20090904-6

Santa Barbara, Calif. — Scientists at UC Santa Barbara have made a significant discovery in understanding the way human embryonic stem cells function.

They explain nature’s way of controlling whether these cells will renew, or will transform to become part of an ear, a liver, or any other part of the human body. The study is reported in the May 1 issue of the journal Cell.

The scientists say the finding bodes well for cancer research, since tumor stem cells are the engines responsible for the growth of tumors. The discovery is also expected to help with other diseases and injuries. The study describes nature’s negative feedback loop in cell biology.

“We have found an element in the cell that controls ‘pluripotency,’ that is the ability of the human embryonic stem cell to differentiate or become almost any cell in the body,” said senior author Kenneth S. Kosik, professor in the Department of Molecular, Cellular & Developmental Biology. Kosik is also co-director and Harriman Chair in Neuroscience Research of UCSB’s Neuroscience Research Institute.

“The beauty and elegance of stem cells is that they have these dual properties,” said Kosik. “On the one hand, they can proliferate — they can divide and renew. On the other hand, they can also transform themselves into any tissue in the body, any type of cell in the body.”

The research team includes James Thomson, who provided an important proof to the research effort. Thomson, an adjunct professor at UCSB, is considered the “father of stem cell biology.” Thomson pioneered work in the isolation and culture of non-human primate and human embryonic stem cells. These cells provide researchers with unprecedented access to the cellular components of the human body, with applications in basic research, drug discovery, and transplantation medicine.

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With regard to human embryonic stem cells, Kosik explained that for some time he and his team have been studying a set of control genes called microRNAs. “To really understand microRNAs, the first step is to remember the central dogma of biology –DNA is the template for RNA and RNA is translated to protein. But microRNAs stop at the RNA step and never go on to make a protein.

“The heart of the matter is that before this paper, we knew that if you want to maintain a pluripotent state and allow self-renewal of embryonic stem cells, you have to sustain levels of transcription factors,” said Kosik. “We also knew that stem cells transition to a differentiated state when you decrease those factors. Now we know how that happens a little better.”

The new research shows that a microRNA — a single-stranded RNA whose function is to decrease gene expression — lowers the activity of three key ingredients in the recipe for embryonic stem cells. This microRNA is known as miR-145. The discovery may have implications for improving the efficiency of methods designed to reprogram differentiated cells into embryonic stem cell-like cells.

As few as three or four genes can make cells pluripotent. “We know what these genes are,” Kosik said. That information was used recently for one of the most astounding breakthroughs of biology of the last couple of years — the discovery of induced pluripotent skin cells.

“You can take a cell, a skin cell, or possibly any cell of the body, and revert it back to a stem cell,” Kosik said. “The way it’s done, is that you take the transcription factors that are required for the pluripotent state, and you get them to express themselves in the skin cells; that’s how you can restore the embryonic stem cell state. You clone a gene, you put it into what’s called a vector, which means you put it into a little bit of housing that allows those genes to get into a cell, then you shoot them into a stem cell. Next, when those genes — those very critical pluripotent cell genes — get turned on, the skin cell starts to change, it goes back to the embryonic pluripotent stem cell state.”

The researchers explained that a rise in miR-145 prevents human embryonic stem cells’ self-renewal and lowers the activity of genes that lend stem cells the capacity to produce other cell types. It also sends the cells on a path toward differentiation. In contrast, when miR-145 is lost, the embryonic stem cells are prevented from differentiating as the concentrations of transcription factors rise.

They also show that the control between miR-145 and the “reprogramming factors” goes both ways. The promoter for miR-145 is bound and repressed by a transcription factor known as OCT4, they found.

“It’s a beautiful double negative feedback loop,” Kosik said. “They control each other. That is the essence of this work.”

Because there is typically less “wiggle room” in the levels of microRNA compared to mRNA, further studies are needed to quantify more precisely the copy numbers of miR-145 and its targets, to figure out exactly how this layer of control really works, Kosik said.

