Drugs Encased in Nanoparticles Travel to Tumors on the Surface of Immune-System Cells

Drugs Encased in Nanoparticles Travel to Tumors on the Surface of Immune-System Cells

  10:53:00 PM    Science Lover

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Clinical trials using patients' own immune cells to target tumors have yielded promising results. However, this approach usually works only if the patients also receive large doses of drugs designed to help immune cells multiply rapidly, and those drugs have life-threatening side effects.

pouches (yellow) to the surfaces of cells. 
(Credit: Darrell Irvine and Matthias Stephan)

Now a team of MIT engineers has devised a way to deliver the necessary drugs by smuggling them on the backs of the cells sent in to fight the tumor. That way, the drugs reach only their intended targets, greatly reducing the risk to the patient.

The new approach could dramatically improve the success rate of immune-cell therapies, which hold promise for treating many types of cancer, says Darrell Irvine, senior author of a paper describing the technique in the Aug. 15 issue of Nature Medicine.

"What we're looking for is the extra nudge that could take immune-cell therapy from working in a subset of people to working in nearly all patients, and to take us closer to cures of disease rather than slowing progression," says Irvine, associate professor of biological engineering and materials science and engineering and a member of MIT's David H. Koch Institute for Integrative Cancer Research.

The new method could also be used to deliver other types of cancer drugs or to promote blood-cell maturation in bone-marrow transplant recipients, according to the researchers.

To perform immune-cell therapy, doctors remove a type of immune cells called T cells from the patient, engineer them to target the tumor, and inject them back into the patient. Those T cells then hunt down and destroy tumor cells. Clinical trials are under way for ovarian and prostate cancers, as well as melanoma.

Although immune-cell therapy is a promising approach to treating cancer, success has been limited by difficulties in generating enough T cells that are specific to the cancer cells and getting those T cells to function properly in the patient.

To overcome those obstacles, researchers have tried injecting patients with adjuvant drugs that stimulate T-cell growth and proliferation. One class of drugs that has been tested in clinical trials is interleukins -- naturally occurring chemicals that help promote T-cell growth but have severe side effects, including heart and lung failure, when given in large doses.

Irvine and his colleagues took a new approach: To avoid toxic side effects, they designed drug-carrying pouches made of fatty membranes that can be attached to sulfur-containing molecules normally found on the T-cell surface.

In the Nature Medicine study, the researchers injected T cells, each carrying about 100 pouches loaded with the interleukins IL-15 and IL-21, into mice with lung and bone marrow tumors. Once the cells reached the tumors, the pouches gradually degraded and released the drug over a weeklong period. The drug molecules attached themselves to receptors on the surface of the same cells that carried them, stimulating them to grow and divide.

Within 16 days, all of the tumors in the mice treated with T cells carrying the drugs disappeared. Those mice survived until the end of the 100-day experiment, while mice that received no treatment died within 25 days, and mice that received either T cells alone or T cells with injections of interleukins died within 75 days.

Irvine's approach to delivering the adjuvant drugs is both simple and innovative, says Dranoff. "The idea of modifying T cells in the lab to make them work better is something many people are exploring through more complicated approaches such as gene modification," he says. "But here, the possibility of just attaching a carefully engineered nanoparticle to the surface of cells could be a much simpler procedure."

While he is now focusing on immune-cell therapy, Irvine believes his cell pouches could be useful for other applications, including targeted delivery of chemotherapy agents. "There are lots of people studying nanoparticles for drug delivery, especially in cancer therapy, but the vast majority of nanoparticles injected intravenously go into the liver or the spleen. Less than 5 percent reach the tumor," says Irvine, who is also a Howard Hughes Medical Institute Investigator.

With a new way to carry drugs specifically to tumors, scientists may be able to resurrect promising drugs that failed in clinical trials because they were cleared from the bloodstream before they could reach their intended targets, or had to be given in doses so high they had toxic side effects.

Irvine and his colleagues also demonstrated that they could attach their pouches to the surface of immature blood cells found in the bone marrow, which are commonly used to treat leukemia. Patients who receive bone-marrow transplants must have their own bone marrow destroyed with radiation or chemotherapy before the transplant, which leaves them vulnerable to infection for about six months while the new bone marrow produces blood cells.

