specialize into a particular cell type, or migrate to help form another part of the embryo, and so on. This transaction, known as signal transduction, is a ceaseless activity of embryonic development.

Scientists have long known that Hedgehog signaling requires the activity of a protein known as Smoothened. This has been demonstrated in animals ranging from insects to humans.

They have also known that defects in Smoothened, which only functions within Hedgehog signaling, are responsible for some cases of human cancers, most prominently a skin cancer known as basal cell carcinoma and a childhood brain cancer known as medulloblastoma -- as well as some birth defects.

However, they have not known how Smoothened executes its function, nor where it is located in the target cell.

Now, through a series of studies conducted in several types of cells in culture, and in zebrafish and mouse embryos, the UCSF scientists have answered both questions. In the process, they have revealed the critical role of a cellular component that until now has been a mystery: an antenna-like structure attached to cells known as the primary cilium.

The primary cilium, it turns out, serves as the fulcrum in a series of acrobatic like moves between the Hedgehog signal and the Smoothened protein. Once Hedgehog has latched on to its receptor on the target cell's surface, it prompts the cell to move Smoothened, located in vesicles around the cell's nucleus, to the primary cilium. The positioning of Smoothened on the cilium, in turn, prompts downstream signaling of Hedgehog signals into the nucleus, where the instructions are issued.

Just how or what the primary cilium is doing to promote Smoothened's activity is not clear, say the researchers. However, its involvement in the process is a revelation.

Scientists elsewhere reported in Nature in (Nov. 6, 2003) that removal of the primary cilium from cells led to defects in neural patterning resulting from Hedgehog signaling. However, they didn't know why.

"This study takes two mysteries- how Smoothened functions and the role of the primary cilium- and suggests a mechanism by which they are connected," says the senior author of the study, Dr. Jeremy Reiter, a fellow in the UCSF Program in Developmental and Stem Cell Biology, which is part of the UCSF Institute for Stem Cell and Tissue Biology.

The implications for medical research, he says, are significant.

Hedgehog signals play an important role in prompting embryonic and adult stem cells to differentiate into some of the specialized cells that make up the body's tissues -- such as those of the brain, pancreas and skin. The new finding, says Reiter, will advance scientists efforts to use signaling molecules to direct the differentiation of embryonic stem cells in the culture dish, with the goal of using them to replace or replenish damaged tissues in patients.

The discovery could be particularly important for neural stem cell research, says Dr. Arnold Kriegstein, director of the UCSF Institute for Stem Cell and Tissue Biology. Kriegstein, a neural stem cell scientist, was not an author on the study.

"Hedgehog signaling plays a critical role in prompting the differentiation of neural stem cells into the various forms of neurons in the brain," he says. "The discovery of the importance of the cilium in Hedgehog signaling should significantly advance our understanding of the mechanisms involved," he says. The finding should fuel research into the causes of certain birth defects (such as holoprosencephaly and limb defects) and cancers, says Reiter. Smoothened is already known to be a proto-oncogene, a normal gene that, if mutated, is capable of causing cancers. But its close involvement with the primary cilium suggests that the latter may also be implicated, suggesting a possible target for therapy.

More broadly, says Reiter, the primary cilium's role in Hedgehog signaling indicates it is likely to function in other signaling pathways, as well.

The scientists moved in on the role of Smoothened and the primary cilium incrementally. First, driven by their interest in Smoothened, they set out to determine where it was expressed in the embryo. They did so by developing highly specific antibodies to the protein and applying them to the tissue of an eight-day mouse embryo. The study revealed that Smoothened was modestly upregulated in cells of the node, an important early organizer tissue within the mouse embryo, and was expressed predominantly along the primary cilium of these nodal cells. This was a significant surprise.

Second, to examine whether Smoothened's movement from vesicles around the nucleus to the cilium was regulated by Hedgehog signals, they carried out two studies, one involving cultured epithelial and fibroblasts cells expressing Smoothened, another involving a mouse embryo. In both cases, one set of cells was exposed to Hedgehog signals. Another set was exposed to cyclopamine, a drug that blocks Smoothened's function. In the cells exposed to the Hedgehog signals, Smoothened moved from the vesicles of the cell body to the cilium. In the cells exposed to cyclopamine, Smoothened was undetectable on the cilium.

Scientists have known that cyclopamine inhibits Hedgehog signaling and can prevent Hedgehog-dependent cancers from spreading. The demonstration that the drug affected Smoothened movement to the cilium suggests how cyclopamine inhibits the Hedgehog pathway, the researchers say, and shows that the correlation between Smoothened on the cilium and pathway activation is very tight.

Third, they examined whether the Smoothened protein included an amino acid sequence that other seven-transmembrane proteins require to move to the primary cilium and, if so, whether this sequence- a so-called "motif"- was essential to its relocation there. The answer to both questions was yes: A study of mouse cells in which Smoothened was mutated to lack the motif revealed that Smoothened no longer moved to the primary cilium.

