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Nanotechnologists Seek Biological Niches

2005; Cell Press; Volume: 123; Issue: 6 Linguagem: Inglês

10.1016/j.cell.2005.12.001

1097-4172

Ivan Amato,

Supramolecular Self-Assembly in Materials

Biologists are embracing nanotechnology—the engineering and manipulating of entities in the 1 to 100 nm range—and are exploiting its potential to develop new therapeutics and diagnostics. Biologists are embracing nanotechnology—the engineering and manipulating of entities in the 1 to 100 nm range—and are exploiting its potential to develop new therapeutics and diagnostics. In late August, behind closed doors at the austere National Academy of Sciences in Washington D.C., Samuel Stupp, a materials scientist and director of the Institute for BioNanotechnology in Medicine at Northwestern University, showed a video clip for a committee evaluating the United States' billion-dollar-a-year National Nanotechnology Program. “Everyone was so quiet watching this movie. It was amazing,” recalls Clayton Teague, director of the National Nanotechnology Coordinating Office. The video's protagonist was a mouse with a damaged spinal cord that could only barely move using its front legs. Stupp's research involves engineering nanomolecules called peptide amphiphiles consisting of a hydrocarbon tail attached to a peptide into which is inserted amino-acid sequences that stimulate neurons to seek new connections with neighboring neurons. As the video clip revealed, two months after the injured mouse received an injection of Stupp's peptide amphiphiles, it was able to move (albeit awkwardly) using all four limbs. In bench tests, these same amphiphiles self-assemble into fibers with diameters in the nanometer-scale range, and the resulting nanofibers form neuron-friendly networks. Analyzing anatomical, molecular, and behavioral results from mice with spinal cord injury, Stupp and his collaborator, Northwestern neurology professor John A. Kessler, suspect that the amphiphiles, which form a gel when they self-assemble, prevent scar tissue from forming, thereby allowing initiation of the regenerative process in the injured spinal cord, a process that normally is blocked. Restoring a measure of mobility to a paralyzed animal, and the potential transfer of such a feat into human patients, is just one dramatic trajectory of the maturing and ever better funded face of biological nanotechnology and its most touted offshoot—nanomedicine. Its practitioners have entered a new era rife with video evidence like Stupp's and Kessler's, patents for sensitive molecular detectors, and nanoparticle-based contrast agents for detecting, monitoring, and treating many illnesses, from breast cancer to cold sores. Nanotechnology pioneer, Richard Smalley, won the Nobel Prize in Chemistry in 1996 for discovering buckyballs—the all-carbon, soccerball-shaped molecules that helped to push nanotechnology into high gear. Smalley, who died of leukemia in October at the age of 62, was one of the most vocal champions of the beneficial potential of nanotechnology, which he argued was destined to solve major societal and medical problems. In many of his speaking engagements since his cancer diagnosis in 1999, he asked rhetorically, “am I part of the last generation to die of cancer, or the first to be saved by nanotechnology?” Now there is a growing roster of biological nanotechnologists who are taking audacious questions like Smalley's to heart and to their laboratories. The word “nanotechnology” conjures up the notion of human innovation and control over objects and processes on the nanometer scale. This is a compelling size range because it is here that many technologically useful properties emerge from assemblies of atoms and molecules. For example, it is in this range that semiconductor materials, such as cadmium selenide, can be designed to emit a rainbow of different colors and thereby serve as biomarkers in cells. The tools of nanotechnology continue to infiltrate molecular biology whose objects of study—DNA, RNA, and proteins—are perhaps the most elegant example of nanoscale constructions. It's important to avoid confusing terms like “nanobiology,” clarifies Jeff Schloss, a program director with the National Human Genome Research Institute and a member of a working group that oversees programs within the year-old research and development framework of the U.S. National Cancer Institute (NCI), known as the Alliance for Nanotechnology in Cancer. After all, Schloss notes, anything in biology that is subcellular in size falls within the nanometer range. A typical protein like hemoglobin is about 5 nm in diameter, DNA's double helix spans about 2 nm, a mitochondrion stretches over a few hundred nm. So, says Schloss, “if we allow the stuff in cells to be considered as nanotechnology, then it becomes hopeless to separate nanotechnology from regular biology.” The distinction between the two, he says, is that “nanotechnology is not nature. Technology is something people make.” Physician Andrew C. von Eschenbach, director of the NCI, has repeatedly identified nanotechnology as central to the Institute's “Challenge Goal” of, in his own words, “eliminating suffering and death from cancer by 2015.” Says Eschenbach: “The NCI is engaged in a concerted effort to harness the power of nanotechnology to radically change the way we diagnose, treat, and prevent cancer.” In 2004, the NCI launched the $144.3 million, 5-year Alliance for Nanotechnology in Cancer. In October of this year, the Alliance announced awards totaling $26.3 million for the first-year funding of seven “centers of cancer nanotechnology excellence.” Each of these brings together interdisciplinary expertise from neighboring academic, medical, and corporate partners to achieve common goals. Such goals include the development of magnetic nanoparticles for brain tumor imaging and treatment and in vivo nanoscale sensing devices that can monitor and report on gene expression. The Alliance also announced $7-million worth of first-year awards for the establishment of a dozen “Cancer Nanotechnology Platform