Nanomedicine
From bioimaging to drug delivery and therapeutics, nanotechnology is poised to change the way doctors practice medicine.
| August 1, 2014
© LAGUNA DESIGN/SCIENCE PHOTO LIBRARY/CORBIS
In a 1959 lecture at Caltech famously dubbed “There’s Plenty of Room at the Bottom,” American physicist and Nobel laureate–to-be Richard Feynman discussed the idea of manipulating structures at the atomic level. Although the applications he discussed were theoretical at the time, his insights prophesied the discovery of many new properties at the nanometer scale that are not observed in materials at larger scales, paving the way for the ever-expanding field of nanomedicine. These days, the use of nanosize materials, comparable in dimension to some proteins, DNA, RNA, and oligosaccharides, is making waves in diverse biomedical fields, including biosensing, imaging, drug delivery, and even surgery.
Nanomaterials typically have high surface area–to-volume ratios, generating a relatively large substrate for chemical attachment. Scientists have been able to create new surface characteristics for nanomaterials and have manipulated coating molecules to fine-tune the particles’ behaviors. Most nanomaterials can also penetrate living cells, providing the basis for nanocarrier delivery of biosensors or therapeutics. When systemically administered, nanomaterials are small enough that they don’t clog blood vessels, but are larger than many small-molecule drugs, facilitating prolonged retention time in the circulatory system. With the ability to engineer synthetic DNA, scientists can now design and assemble nanostructures that take advantage of ?Watson-Crick base pairing to improve target detection and drug delivery.
Both the academic community and the pharmaceutical industry are making increasing investments of time and money in nanotherapeutics. Nearly 50 biomedical products incorporating nanoparticles are already on the market, and many more are moving through the pipeline, with dozens in Phase 2 or Phase 3 clinical trials. Drugmakers are well on their way to realizing the prediction of Christopher Guiffre, chief business officer at the Cambridge, Massachusetts–based nanotherapeutics company Cerulean Pharma, who last November forecast, “Five years from now every pharma will have a nano program.”
Nano-sight
Technologies that enable improved cancer detection are constantly racing against the diseases they aim to diagnose, and when survival depends on early intervention, losing this race can be fatal. While detecting cancer biomarkers is the key to early diagnosis, the number of bona fide biomarkers that reliably reveal the presence of cancerous cells is low. To overcome this challenge, researchers are developing functional nanomaterials for more sensitive detection of intracellular metabolites, tumor cell–membrane proteins, and even cancer cells that are circulating in the bloodstream. (See “Fighting Cancer with Nanomedicine,” The Scientist, April 2014.)
The extreme brightness, excellent photostability, and ready modulation of silica nanoparticles, along with other advantages, make them particularly useful for molecular imaging and ultrasensitive detection.
Silica nanoparticles are one promising material for detecting specific molecular targets. Dye-doped silica nanoparticles contain a large quantity of dye molecules housed inside a silica matrix, giving an intense fluorescence signal that is up to 10,000 times greater than that of a single organic fluorophore. Taking advantage of Förster Resonance Energy Transfer (FRET), in which a photon emitted by one fluorophore can excite another nearby fluorophore, researchers can synthesize fluorescent silica nanoparticles with emission wavelengths that span a wide spectrum by simply modulating the ratio of the different dyes—the donor chromophore and the acceptor chromophore. The extreme brightness, excellent photostability, and ready modulation of silica nanoparticles, along with other advantages, make them particularly useful for molecular imaging and ultrasensitive detection.
THE NANOMEDICINE CABINET: Scientists are engineering nanometer-size particles made of diverse materials to aid in patient care. The unique properties of these structures are making waves in biomedical analysis and targeted therapy.See full infographic: JPG | PDF© TAMI TOLPAOther materials that are under investigation as nanodetectors include graphene oxide (GO), the monolayer of graphite oxide, which has unique electronic, thermal, and mechanical properties. Semiconductor-material quantum dots (QDs), now being developed by Shuming Nie’s group at Emory University, exhibit quantum mechanical properties when covalently coupled to biomolecules and could improve cancer imaging and molecular profiling.1 Spherical nucleic acids (SNAs), in which nucleic acids are oriented in a spherical geometry, scaffolded on a nanoparticle core (which may be retained or dissolved), are also gaining traction by the pioneering work of Chad Mirkin’s group at Northwestern University.2 (Seeillustration.)
Nanoparticles are also proving their worth as probes for various types of bioimaging, including fluorescence, magnetic resonance imaging (MRI), computed tomography (CT), and positron emission tomography (PET). For instance, Xiaoyuan Chen, now at the National Institutes of Health’s National Institute of Biomedical Imaging and Bioengineering, and Hongjie Dai of Stanford University have developed carbon nanotubes for performing PET scans in mice. When modified with the macromolecule polyethylene glycol to improve biocompatibility, the nanotubes were very stable and remained in circulation for days, far longer than the few hours typical of many molecular imaging agents.3 Further modification with a short-peptide targeting ligand called RGD caused the nanotubes to selectively accumulate in tumors that overexpressed integrin, the molecular target of RGD, enabling precise tumor imaging.