Finally, to determine Smoothened's function, they tested the mutant form of Smoothened that no longer could move to the primary cilium in epithelial cells in culture and in zebrafish embryos to see if the protein still functioned. It did not.

"Thus, not only does Smoothened ciliary localization depend up on Hedgehog signaling, but Hedgehog signaling depends on a Smoothened ciliary localization motif," says Reiter.

"Whether Smoothened functions at the cilium in all cell types remains to be determined. In addition, how Smoothened activates the Hedgehog pathway at the cilium remains unclear," he says. "But the current finding lays the groundwork for future studies that could ultimately have clinical benefit."

The study was funded by the National Institutes of Health, the Sandler Foundation and the March of Dimes.

http://www.emaxhealth.com/51/5097.html

Cholesterol gets 'thumbs up' for role in digit development
April 22 (EurekAlert): When a new mother counts her newborn's fingers and toes, she probably doesn't realize that cholesterol may be to thank for baby's complete set of 20 digits.

Although cholesterol has a bad rap as the sticky, fatty substance responsible for clogging arteries, Vanderbilt University Medical Center researchers recently found that the attachment of cholesterol to an important developmental protein controls the development of fingers and toes in mice. Without cholesterol, mice developed extra digits, as well as digits in the wrong places.

The new study published online in the Proceedings of the National Academy of Sciences (PNAS) last week helps to clear up some of the conflicting data about cholesterol's controversial role in limb development, said senior author on the study, Chin Chiang, Ph.D., associate professor of Cell and Developmental Biology.

The developmental protein at work here, named Sonic hedgehog after the video game character, was discovered in the early 1990s and shown to have important roles in patterning the developing embryo, including proper digit patterning. Chiang led the early studies showing that mice without Sonic hedgehog developed only a single digit â€" a thumb on the front paw (or a "big toe"on the back paw).

The Sonic hedgehog protein is produced by a specialized group of cells located at the posterior part of the developing limb bud, which eventually develops into the pinkie finger or toe. At the site of its synthesis, Sonic hedgehog concentrations are high. It then diffuses out across the developing limb bud, and the declining concentrations (or gradient) of the protein dictate the identity of the other digits.

"Questions have remained about what regulates the Sonic hedgehog gradient,"said Chiang. "And we've been working on that for a number of years."One clue about this regulation came when other researchers discovered Sonic hedgehog's rather unusual requirement â€" the protein had to have a cholesterol molecule attached to work properly.

"In fact, Sonic hedgehog is the only protein known to be modified by cholesterol,"Chiang said.

Because cholesterol is typically found in cell membranes and thought to 'tether' proteins to cells, scientists speculated that cholesterol might inhibit the movement of Sonic hedgehog through the developing tissue. This unique modification might explain why concentrations of the protein were high at the site of its production and then tapered off with increasing distance from the synthesis site.

But previous studies in mice suggested that cholesterol promoted the movement of Sonic hedgehog, a counterintuitive proposal given cholesterol's supposed tethering ability.

To try and clear up cholesterol's role in digit patterning and the Sonic hedgehog gradient, Chiang and colleagues created mice with an altered form of the Sonic hedgehog protein to which cholesterol cannot attach.

They found that mice lacking cholesterol-modified Sonic hedgehog developed with malformed and ectopic, or out of place, digits. The second, or "index,"digits were stunted and misshapen, appearing more similar to a thumb than a normal second digit.

The researchers also examined mice in which only half of their Sonic hedgehog proteins could attach to cholesterol. Those mice developed normal digits 2 through 5 (index through pinkie), but had duplication of these digits anteriorly.

The findings suggested that Sonic hedgehog without cholesterol traveled further than normal, triggering the anomalous digit duplications.

Chiang and colleagues confirmed this microscopically, showing that the protein spread out more evenly across the limb bud (a weaker gradient) in mice lacking cholesterol-modified Sonic hedgehog compared to normal animals.

"We found that, without cholesterol, Sonic hedgehog moves more readily, far from its site of synthesis, all the way to the anterior part of the limb bud where it is normally never detected,"Chiang explained. When Sonic hedgehog travels to tissue where it normally would be absent (as it does when cholesterol is missing), extra digits may form â€" a condition known as polydactyly.

Although the causes of polydactyly in humans are not fully understood, mutations in some part of the Sonic hedgehog signaling pathway are high on the list of suspects.

In addition to limb deformities, errors in Sonic hedgehog signaling are involved in a number of other human conditions including cancer and a condition known as holoprosencephaly, a congenital malformation of the forebrain. Chiang is currently examining the role of cholesterol-modified Sonic hedgehog in the developing brain and spinal cord.