Partnerships,” which fund research collaborations focused on developing technologies in six program areas. These areas include the development of in vivo imaging tools and methods to assess treatment efficacy in real time. In addition to the NCI Alliance for Nanotechnology in Cancer, the NIH has launched a related R&D framework, known as the NIH Nanomedicine Roadmap Initiative, which has earmarked about $80 million through 2009 for medically directed biology research on the nanoscale. So far, the Alliance has announced awards of nearly $6 million, which will fund the creation of four Nanomedicine Development Centers. Among the awardees is the Center for Design of Biomimetic Nanoconductors based at the University of Illinois, Urbana-Champaign. There, researchers will investigate the basic biology, manipulation, and design of ion channels, protein-based pores, and other nanoscale portals by which ions, chemical signals, and other molecules can move across cell membranes. A primary goal of the center will be to cull ideas from biological systems for the design of biobatteries that could be useful in implantable devices such as artificial retinas. The European research and development community also is gearing up for a strategic push into nanomedicine. In September, stakeholders in industry, private and public research centers, and academia released a “vision paper” entitled “European Technology Platform on NanoMedicine: Nanotechnology for Health.” According to a spokesman for the European Commission's Research Directorate-General's office, this document could lead to the specification of nanomedicine as a priority area in the European Union's ongoing Framework Programmes, the latest of which encompasses 17.5 billion Euros. Even as these large-scale R&D formats are designed and funded, Fortune 500 companies and startups are racing to develop and market biological nanotechnologies. Meanwhile, the U.S. Food and Drug Administration (FDA), the Environmental Protection Agency, and other regulatory agencies in the U.S. and abroad struggle to identify potential health, environmental, and occupational hazards that could come with these new and still poorly understood products. These same agencies also are working toward protocols to manage those risks. Reports by market analysis firms claim that there are more than 100 nano-based biomedical products—including drugs, drug-delivery systems, therapeutic devices, and diagnostic tests—in various stages of development and commercialization. In January this year, the FDA approved Abraxane—described by the NCI “as the first of a new class of ‘protein-bound particle’ drugs”—for treating metastatic breast cancer patients who do not respond to chemotherapy or whose cancer returns after such treatment. Developed by American Pharmaceutical Partners in Schaumburg, Illinois and Santa Monica-based American Bioscience, Abraxane consists of molecules of the chemotherapy agent taxol (paclitaxel) attached to the blood serum protein albumin to form injectable medicinal nanoparticles. Phase III clinical trials in 454 breast cancer patients showed that Abraxane was better than Taxol alone at reducing the size of tumors (about 33% reduction for Abraxane treatment compared with 19% for Taxol alone) and at slowing tumor progression (about 22 weeks with no worsening of breast cancer for Abraxane compared with about 16 weeks for Taxol). In the standard Taxol treatment, the active drug (which is highly insoluble in water) is dissolved in a solvent derived from castor oil that can have serious side effects in patients. Because of its lower toxicity, Abraxane treatment can safely deliver more active drug to tumors. Among other investigational nanomedicines in clinical testing is Combidex, a magnetic nanoparticle that can be used to make certain cancers stand out more obviously in magnetic resonance imaging scans. These particles consist of nanoscale clumps of iron oxide wrapped in a carbohydrate shell, resulting in nanoparticles with a diameter of 40 nm. Because these nanoparticles are taken up by macrophages (immune cells present in healthy lymph nodes but not in cancerous nodes), the particles accumulate selectively in noncancerous lymph nodes. In MRI scans, therefore, healthy lymph nodes produce bigger signals, which makes them appear dark compared to cancerous nodes, which appear lighter. One aim of the technique is to help distinguish nodes that are enlarged due to cancer from those enlarged due to inflammation. Made by Advanced Magnetics Inc. in Cambridge, Massachusetts, the FDA issued this past March a so-called “approvable letter,” which indicates that the new nanoparticle-based contrast agent can be approved as long as the company provides additional clinical data. Abraxane and Combidex represent perhaps the largest class of nanoscale products in various stages of development, that is, nanoparticles. By mixing and matching various cores—made of polymers, metal oxides, semiconductor materials like cadmium selenide, and carbon nanotubes—with different coatings and molecular appendages (that add capabilities such as tissue-targeting, drug-delivery, imaging enhancement, and selective ability to absorb laser energy), researchers are developing a catalog of nanoparticles for medical applications. One of the more visible players in this arena is James Baker, director of the Michigan Nanotechnology Institute for Medicine and Biological Sciences. It was the known dangers of another emerging molecular technology in the 1990s, gene therapy, that prompted his entry into the field of nanotechnology research. He was dismayed by the death in 1999 of 18-year-old Jesse Gelsinger during a controversial human trial of an experimental gene therapy treatment. Baker says, “at that point, it struck me that what was really needed was synthetic systems” that would be less likely to elicit dangerous immune responses. For many medical applications, he knew the most basic structural criterion that such systems would have to meet: “You have to be in a certain size range to get out of blood vessels and into cells—bigger than 25 nm but smaller than 70 