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Human embryonic stem cells are poised between a proliferative state with the potential to become any cell in the body and a differentiated state with a more limited ability to proliferate. To maintain this delicate balance embryonic stem cells express a set of factors, including OCT4, SOX2, and KLF4, to control multiple genes that sustain the proliferative pluripotent state. These genes can be repressed by a tiny RNA called miR-145, and in turn, one of the transcription factors, OCT4, can repress miR-145. Thus, a double negative feedback loop sets the delicate balance. 

Kosik credits the lion’s share of this discovery to first author Na Xu, a postdoctoral fellow who is also supported by the California Institute for Regenerative Medicine (CIRM). “Na Xu deserves enormous credit for this work,” said Kosik. “She performed nearly every experiment in the paper and was the major contributor to the ideas in the paper.” Meanwhile, Thales Papagiannakopoulos, a graduate student working in the Kosik lab, was very generous in helping Na Xu with one of the experiments. He helped with one of several proofs that showed that the targets of miR-145 are the three transcription factors that are being reported, explained Kosik.

Thomson provided one of several proofs for the control point of miR-145 expression, said Kosik.

20090904-5

CortlandtForum.com, September 3, 2009  —  Pomegranate juice may help slow progression of prostate cancer in men who experience rising PSA levels following radical prostatectomy or radiotherapy, data suggest.

Researchers at the University of California at Los Angeles (UCLA) presented long-term data from a phase 2 trial involving 48 men who had rising PSA levels after prostate cancer treatment. To be eligible for the study, subjects had to have a PSA level greater than 0.2 ng/mL but less than 5. The men had a pretreatment Gleason score of 7 or less. Participants drank eight ounces of pomegranate juice daily (570 mg total polyphenol gallic acid equivalents).

Interim results previously published in Clinical Cancer Research (2006;12:4018-4026) showed a significant increase in the mean PSA doubling times after treatment with pomegranate juice: from 15 months at baseline to 54 months post-treatment. Following these positive results, the study was amended to allow subjects to continue treatment and undergo evaluation at three-month intervals until disease progression. At the end of six years, the mean PSA doubling time was 60 months post-treatment, according to investigators.

In the sixth year of treatment, 15 patients (31%) remained in the study, with a median follow-up of 30 months post-treatment (maximum 64 months). These patients had a significantly greater PSA doubling time and larger decline in median PSA slope than subjects no longer in the study.

“We are now in the seventh year of this study. This is quite unique in clinical research-to have such a long length of follow-up in a phase 2 study,” said lead investigator Allan Pantuck, MD, Associate Professor of Urology at the David Geffen School of Medicine at UCLA. “We have preclinical and clinical data that continue to suggest a slowing down of PSA doubling times in patients with prostate cancer.” The effect of pomegranate juice on PSA doubling times appears to be durable, he said.

Dr. Pantuck noted that the trial so far suggests that daily consumption of pomegranate juice for more than five years appears to be safe and to produce no untoward adverse effects. In addition, data show that some patients may be more sensitive than others to the effects of pomegranate juice.

He pointed out that pomegranate juice is being studied for many medical indications- “everything from cancer to heart disease. However, it is still too early to say it is an elixir of life or that we have proven that there is a benefit. We currently have positive results that have justified the time and expense and effort to study the juice in a phase 3 study, and we hope to have these definitive results shortly.”

The investigators reported study findings at the recent American Urological Association Annual Meeting in Chicago.

“This study suggests that pomegranate juice may effectively slow the progression of prostate cancer after unsuccessful treatment,” said American Urological Association spokesperson Christopher Amling, MD, Division Chief of Surgery at Oregon Health & Science University in Portland. “This finding and other ongoing research might one day reveal that pomegranate juice is an effective prostate cancer preventative agent as well.”

20090904-2

That’s why new flu vaccines are produced every year, carefully tailored to fight the specific strains predicted to be the most widespread around the world in the coming flu season. It’s also why vaccines don’t work against pathogens that are constantly mutating, resulting in a broad spectrum of strain variants against which conventional vaccines are ineffective.