Delivering drugs that accelerate blood-cell production along with the bone-marrow transplant could shorten the period of immunosuppression, making the process safer for patients, says Irvine. In the Nature Medicine paper, his team reports successfully enhancing blood-cell maturation in mice by delivering one such drug along with the cells.

Irvine is now starting to work on making sure the manufacturing process will yield nanoparticles safe to test in humans. Once that is done, he hopes the particles could be used in clinical trials in cancer patients, possibly within the next two or three years.

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Fly Cells Flock Together, Follow the Light

Fly Cells Flock Together, Follow the Light

  12:52:00 AM    Science Lover

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Scientists at Johns Hopkins report using a laser beam to activate a protein that makes a cluster of fruit fly cells act like a school of fish turning in social unison, following the lead of the one stimulated with light.

Still image from a video clip showing a photo-activatable 
form of Rac. (Credit: Image courtesy of Denise Montell Lab)

The study of this unexpected cell movement, reported May 16 in Nature Cell Biology,holds potential importance for understanding embryonic development, wound healing and tumor metastasis -- the process by which tumor cells acquire the ability to invade surrounding tissues and migrate long distances to colonize lymph nodes, bones and other distant organs.

The research dramatically demonstrates, the researchers say, the collective direction-sensing behavior of live cells in intact tissue, and a means of controlling protein behavior in a living organism by shining a focused beam of light precisely on the parts of cells where they want the protein to be active.

"Our little system in the fruit fly is an elegant example of cells behaving socially in their natural environment -- surrounded by other cells," says Denise Montell, Ph.D., a professor of biological chemistry and director of the Center for Cell Dynamics at the Johns Hopkins University School of Medicine. "You can't capture this behavior if you look at individual cells in a culture dish."

The "social" migrating behavior among a cluster of cells in the fly ovary surprised the research team, which was using a new laser light tool to manipulate protein activity.

"People tend to think of cancer as single cells breaking off from the tumor and migrating away," Montell says, but it's likely that this collective form of movement is important, at one phase or another, in the spread of tumors."

A better understanding of how and why cells move can facilitate the development of new treatments not only for cancer but other disorders characterized by aberrant cell behavior.

Developed in the laboratory of Klaus Hahn, Ph.D., Thurman Professor of Pharmacology at the University of North Carolina at Chapel Hill, the light-activation technique previously had been shown to control cell movement in cultured mammalian cells. The Hopkins-led study provides proof of principle that a non-toxic light alone can activate a protein in live organisms, allowing researchers to safely control when and where cells move.

The Hopkins team conducted their study on a cluster of six so-called border cells in the fly ovary, cells the team has long studied and which are important to the fly because if they don't migrate, females are sterile. In addition they serve as a model for understanding the mechanisms that control collective cell movements in general, which occur during normal embryonic development, wound healing and in tumor metastasis.

First, they genetically altered the border cells so that they were lacking the ability to respond to naturally occurring chemical attractants that normally control their movement. Then they used a fly protein known as Rac, which was fused to a photoactivatable (PA) plant protein, a creation engineered by Hahn's lab. The PA-Rac, which remains inert in the dark, reacts to light because the plant protein changes shape and allows Rac to become active, causing the cells to move.

Because a beam of laser light can be much smaller than a cell, the team was able to activate Rac not only in one single cell, but also in one part of one cell, Montell says: "The other cool thing is this is reversible, so as soon as you take the light away, the PA-Rac wraps back up and turns itself off."

Following up on previous research, the team wanted to find out if Rac would be sufficient to set the direction of movement of cells within live tissue.

When they shined a laser beam on various individual cells, the entire cluster responded by moving in directions that it wouldn't under normal conditions: sideways, for instance, and even in reverse. In short, they followed the light.

"When we activated Rac in even one part of one of these cells -- and not in the cell that would be the leader if all was normal -- it was as if all the other cells said, Aha! You've got more Rac activity so we're heading your way," Montell says. "It's amazing to me that somehow the cells sense each others' levels of Rac activity and collectively decide which way to go.

Authors on the paper, in addition to Montell and Hahn, are Xiaobo Wang from Johns Hopkins and Yi Wu from UNC.

Funding was provided by the National Institutes of Health and the Cell Migration Consortium.

