20091202-2

Cell mates: This microfluidics chip (top) contains 1,536 minute wells, each designed to force contact between two fluorescently labeled cells. An immune cell and a tumor cell are forced into contact inside a single well of a microfluidics chip (bottom). The setup allows researchers to identify, isolate, and study immune cells with particularly potent anticancer characteristics.     Credit: COCHISE Project
 

A new biosensor can spot potent cancer-killing immune cells.
 

MIT Technology Review, December 1, 2009, by Lauren Gravitz  —  Scientists have long known that the human immune system has a method for detecting and destroying precancerous cells. But finding the cells behind this defense mechanism in order to study and perhaps even mimic them has proved quite the challenge. Since the malignant precancerous cells are eradicated before we even know they exist, identifying the cells that killed them seemed nearly impossible. Now European researchers have built a microfluidic biosensor that traps single immune cells together with single tumor cells, allowing the researchers to pick the most potent of these cancer killers out of a crowded field.

The project, called Cell On CHIp bioSEnsor (COCHISE), was initiated by microsystems engineer Roberto Guerrieri at the University of Bologna, Italy. Guerrieri noticed that immunologists had no way to identify and isolate those rare immune cells, or lymphocytes, with antitumor properties–only about one in every 1,000 immune cells has such properties.

Together with postdoctoral researcher Massimo Bocchi, Guerrieri created a microfluidics platform with an array of 1,536 microwells. In each well, electric fields force contact between a fluorescently labeled tumor cell and a labeled immune cell. An automated system then scans the array and detects wells in which the tumor cell’s color has disappeared, thereby identifying the lymphocytes that are likely most effective against the leukemia and lymphoma cancers they tested.

The researchers then collect the individual cells that have triumphed over the tumor cells andprovide them to immunologists for study and propagation. “Analyzing a cell we know is active is a large step for research, because you can correlate expression of cytokines or gene expression,” Bocchi says. “You can then identify genetic properties that are probably responsible for the cell being active against the tumor.” He notes that this could one day be used to find new drugs to fight the disease.

Guerrieri and his colleagues are also working to clone entire cell lines from these single, potent lymphocytes. They plan to see if the resulting daughter cells maintain the same anticancer properties. If so, such an approach could be useful for developing cancer vaccines based on a transplant of a patient’s own lymphocytes, the researchers say.

As far as the biosensor is concerned, “the design itself is not really new,” says Luke Lee, director of the Biomolecular Nanotechnology Center at the University of California at Berkeley. Others have developed similar designs, although Lee notes that none are as user-friendly as the COCHISE system. Unlike the other devices, Lee says, the biosensor devised by Guerrieri and Bocchi and their collaborators offers a way to cleanly deliver cells to the chip and manipulate them. “Most demonstrations aren’t as clean as this,” he says.

“It seems like an interesting technology,” says Yale University hematology-oncology specialist Madhav Dhodapkar. Despite the technology’s promise, however, he notes that problems can arise when tumor cells are removed from the environment that surrounds them inside the body. “We should not lose track of the complexity of cancer,” Dhodapkar says. “A tumor cell separated from its microenvironment does not have the same biology, so studying interactions by taking cells out of their microenvironment has caveats.”

Dhodapkar still believes the technology holds great promise for immunologists, cell biologists, and other researchers. “I think the biggest advantage of this technology may very well be that it will allow an opportunity to ask really detailed questions of cell-to-cell interactions that otherwise are much harder to do,” he says. “If it pans out, it could be a very useful tool. Not just for cancer but for many other platforms.”

That’s precisely what Bocchi is hoping. “When we completed the project, we observed that the tool wasn’t just for immunology but a more general platform that could run a large number of applications,” he says. Among other uses, he points to gene therapy and even the study of microalgae–one of the great biofuel hopes.

In 2006, just as COCHISE was getting off the ground, Bocchi started a company called MindSeeds to develop and commercialize the technology. Before it can go much further, the company still needs to find ways to scale up the technology–its automated platform currently examines only one cell at a time–and to standardize the technology so that every experiment can be repeated to yield the same results. “Because we’re not bound to a specific type of cell, we can potentially address several markets, and several fields,” Bocchi says.

Nanotechnology-based drug delivery offers new treatment options for deadly pancreatic cancers.
 