"We are finding some surprises,"Chiang said, "suggesting that the function of cholesterol is different in these different tissues." The continued study of the wide-ranging actions of Sonic hedgehog promises to expose the incredible secrets of the developing embryo and could provide clues for preventing devastating birth defects.

http://www.hindu.com/thehindu/holnus/008200604221440.htm

Public release date: 22-May-2006
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Contact: John D. Pastor
jpastor@vpha.health.ufl.edu
352-273-5815
University of Florida

How ancient whales lost their legs, got sleek and conquered the oceans

When ancient whales finally parted company with the last remnants of their legs about 35 million years ago, a relatively sudden genetic event may have crowned an eons-long shrinking process.
An international group of scientists led by Hans Thewissen, Ph.D., a professor of anatomy at Northeastern Ohio Universities College of Medicine, has used developmental data from contemporary spotted dolphins and fossils of ancient whales to try to pinpoint the genetic changes that could have caused whales, dolphins and porpoises to lose their hind limbs.

More than 50 million years ago the ancestors of whales and dolphins were four-footed land animals, not unlike large dogs. They became the sleek swimmers we recognize today during the next 15 million years, losing their hind limbs in a dramatic example of evolutionary change.

"We can see from fossils that whales clearly lived on land - they actually share a common ancestor with hippos, camels and deer," said team member Martin Cohn, Ph.D., a developmental biologist and associate professor with the UF departments of zoology and anatomy and cell biology and a member of the UF Genetics Institute. "Their transition to an aquatic lifestyle occurred long before they eliminated their hind limbs. During the transition, their limbs became smaller, but they kept the same number and arrangement of hind limb bones as their terrestrial ancestors."

In findings to be published this week in the Proceedings of the National Academy of Sciences, scientists say the gradual shrinkage of the whales' hind limbs over 15 million years was the result of slowly accumulated genetic changes that influenced the size of the limbs and that these changes happened sometime late in development, during the fetal period.

However, the actual loss of the hind limb occurred much further along in the evolutionary process, when a drastic change occurred to inactivate a gene essential for limb development. This gene - called Sonic hedgehog - functions during the first quarter of gestation in the embryonic period of the animals' development, before the fetal period.

In all limbed vertebrates, Sonic hedgehog is required for normal limbs to develop beyond the knee and elbow joints. Because ancient whales' hind limbs remained perfectly formed all the way to the toes even as they became smaller suggests that Sonic hedgehog was still functioning to pattern the limb skeleton.

The new research shows that, near the end of 15 million years, with the hind limbs of ancient whales nonfunctional and all but gone, lack of Sonic hedgehog clearly comes into play. While the animals still may have developed embryonic hind limb buds, as happens in today's spotted dolphins, they didn't have the Sonic hedgehog required to grow a complete or even partial limb, although it is active elsewhere in the embryo.

The team also showed why Sonic hedgehog became inactive and all traces of hind limbs vanished at the end of this stage of whale evolution, said Cohn. A gene called Hand2, which normally functions as a switch to turn on Sonic hedgehog, was shown to be inactive in the hind limb buds of dolphins. Without it, limb development grinds to a halt.

"By integrating data from fossils with developmental data from embryonic dolphins, we were able to trace these genetic changes to the point in time when they happened," Thewissen said.

"Studies on swimming in mammals show that a sleek body is necessary for efficient swimming, because projecting organs such as rudimentary hind limbs cause a lot of drag, and slow a swimmer down," said Thewissen, who spends about a month every year in Pakistan and India collecting fossils that document the land-to-water transition of whales.

Researchers say the findings tend to support traditional evolutionary theory, a la Charles Darwin, that says minor changes over vast expanses of time add up to big changes. And while Sonic hedgehog's role in the evolution of hind limbs in ancient whales is becoming apparent, it is still not fully defined.

"It's clear when ancient whales lost all vestiges of the limb it was probably triggered by loss of Sonic hedgehog," said Clifford Tabin, Ph.D., a professor of genetics at Harvard Medical School who was not involved in the research. "But it's hard to say for certain because you're looking at events long after they occurred. As they suggest, there could have been a continual decrease in Sonic as the limbs reduced until the modern version of the animal arrived."

The study itself, combining fossil and developmental data, is notable, Tabin said.

"Whales went through this remarkable transformation to become more like the ancestral fish," Tabin said. "Convergence of evolutionary studies and developmental genetics give us another piece in this growing tapestry of how genetic changes lead to morphological change. It is a remarkable process that was achieved simply and led to profound consequences in how whales were able to survive. Only now in the last five years are we developing this understanding of how the world of evolution is controlled genetically."


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In addition to UF and Northeastern Ohio Universities College of Medicine, scientists from the Natural History Museum of Los Angeles County and the Indian Institute of Technology were involved in the research. Financial support was provided by the National Science Foundation, the National Institutes of Health and the Indian Department of Science and Technology, New Delhi.

http://www.eurekalert.org/pub_releases/2006-05/uof-haw052206.php