A video clip of a photo-activatable form of Rac is available at: http://www.icm.com/montell/MovieS5%28RacQ61Lforwardandrev%29.mov

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Nanoparticles: Golden Bullet for Cancer?

Nanoparticles: Golden Bullet for Cancer?

  10:03:00 AM    Science Lover

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In a lecture he delivered in 1906, the German physician Paul Ehrlich coined the term Zuberkugel, or "magic bullet," as shorthand for a highly targeted medical treatment.

Infrared images made while tumors were irradiated with a laser show that in nanocage-injected mice (left), the surface of the tumor quickly became hot enough to kill cells. In buffer-injected mice (right), the temperature barely budged. This specificity is what makes photothermal therapy so attractive as a cancer therapy. (Credit: WUSTL)

Magic bullets, also called silver bullets, because of the folkloric belief that only silver bullets can kill supernatural creatures, remain the goal of drug development efforts today.

A team of scientists at Washington University in St. Louis is currently working on a magic bullet for cancer, a disease whose treatments are notoriously indiscriminate and nonspecific. But their bullets are gold rather than silver. Literally.

The gold bullets are gold nanocages that, when injected, selectively accumulate in tumors. When the tumors are later bathed in laser light, the surrounding tissue is barely warmed, but the nanocages convert light to heat, killing the malignant cells.

In an article just published in the journal Small, the team describes the successful photothermal treatment of tumors in mice.

The team includes Younan Xia, Ph.D., the James M. McKelvey Professor of Biomedical Engineering in the School of Engineering and Applied Science, Michael J. Welch, Ph.D., professor of radiology and developmental biology in the School of Medicine, Jingyi Chen, Ph.D., research assistant professor of biomedical engineering and Charles Glaus, Ph.D., a postdoctoral research associate in the Department of Radiology.

"We saw significant changes in tumor metabolism and histology," says Welch, "which is remarkable given that the work was exploratory, the laser 'dose' had not been maximized, and the tumors were 'passively' rather than 'actively' targeted."

Why the nanocages get hot

The nanocages themselves are harmless. "Gold salts and gold colloids have been used to treat arthritis for more than 100 years," says Welch. "People know what gold does in the body and it's inert, so we hope this is going to be a nontoxic approach."

"The key to photothermal therapy," says Xia, "is the cages' ability to efficiently absorb light and convert it to heat. "

Suspensions of the gold nanocages, which are roughly the same size as a virus particle, are not always yellow, as one would expect, but instead can be any color in the rainbow.

They are colored by something called a surface plasmon resonance. Some of the electrons in the gold are not anchored to individual atoms but instead form a free-floating electron gas, Xia explains. Light falling on these electrons can drive them to oscillate as one. This collective oscillation, the surface plasmon, picks a particular wavelength, or color, out of the incident light, and this determines the color we see.

Medieval artisans made ruby-red stained glass by mixing gold chloride into molten glass, a process that left tiny gold particles suspended in the glass, says Xia.

The resonance -- and the color -- can be tuned over a wide range of wavelengths by altering the thickness of the cages' walls. For biomedical applications, Xia's lab tunes the cages to 800 nanometers, a wavelength that falls in a window of tissue transparency that lies between 750 and 900 nanometers, in the near-infrared part of the spectrum.

Light in this sweet spot can penetrate as deep as several inches in the body (either from the skin or the interior of the gastrointestinal tract or other organ systems).

The conversion of light to heat arises from the same physical effect as the color. The resonance has two parts. At the resonant frequency, light is typically both scattered off the cages and absorbed by them.

By controlling the cages' size, Xia's lab tailors them to achieve maximum absorption.

Passive targeting

"If we put bare nanoparticles into your body," says Xia, "proteins would deposit on the particles, and they would be captured by the immune system and dragged out of the bloodstream into the liver or spleen."

To prevent this, the lab coated the nanocages with a layer of PEG, a nontoxic chemical most people have encountered in the form of the laxatives GoLyTELY or MiraLAX. PEG resists the adsorption of proteins, in effect disguising the nanoparticles so that the immune system cannot recognize them.

Instead of being swept from the bloodstream, the disguised particles circulate long enough to accumulate in tumors.