MIT Technology Review, December 1, 2009, by Erika Jonietz  —  Nanoparticles that deliver two or more drugs simultaneously can significantly shrink pancreatic cancer tumors and also reduce its spread, say researchers at Massachusetts General Hospital. Tayyaba Hasan, who is also a professor of dermatology at Harvard Medical School, led the development and testing of two “nanocells.” These nanocells combine light-based therapy with molecules that inhibit the growth of cancer cells or of the blood vessels that feed them.

Though the particles have only been studied in mice so far, the cancer-research community is excited. Pancreatic cancer remains one of the deadliest and hardest cancers to treat; mortality rates have changed very little in the last 30 years. After diagnosis, patients tend to live only six months, and less than 5 percent survive for five years. “In terms of a patient population, there is very little we can do for them once we find the cancer,” says Craig Thompson, director of the Abramson Cancer Center at the University of Pennsylvania.

Hasan and two research fellows in her lab, Prakash Rai and Lei Z. Zheng, presented their initial results on November 17 at the International Conference on Molecular Targets and Cancer Therapeutics, held jointly by the American Association for Cancer Research, the U.S. National Cancer Institute (NCI), and the European Organization for Research and Treatment of Cancer.

The team’s first type of nanocell is designed to effectively starve tumors by cutting off their blood supply. They trapped a photosensitive drug called verteporfin, which creates toxic oxygen radicals when exposed to specific wavelengths of light, inside solid polymer nanoparticles. Those nanoparticles were then encapsulated in lipid particles along with bevacizumab, an antibody that specifically inhibits the growth of new blood vessels by blocking a protein called VEGF. Both verteporfin and bevacizumab are already approved by the U.S. Food and Drug Administration. Bevacizumab is approved for the treatment of advanced cancers of the colon, breast, lung, and kidney; it’s marketed by Genentech as Avastin. Verteporfin is used to eliminate abnormal blood vessels in wet-form macular degeneration. It’s sold as Visudyne by Novartis.

In a previous small-scale clinical trial, verteporfin alone increased the median survival of pancreatic cancer patients from six months to nine months. Adding Avastin, however, did not increase survival time–possibly because the Avastin killed off the tumor’s blood vessels, making it difficult to get enough of the photosensitive drug to the cancer.

In contrast, when the nanocells are injected intravenously, they deliver both drugs directly to the inside of cancer cells. Blood vessels in normal tissue are impermeable to the nanoparticles, but blood vessels in tumors are “leakier,” with much larger pores that allow the nanoparticles to pass through. As a result, the nanoparticles accumulate inside tumors and deliver more of their payload to the cancer cells than to healthy cells. The nanocells provide a higher effective dose of drug to the tumors as well as fewer side effects because the researchers used a lower dose of both drugs than usual.

The team implanted human pancreatic cancer cells in mice and allowed tumors to grow. They then injected the mice with a single dose of the nanocells and exposed the tumor to long-wavelength light. Mice given this single treatment showed a greater reduction in their tumor size than mice treated with either drug alone. The mice treated with the nanocells also had at least two times fewer metastases to the liver, lungs, and lymph nodes. “Injecting these things as separate entities is not as effective as combining them into one construct,” says Hasan.

Hasan believes that’s because the nanocells actually fuse with the tumor cells and deliver the Avastin inside the cell, instead of just to the outside. And though Hasan’s lab has not done any toxicity studies, she hopes that the nanocells’ preferential accumulation inside of tumors may decrease the drug’s side effects, which can be quite dangerous. As many as 30 percent of patients receiving Avastin suffer cardiovascular side effects, including dangerously high blood pressure, stroke, and heart failure.

Shiladitya Sengupta, an assistant professor of medicine and health sciences and technology at Harvard Medical School, calls the results of Hasan’s mouse experiments “dramatic.” He says, “In the context of pancreatic cancer, [the results are] outstanding, because there’s no therapy.”

Sengupta did not participate in Hasan’s research, but he originated the idea of drug delivery using nanocells. Technology Review recognized him for this idea with a 2005 TR35 award. He cofounded Cerulean Pharma to commercialize the nanocell platform and other nanopharmaceutical delivery methods. But one tricky aspect of the technology is that it must be individually optimized for every new combination of drugs, he notes.

Hasan’s team has already developed a second nanocell designed to prevent pancreatic cancers from developing resistance to chemotherapy, a very common problem. Other researchers have identified two proteins, EGFR and MET, as particularly important in the development and growth of pancreatic cancer. In fact, in cancer cell lines in the lab, when biologists block EGFR, the cells increase their production of MET, and vice versa. So to better control the tumors, Hasan’s team set out to target EGFR and MET simultaneously, while again hitting the tumor with light to increase the effectiveness of the treatment.