nm,” he says. That's right in the middle of the classic size range—1 to 100 nm—that the nanotechnology community has defined as its own. By the mid-1990s, Baker and a few dozen colleagues in a variety of disciplines began pooling their expertise with the goal of developing medically valuable nanoparticles whose structures and properties were sufficiently uniform and well characterized to meet stringent FDA requirements for any drug candidate. In the late 1990s, the NCI awarded Baker and his colleagues a $12 million grant to, in Baker's words, “develop a platform that essentially would diagnose a cancer, treat it, image the tumor, and also see if the tumor was responding.” (Baker also is the principal investigator for one of the Cancer Nanotechnology Platform Partnerships recently awarded by NCI.) The nanoparticles that Baker and his colleagues are developing as drug delivery vehicles are called dendrimers. A dendrimer is an unusually well-defined polymer structure built up using simple monomeric units from a central core in precisely controlled steps that sequentially add layers. This synthetic procedure leads to spherical bush-like structures of specified and uniform nanoscale sizes. Conventional polymers grow into linear and often branching structures, but the individual molecules range vastly in size, a nonuniformity that would be unlikely to pass muster with the FDA. The most advanced dendrimer designed by Baker's team so far is based on a PAMAM (polyamidoamine) dendrimer that contains a mechanism for directing the dendrimer to cancer cells and a means for delivering an anticancer drug, such as methotrexate, once the nanoparticle has reached its target. “Targeting drugs directly to cancer cells reduces the amount that gets to normal cells, increases the drug's anticancer effect, and reduces its toxicity,” Baker says. To achieve such targeting, the researchers attach molecules of folate to chemical “hooks” built into the PAMAM nanoparticles. Because certain types of cancer cells overexpress folate receptors, dendrimers bearing folate molecules are able to home to these cells and stick to them. First, the investigators showed that when folate-bearing PAMAM nanoparticles carrying fluorescent marker molecules were injected into mice bearing certain human tumors the dendrimers became sequestered in the cancerous tissue. With up to 120 attachment points for molecular appendages, the PAMAM dendrimers can be readily engineered to carry chemotherapy agents such as methotrexate. In the June 15 issue of Cancer Research, Baker and his collaborators report that between 30 and 40 percent of the mice receiving the dendrimer-methotrexate nanoparticles lived for the trial's entire 99 day duration, whereas all mice in the control group died in that time period. By Baker's calculations, a three month delay in tumor growth for a mouse may be equivalent to a three year delay in the progression of a tumor in human patients. Their therapeutic potential notwithstanding, the question arises of whether nanoscale particles pose toxicological dangers that nobody has yet anticipated. The small portfolio of toxicology studies in hand so far suggest that nanoparticles, such as buckyballs and related all-carbon particles known as buckytubes, indeed can pose respiratory hazards in a similar way to nanoscale soot particles in diesel exhaust. But the higher-stakes question is whether the various nanoscale particles researchers are developing, and that are winding their way into products on store shelves and doctors offices, harbor toxicities and environmental hazards that are new. For example, could Stupp's amphiphiles end up migrating from the point of injection and self-assemble elsewhere in ways that, say, interfere with blood circulation or the filtering capacity of the kidneys? “We know nanoparticles can pass through the blood brain barrier. We know they can make it through the placenta,” says Toni Marechaux, director of the Board on Manufacturing and Engineering Design of the National Research Council, a body of the National Academy of Sciences. Like many in the nanotechnology community, Marechaux and her colleagues worry that there may be heretofore unrecognized hazards posed by nanotechnologies as they enter widespread use, including those already on the market, such as sunscreens containing nanoscale particles of titanium dioxide. The toxicology studies have yet to be done, Marechaux notes. “We just don't know what the dangers might be.” That's true for every new nanoscale entity under development. Researchers in academia, industry, and government have only just begun to design and undertake health and environmental hazard studies for nanotechnologies. Early data indicate that nanoscale particles can pass through the skin and, in the case of carbon nanotubes, can lead to inflammation when inhaled into the lung, leading to the formation of small scar patches called granulomas. According to a National Toxicology Program fact sheet on nanoparticles, “particle size can impart toxicity equally if not more so than chemical composition,” a phenomenon “that hints at the complexity of the topic.” What's more, “there are indications…that manufactured nanoscale materials may distribute in the body in unpredictable ways.” “I am concerned about what could happen,” says Baker, a member of the Nanotechnology Technical Advisory Group of the President's Council of Advisors on Science and Technology. “I think it's crazy to have abject fear, but we need to be careful. If we in the field are claiming that by making particles nanoscale, we impart something unique to them, then we can't argue at the same time that we know they are safe.” The world is full of dangerous medicines and technologies that are beneficial because their toxicities and other hazards are known and manageable and their benefits are deemed to outweigh their risks, points out Teague, director of the National Nanotechnology Coordinating Office. With that reality long in place, he and other biological nanotechnology devotees believe that the same ought to hold for the nanoscale wonders that they are making real.