“That’s the Achilles’ heel of vaccines,” says Michael Mahan, a bacterial geneticist in UCSB’s Department of Molecular, Cellular, and Developmental Biology. Together with Douglas Heithoff – Mahan’s former student, now a research scientist in the department – and colleagues at the University of Utah, he’s working to develop a new generation of vaccines, termed cross-protective, that protect against many strains of a given pathogen.

In the past, vaccines have been developed using a somewhat empirical approach. “They work, but you usually don’t know why,” Heithoff says. “We’re trying to use a mechanistic approach toward vaccine design.”

To do that, they first took a careful look at how microbes operate during infection. “We’re working to understand the mechanisms of disease at the molecular level,” Mahan says, “in order to make better medicines.”

They’ve been studying Salmonella, a bacterium that is found in the digestive tracts of mammals, reptiles, birds, and insects, and is spread in feces. Salmonella infects up to 1.5 billion people annually worldwide, with more than a million cases in the U.S. alone. There are around 2,500 known strains of the bacterium, and any given strain can cause sickness in some hosts but not in others. In humans, different strains of Salmonella can cause food poisoning, blood poisoning (sepsis), and typhoid fever.

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For most people, gastroenteritis caused by Salmonella is unpleasant, but short-lived. In the elderly, the very young, and people whose immune systems are compromised by HIV or cancer treatments, however, an infection can be fatal. In these severe cases, bacteria may spread from the intestines to the blood and then to other organs.

Humans usually pick up Salmonella from contaminated beef or chicken. Vegetarians aren’t immune though, since Salmonella carried in animal waste can contaminate fields where vegetables are grown and facilities where food is processed or prepared. A Salmonella outbreak that hit headlines early this year was traced to contaminated peanut butter and other products containing peanuts. It infected hundreds of people in dozens of states and has been linked to at least nine deaths.

One strain of Salmonella causes typhoid fever, a sometimes-fatal illness that is very rare in the U.S., but affects more than 20 million people in the developing world each year, according to the Centers for Disease Control. That strain is only known to infect humans, and spreads as a result of poor water sanitation and hygiene practices.

There is a vaccine against typhoid, developed decades ago, but it’s not particularly effective, Mahan says. It offers protection only against a few related strains of typhoid-causing Salmonella, so it’s no help in fighting the other strains that can sicken or kill.

In their earlier work on Salmonella, Mahan and Heithoff studied how the bacterium is able to lurk benignly in some hosts, and then rapidly wreak havoc once it finds its way into others. They discovered an enzyme that acts as a master switch. This switch controls “many, many virulence functions,” Mahan says, allowing the bacterium to quickly transform from harmless hitchhiker to deadly invader.

While this switch is a great asset for the microbe, it’s also its greatest vulnerability -and a terrific target for a new vaccine. Mahan and Heithoff created one by inactivating the switch, thereby disarming the bacterium. They used this crippled Salmonella – which can’t cause illness, but still provokes an immune response – as a vaccine.

They’ve tested their new Salmonella vaccine in mice, chickens, and cows, and found that it gave the animals immunity to more than 20 strains of Salmonella isolated from various infected animals from around the world.

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Mahan and Heithoff’s vaccine has another advantage over those that are currently in use: It doesn’t cause an increase in a specific type of inhibitory immune cells-cells that are associated with immune declines in cancer patients. The researchers have also shown that these inhibitory cells become more abundant with the normal aging process, which Mahan says, “may explain why the aged are more susceptible to disease and why they are difficult to effectively vaccinate,” He continued, “We’re currently working on interventions that negate these inhibitory cells-those interventions have the potential to reduce disease susceptibility and increase vaccination efficiency in the elderly.”

Although the researchers have focused on developing a new vaccine for Salmonella, the same kind of master switch is found in other dangerous bacteria, including those that cause cholera, dysentery and the plague. Mahan believes that one day cross-protective vaccines will be developed against those and other infectious microbes.

Viruses, like those that cause the flu, are a trickier target, Mahan says, “because they rapidly mutate and have very little genetic material.” He’s optimistic, however, that a new generation of human vaccines can be developed against both bacteria and viruses, offering a broader, more effective defense against multiple strains of microbes, rather than against just one or two variants.