A growing tumor must develop its own blood supply to prevent its core from being starved of oxygen and nutrients. But tumor vessels are as aberrant as tumor cells. They have irregular diameters and abnormal branching patterns, but most importantly, they have thin, leaky walls.

The cells that line a tumor's blood vessel, normally packed so tightly they form a waterproof barrier, are disorganized and irregularly shaped, and there are gaps between them.

The nanocages infiltrate through those gaps efficiently enough that they turn the surface of the normally pinkish tumor black.

A trial run

In Welch's lab, mice bearing tumors on both flanks were randomly divided into two groups. The mice in one group were injected with the PEG-coated nanocages and those in the other with buffer solution. Several days later the right tumor of each animal was exposed to a diode laser for 10 minutes.

The team employed several different noninvasive imaging techniques to follow the effects of the therapy. (Welch is head of the oncologic imaging research program at the Siteman Cancer Center of Washington University School of Medicine and Barnes-Jewish Hospital and has worked on imaging agents and techniques for many years.)

During irradiation, thermal images of the mice were made with an infrared camera. As is true of cells in other animals that automatically regulate their body temperature, mouse cells function optimally only if the mouse's body temperature remains between 36.5 and 37.5 degrees Celsius (98 to 101 degrees Fahrenheit).

At temperatures above 42 degrees Celsius (107 degrees Fahrenheit) the cells begin to die as the proteins whose proper functioning maintains them begin to unfold.

In the nanocage-injected mice, the skin surface temperature increased rapidly from 32 degrees Celsius to 54 degrees C (129 degrees F).

In the buffer-injected mice, however, the surface temperature remained below 37 degrees Celsius (98.6 degrees Fahrenheit).

To see what effect this heating had on the tumors, the mice were injected with a radioactive tracer incorporated in a molecule similar to glucose, the main energy source in the body. Positron emission and computerized tomography (PET and CT) scans were used to record the concentration of the glucose lookalike in body tissues; the higher the glucose uptake, the greater the metabolic activity.

The tumors of nanocage-injected mice were significantly fainter on the PET scans than those of buffer-injected mice, indicating that many tumor cells were no longer functioning.

The tumors in the nanocage-treated mice were later found to have marked histological signs of cellular damage.

Active targeting

The scientists have just received a five-year, $2,129,873 grant from the National Cancer Institute to continue their work with photothermal therapy.

Despite their results, Xia is dissatisfied with passive targeting. Although the tumors took up enough gold nanocages to give them a black cast, only 6 percent of the injected particles accumulated at the tumor site.

Xia would like that number to be closer to 40 percent so that fewer particles would have to be injected. He plans to attach tailor-made ligands to the nanocages that recognize and lock onto receptors on the surface of the tumor cells.

In addition to designing nanocages that actively target the tumor cells, the team is considering loading the hollow particles with a cancer-fighting drug, so that the tumor would be attacked on two fronts.

But the important achievement, from the point of view of cancer patients, is that any nanocage treatment would be narrowly targeted and thus avoid the side effects patients dread.

The TV and radio character the Lone Ranger used only silver bullets, allegedly to remind himself that life was precious and not to be lightly thrown away. If he still rode today, he might consider swapping silver for gold.

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Tumors Feel The Deadly Sting Of Nanobees

Tumors Feel The Deadly Sting Of Nanobees

  1:59:00 AM    Science Lover

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When bees sting, they pump poison into their victims. Now the toxin in bee venom has been harnessed to kill tumor cells by researchers at Washington University School of Medicine in St. Louis. The researchers attached the major component of bee venom to nano-sized spheres that they call nanobees.

Bee on a finger. Researchers have recently harnessed the toxin in bee venom
to kill tumor cells. (Credit: iStockphoto/Tatiana Buzuleac)

In mice, nanobees delivered the bee toxin melittin to tumors while protecting other tissues from the toxin's destructive power. The mice's tumors stopped growing or shrank. The nanobees' effectiveness against cancer in the mice is reported in advance online publication Aug. 10 in the Journal of Clinical Investigation.

"The nanobees fly in, land on the surface of cells and deposit their cargo of melittin which rapidly merges with the target cells," says co-author Samuel Wickline, M.D., who heads the Siteman Center of Cancer Nanotechnology Excellence at Washington University. "We've shown that the bee toxin gets taken into the cells where it pokes holes in their internal structures."