This second nanocell required a more sophisticated design. Rai started with a small molecule called PHA-66572, which inhibits the MET protein, and confined it in the same sort of solid polymer nanoparticle used in the first nanocell. He then surrounded those nanoparticles with cetuximab, an antibody that blocks EGFR. Finally, he incorporated Visudyne into a lipid sphere that he used to encapsulate these two layers.

Zheng says that tumors shrank dramatically in mice that had been implanted with pancreatic cancers and then given a single injection of the nanocells followed by light therapy. He is still measuring the effects on metastasis, but since the MET protein is active in most cancers that have metastasized (not just pancreatic cancer), the researchers are optimistic that the growth-factor nanocells will significantly decrease the number and size of metastases as well.

Zheng says that these results are particularly encouraging because of the apparent reduction in toxicity of the drugs. Pfizer developed PHA-66572 specifically to block MET in cancer cells, but it proved so toxic that the company abandoned the drug. In contrast, Zheng says that the animals that he gave the nanocell maintained normal activity levels and didn’t lose weight.

Hasan hopes that both nanocells will be tested in pancreatic cancer patients within just a few years. Because Avastin and Visudyne are already FDA-approved, their two-part nanocell will likely be the first tested, probably in about two years, but perhaps as soon as a year from now, she says.

The NCI is already conducting toxicology tests of the Avastin-Visudyne nanocell as part of a new drug application to the FDA. The growth factor nanocell should enter the clinic “soon after,” Hasan says. The key is finding the best MET inhibitor, and Hasan says that other researchers are already testing several promising candidates.

20091202-1

DEADLY AGENT The drug-resistant ST313 bacterium, in red.

The strain, a variant of Salmonella typhimurium, is named ST313. 

 

The New York Times, December 2, 2009, by Donald G. McNeil Jr  —  A new drug-resistant strain of bacteria has emerged in the last decade in Africa and is causing unusual numbers of deaths there, British and African researchers said on Monday.

The strain, a variant of Salmonella typhimurium, is named ST313. Its genome was decoded by researchers from the Wellcome Trust Sanger Institute and researchers in Kenya and Malawi.

While most salmonella bacteria cause diarrhea and are rarely fatal, this one causes death in one of four cases among children and vulnerable adults in some African regions, the researchers said. Many of its victims have been weakened by the AIDS virus, anemia, malaria or malnutrition.

Salmonella normally circulates in animals and reaches humans via food poisoning. (Consumer Reports said Monday that two-thirds of the chickens it had tested had campylobacter or salmonella, though not of this new strain.)

But after sequencing the bacterial DNA found in about 50 Africans with severe infections, the researchers said the ST313 strain appeared to be mutating to circulate in humans independently of animals, as, for example, drug-resistant staph infections now do.

ST313 “has rapidly gained resistance to many of the commonly used antibiotics in the field,” said Dr. Chisomo Msefula, a researcher, and the multi-drug-resistant form seems to be becoming dominant in parts of Africa as antibiotics knock out competitors.

The paper’s authors said poor countries needed greater access to sophisticated genetic sequencing machines that could spot tiny DNA mutations like the ones making this germ so lethal.

December 2, 2009, by Gabe Mirkin MD  —  Jens Bangsbo of the University of Copenhagen has shown that if you want to run, cycle or swim faster at any distance, you have to train at a pace that is almost as fast as you can move

(Journal of Applied Physiology, November 2009).  He asked

competitive distance runners to reduce their mileage by 25 percent,

and to run 8 to 12  30-second sprints  2-3 times a week, with some

additional 0.6-0.8 mile sprints 1 or 2 times per week, for 6 to 9

weeks. The control group of runners continued their regular training

program, and showed no improvement.  The sprint group improved both

their 3K (1.8 mile)  and 10K (6 mile) race times by more than three

percent (more than a minute in the 10-K race).  Half of them ran

their best times ever, even though many had been racing for more

than five years.

 

Two years ago, Dr. Bangsbo did ground-breaking research

supporting the leading theory that exhaustion of the sodium-

potassium pump is the major cause of muscle fatigue during

exercise (Acta Physiologica, November 2007).  In this new study,

he shows how sprint training improves a muscle’s capacity to

pump potassium back inside muscle cells during exercise, which

helps all athletes run or cycle faster in competition, even in

endurance events such as marathons and multi-day bicycle races.