His biggest reservation is that currently unknown pathogens could turn up in humans, surprising us and leaving us ill-prepared for their attack. “Cross-protective vaccines may work against a wide range of strains of germs, but what about the ‘bugs’ we don’t know about?” he asks.

In the meantime, Mahan, Heithoff, and their colleagues are continuing to test their Salmonella vaccine in animals, with good results. Used in humans, a vaccine effective against many strains of the bacterium could save thousands of lives, and could spare millions of other people from very unpleasant illnesses.

Mahan and Heithoff predict that cross-protective Salmonella vaccines will one day be approved for human use, but “The first of them is at least 10 years away…” Heithoff says.

Human benefits from their research, however, are closer than that: when their new vaccine is cleared for use in livestock-which “isn’t too far in the future,” according to Mahan- it will reduce human food-borne illnesses by reducing levels of Salmonella in farm animals. That, in turn, will ensure that less of the pathogen finds its way into kitchens and restaurants, and ultimately into people’s digestive tracts.

“We have to make the food supply safer,” Mahan says. “It’s unacceptable to me that a child could eat a cheeseburger and die, or spend the rest of his or her life on dialysis. We can do better, and we’re making good progress…”

Links:

Michael Mahan’s homepage:
lifesci.ucsb.edu/mcdb/faculty/mahan

UCSB’s Department of Molecular, Cellular and Developmental Biology:
lifesci.ucsb.edu/mcdb

NIH’s Medline Plus page on Salmonella:
nlm.nih.gov/medlineplus/salmonellainfections.html

CDC Salmonella home page:
cdc.gov/salmonella

By Cole Petrochko, Staff Writer, MedPage Today
Published: September 03, 2009 

WASHINGTON — A second company has been given an FDA go-ahead to market an as-yet-unapproved drug to treat Gaucher’s disease, helping to remedy a shortage of the only agent approved for the condition.

Drug manufacturer Shire, based in the U.K., has submitted a new drug application for its enzyme replacement drug velaglucerase at the request of the FDA.

The request is an attempt to provide treatment for patients after a viral contamination in June shut down the Genzyme plant that manufactures the mammalian cell-derived glucocerebrosidase analog imiglucerase (Cerezyme).

In August, the FDA cleared the plant cell-derived recombinant form of glucocerebrosidase from manufacturer Protalix for use while phase III trials are ongoing. (See FDA Allows Pre-Approval Use of Gaucher Treatment)

Velaglucerase will be available free under the FDA preapproval protocol, just as the Protalix drug will be.

However, because velaglucerase has already filed for approval, a move Protalix may be months away from, patients on velaglucerase may have to pay for it sooner than expected. Genzyme’s imiglucerase costs up to $250,000 a year.

Gaucher’s disease, a lysosomal storage disorder that causes hematologic abnormalities such as anemia and enlargement of the liver and spleen, affects about 2,500 people in the U.S.

The final phase III trials of velaglucerase met all of Shire’s primary and secondary endpoints, according to a company release.

The studies enrolled both treatment-naive and previously treated patients with type I Gaucher’s disease, comparing hemoglobin concentration and platelet count between patients taking velaglucerase and those taking imiglucerase.

Approximately 1% of patients developed antibodies to velaglucerase during the trials.

Velaglucerase is produced with the exact human amino acid sequence and a human glycosylation pattern. The drug supplements or replaces beta-glucocerebrosidase, an enzyme that catalyzes the hydrolysis of glucocerebroside and corrects the pathophysiology of Gaucher’s disease.

Although Genzyme has restarted manufacturing of imiglucerase, the product is not expected to ship until November at the earliest.

By John Gever, Senior Editor, MedPage Today
Published: August 17, 2009

An investigational therapy for Gaucher’s disease may be given to patients under an FDA-approved treatment protocol while the currently approved treatment remains in short supply.