Melittin is a small protein, or peptide, that is strongly attracted to cell membranes, where it can form pores that break up cells and kill them.

"Melittin has been of interest to researchers because in high enough concentration it can destroy any cell it comes into contact with, making it an effective antibacterial and antifungal agent and potentially an anticancer agent," says co-author Paul Schlesinger, M.D., Ph.D., associate professor of cell biology and physiology. "Cancer cells can adapt and develop resistance to many anticancer agents that alter gene function or target a cell's DNA, but it's hard for cells to find a way around the mechanism that melittin uses to kill."

The scientists tested nanobees in two kinds of mice with cancerous tumors. One mouse breed was implanted with human breast cancer cells and the other with melanoma tumors. After four to five injections of the melittin-carrying nanoparticles over several days, growth of the mice's breast cancer tumors slowed by nearly 25 percent, and the size of the mice's melanoma tumors decreased by 88 percent compared to untreated tumors.

The researchers indicate that the nanobees gathered in these solid tumors because tumors often have leaky blood vessels and tend to retain material. Scientists call this the enhanced permeability and retention effect of tumors, and it explains how certain drugs concentrate in tumor tissue much more than they do in normal tissues.

But the researchers also developed a more specific method for making sure nanobees go to tumors and not healthy tissue by loading the nanobees with additional components. When they added a targeting agent that was attracted to growing blood vessels around tumors, the nanobees were guided to precancerous skin lesions that were rapidly increasing their blood supply. Injections of targeted nanobees reduced the extent of proliferation of precancerous skin cells in the mice by 80 percent.

Overall, the results suggest that nanobees could not only lessen the growth and size of established cancerous tumors but also act at early stages to prevent cancer from developing.

"Nanobees are an effective way to package the useful, but potentially deadly, melittin, sequestering it so that it neither harms normal cells nor gets degraded before it reaches its target," Schlesinger says.

If a significant amount of melittin were injected directly into the bloodstream, widespread destruction of red blood cells would result. The researchers showed that nanoparticles protected the mice's red cells and other tissues from the toxic effects of melittin. Nanobees injected into the bloodstream did not harm the mice. They had normal blood counts, and tests for the presence of blood-borne enzymes indicative of organ damage were negative.

When secured to the nanobees, melittin is safe from protein-destroying enzymes that the body produces. Although unattached melittin was cleared from the mice's circulation within minutes, half of the melittin on nanobees was still circulating 200 minutes later. Schlesinger indicates that is long enough for the nanobees to circulate through the mice's bloodstream 200 times, giving them ample time to locate tumors.

"Melittin is a workhorse," says Wickline, also professor of medicine in the Cardiovascular Division and professor of physics, of biomedical engineering and of cell biology and physiology. "It's very stable on the nanoparticles, and it's easily and cheaply produced. We are now using a nontoxic part of the melittin molecule to hook other drugs, targeting agents or imaging compounds onto nanoparticles."

The core of the nanobees is composed of perfluorocarbon, an inert compound used in artificial blood. The research group developed perfluorocarbon nanoparticles several years ago and have been studying their use in various medical applications, including diagnosis and treatment of atherosclerosis and cancer. About six millionths of an inch in diameter, the nanoparticles are large enough to carry thousands of active compounds, yet small enough to pass readily through the bloodstream and to attach to cell membranes.

"We can add melittin to our nanoparticles after they are built," Wickline says. "If we've already developed nanoparticles as carriers and given them a targeting agent, we can then add a variety of components using native melittin or melittin-like proteins without needing to rebuild the carrier. Melittin fortunately goes onto the nanoparticles very quickly and completely and remains on the nanobee until cell contact is made."

The flexibility of nanobees and other nanoparticles made by the group suggests they could be readily adapted to fit medical situations as needed. The ability to attach imaging agents to nanoparticles means that the nanoparticles can give a visible indication of how much medication gets to tumors and how tumors respond.

"Potentially, these could be formulated for a particular patient," Schlesinger says. "We are learning more and more about tumor biology, and that knowledge could soon allow us to create nanoparticles targeted for specific tumors using the nanobee approach."

Funding from the National Institutes of Health and the American Heart Association supported this research.

If you like this post, buy me a Pittza at $1!

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