 

A muscle can contract only if it has an electrical charge

across the muscle cell membrane.  This electrical charge comes

mainly from having sodium primarily outside the cell and

potassium primarily inside the cell.  This higher concentration of

sodium outside the cell and higher concentration of potassium

inside the cell is maintained by sodium-potassium pumps in the

cell membranes.  The pumps get their energy from an enzyme

called ATPase.

 

When the brain sends electrical signals along nerves

leading to each muscle fiber, sodium moves rapidly into muscle

cells followed by an equivalent movement of potassium out of the

cells, causing the muscle fibers to contract.  However, the sodium-

potassium pump cannot pump potassium back into the cells as

fast as the rapidly-contracting muscle cells move potassium out.

 

Dr. Bangsbo showed that during rapid contractions,

muscle cells lose potassium so fast that there is a doubling of the

potassium outside cells in less than a minute.  The electrical

charge between the inside and outside of muscle cells is reduced,

and they contract with much less force until finally they cannot

contract at all.  During continuous contractions of muscles, the

loss of force from a muscle contraction is directly proportional

to the amount of potassium that goes outside the cells.

 

Over time, repeated muscle contractions themselves will

markedly increase the ability of the sodium-potassium pump to

pump potassium into cells.  The greater the force on a muscle

during training, the more effectively the potassium pump can

pump potassium back into muscles, resulting in greater endurance

for the athlete. So intense training is necessary for endurance,

and any training strategy that increases the number of intense

workouts will give the athlete greater endurance.

 

You can also increase the effectiveness of  the sodium

potassium pumps by being excited before a race (which increases

adrenalin), and by eating before and during races (which raises

insulin levels).  Hormones known to strengthen the sodium-

potassium pump, and therefore to increase endurance, include

adrenalin, insulin, insulin-like growth factor I, calcitonins,

amylin, thyroid, testosterone and cortisones.

 

How to apply this information to your training program:

 

You cannot gain maximum endurance just with continuous

exercise. To improve your potassium-sodium pumps, you have to

put maximum force on your muscles. This requires some form of

interval training.  (CAUTION: Intense exercise can kill a person

with blocked arteries to the heart; check with your doctor before

increasing the intensity of your program.

 

 Intervals are classified as short intervals that take fewer

than 30 seconds and do not generate significant amounts of lactic

acid; and long intervals that take more than two minutes and

generate large amounts of lactic acid.  The longest you can

exercise with maximal force on muscles is about 30 seconds. All

competitive athletes should do some sort of 30-second interval.

Nobody knows how often you have to do this, but most runners

and cyclists do short intervals once or twice a seek.  You probably

should do long intervals also.  However, applying near-maximal

force on muscles for more than 30 seconds causes considerable

muscle damage, so you have to allow muscles to recover by

doing slow training for one or two days afterwards.

 

Since short intervals do not accumulate much lactic acid,

you can do a large number of repetitions during a single workout.

Long intervals cause a tremendous amount of muscle damage, so

you can only do a few long intervals during a workout.  A sound

endurance program should include a lot of slow miles, one or two

workouts with many short intervals, and probably at least one

workout that includes a few long intervals each week.

The New York Times, December 2, 2009, by Anahad O’Connor  —  It has long been said that regular physical activity and better sleep go hand in hand. Burn more energy during the day, the thinking goes, and you will be more tired at night.

But only recently have scientists sought to find out precisely to what extent. One extensive study published this year looked for answers by having healthy children wear actigraphs – devices that measure movement – and then seeing whether more movement and activity during the day meant improved sleep at night. The results should be particularly enlightening to parents.

The study found that sleep onset latency – the time it takes to fall asleep once in bed – ranged from as little as roughly 10 minutes for some children to more than 40 minutes for others. But physical activity during the day and sleep onset at night were closely linked: every hour of sedentary activity during the day resulted in an additional three minutes in the time it took to fall asleep at night. And the children who fell asleep faster ultimately slept longer, getting an extra hour of sleep for every 10-minute reduction in the time it took them to drift off.

Studies on adults have reached generally similar results, showing that an increase in physical activity improves sleep onset and increases sleep duration, particularly in people who have trouble sleeping.

THE BOTTOM LINE Studies suggest that being more physically active can lead to better sleep.