Protalix BioTherapeutics said the FDA had given the go-ahead for physicians to prescribe its plant cell-derived recombinant form of glucocerebrosidase, the enzyme that is deficient in patients with Gaucher’s disease, under a special protocol while the firm’s ongoing phase III trial is underway.

The company will provide the drug free to patients enrolled in the protocol.

The only Gaucher’s disease treatment now approved is made by Genzyme, which had to halt production in June because of viral contamination in its Boston manufacturing plant.

Genzyme announced last week that it had restarted production of its mammalian cell-derived form of glucocerebrosidase, trade-named Cerezyme, but shipments of the finished product could not begin until November at the earliest.

A shortage situation is likely to persist through the end of the year, the company said.

It has now implemented a dose conservation program “to try to ensure that the most vulnerable patients continue to receive Cerezyme,” the firm said.

The product is now available only to two patient populations: patients with Gaucher disease type 1 who are 18 or younger, and patients with Gaucher disease types 2 and 3.

Genzyme has also created an emergency access program, through which physicians may apply to receive Cerezyme for patients who are in life-threatening situations.

A Protalix spokesperson said it had enough of its glucocerebrosidase product on hand to supply “a couple hundred” patients unable to receive Cerezyme during the shortage period.

However, patients who receive the drug under the newly approved protocol can stay on it until full FDA approval is awarded, the firm said.

Protalix, based in Israel, expects to complete the phase III trial in September. It said summary results would be made public in October and a U.S. marketing application filed by the end of this year.

About 2,500 people in the U.S. are estimated to have Gaucher’s disease, a lysosomal storage disorder that causes hematologic abnormalities such as anemia as well as enlargement of the liver and spleen.

20090904-1

A female apple maggot fly, Rhagoletis pomonella, implants an egg into an apple. Wasps that attack the flies and eat their larvae appear to be changing on a genetic level in the same way that the flies themselves appear to be changing genetically. (Credit: Rob Oakleaf) 

U of Notre Dame, ScienceDaily.com – A team of researchers are reporting the ongoing emergence of a new species of fruit fly–and the sequential development of a new species of wasp–in the February 6 issue of the journal Science.

Jeff Feder, a University of Notre Dame biologist, and his colleagues say the introduction of apples to America almost 400 years ago ultimately may have changed the behavior of a fruit fly, leading to its modification and the subsequent modification of a parasitic wasp that feeds on it.

The result is a chain reaction of biodiversity where the modification of one species triggers the sequential modification of a second, dependent species.

“It’s a nice demonstration of how the initial speciation of one organism opens up an opportunity for another species in the ecosystem to speciate in kind,” said Feder. “Biodiversity in essence is the source for new biodiversity.”

For almost 250 years after the introduction of apples to North America, insects referred to as hawthorn flies, Rhagoletis pomonella, continued to meet on the small, red fruit of hawthorn trees to mate and lay eggs. Then, in the mid-1800s, some of these “hawthorn flies” began to mate and lay eggs on apples instead. According to Feder, the flies attracted to apples eventually became genetically differentiated from the flies attracted to hawthorns, and so did the wasps that live on the flies’ larvae.

The genetic distinctions mainly show up as gene frequency differences between the flies and their associated wasp populations rather than fixed, all or none, differences. This is consistent with the process by which new biological species arise.

“The Diachasma alloeum wasp that we studied is just one of several wasps that spend a significant portion of their lives attached to hawthorn and apple flies,” said Feder. “We have preliminary evidence that one of the other wasps also may be forming specialized races on the flies, but it is too early to tell definitively.”

“What is startling is how fast populations can ecologically adapt to new habitats and begin to evolve into different species in front of our eyes,” he said.

Feder says the research is important because it provides insights into solving Darwin’s mystery of the origins of new species. “Clues can be found right before us as we sit on our deck chairs barbecuing and drinking pop. All we have to do is open our eyes and we can see new life forms coming into being in that scraggly old apple tree in our backyard.”

Notre Dame biologists Andrew Forbes and Tom Powell, along with University of Florida entomologist Lukasz Stelinski and Michigan State University biologist James Smith also worked on this project.

The National Science Foundation supports the research.