To further increase the specificity of nanodetectors, researchers can add recognition probes such as aptamers—short synthetic nucleic acid strands that bind target molecules. For example, we conjugated gold nanoparticles with aptamers that had been identified through iteratively screening DNA probes using living cancer cells.4 Circulating tumor cells (CTCs) are shed into the bloodstream from primary tumors and provide a potential target for early cancer diagnosis. However, CTCs are rare, with blood concentrations of typically fewer than 10 cells per milliliter of blood. Collaborating with physicians to profile samples from leukemia patients, we demonstrated that aptamers are capable of differentiating among different subtypes of leukemia, as well as among patient samples before and after chemotherapy (unpublished data). In addition to leukemia, we have selected aptamers specific to cancers of the lung, liver, ovaries, colon, brain, breast, and pancreas, as well as to bacterial cells. Other researchers have developed nanoparticles with numerous and diverse surface aptamers, enabling them to bind their targets more efficiently and securely.
Eye on the target
NANOCAPSULES: A false-color transmission electron micrograph of liposomes, spherical particles composed of a lipid bilayer around a central cavity that can be engineered to deliver both hydrophobic and hydrophilic drugs to specific cells in the body© DAVID MCCARTHY/SCIENCE SOURCEThe prototype of targeted drug delivery can be traced back to the concept of a “magic bullet,” proposed by chemotherapy pioneer and 1908 Nobel laureate Paul Ehrlich. Ehrlich envisioned a drug that could selectively target a disease-causing organism or diseased cells, leaving healthy tissue unharmed. A century later, researchers are developing many types of nanoscale “magic bullets” that can specifically deliver drugs into target cells or tissues.
Doxil, the first nanotherapeutic approved by the US Food and Drug Administration, is a liposome (~100 nm in diameter) containing the widely used anticancer drug doxorubicin. The therapy takes advantage of the leaky blood vasculature and poor lymphatic drainage in tumor tissues that allow the nanoparticles to squeeze from blood vessels into a tumor and stay there for hours or days. Scientists have also been developing nanotherapeutics capable of targeting specific cell types by binding to surface biomarkers on diseased cells. Targeting ligands range from macromolecules, such as antibodies and aptamers, to small molecules, such as folate, that bind to receptors overexpressed in many types of cancers.
Aptamers in particular are a popular tool for targeting specific cells. Aptamer development is efficient and cost-effective, as automated nucleic acid synthesis allows easy, affordable chemical synthesis and modification of functional moieties. Other advantages include high stability and long shelf life, rapid tissue penetration based on the relatively small molecular weights, low immunogenicity, and ease of antidote development in the case of an adverse reaction to therapy by simply administering an aptamer’s complementary DNA. We have demonstrated the principle of modifying aptamers on the surfaces of doxorubicin-containing liposomes, which then selectively delivered the drug to cultured cancer cells.5
Recent advances in predicting the secondary structures of a DNA fragment or interactions between multiple DNA strands, as well as in technologies to automatically synthesize predesigned DNA sequences, has opened the door to more advanced applications of aptamers and other DNA structures in nanomedicine. For instance, we have developed aptamer-tethered DNA nanotrains, assembled from multiple copies of short DNA building blocks. On one end, an aptamer moiety allows specific target cell recognition during drug delivery, and a long double-stranded DNA section on the other end forms the “boxcars” for drug loading. The nanotrains, which can hold a high drug payload and specifically deliver anticancer drugs into target cancer cells in culture and animal models,6 could reduce drug side effects while inhibiting tumor growth. Alternatively, Daniel Anderson of MIT engineered a tetrahedral cage of DNA, often called “DNA origami,” for folate-mediated targeted delivery of small interfering RNAs (siRNAs) to silence some tumor genes.7 And Mirkin’s SNAs can similarly transport siRNAs as “guided missiles” to knock out overexpressed genes in cancer cells. Mirkin’s group also recently demonstrated that the SNAs were able to penetrate the blood-brain barrier and specifically target genes in the brains of glioblastoma animal models.2 Peng Yin of Harvard Medical School and the Wyss Institute and others are now building even more complex DNA nanostructures with refined functions, such as smart biomedical analysis.8
Conventional assembly of such DNA nanostructures exploits the hybridization of a DNA strand to part of its complementary strand. In addition, we have discovered that DNA nanostructures called nanoflowers because they resemble a ring of nanosize petals, can be self-assembled through liquid crystallization of DNA, which typically occurs at high concentrations of the nucleic acid.9 Importantly, these DNA nanostructures can be readily incorporated with components possessing multiple functionalities, such as aptamers for specific recognition, fluorophores for molecular imaging, and DNA therapeutics for disease therapy.
Another example of novel nanoparticles is DNA micelles, three-dimensional nanostructures that can be readily modified to include aptamers for specific cell-type recognition, or DNA antisense for gene silencing. The lipid core and sphere of projecting nucleic acids can enter cells without any transfection agents and have high resistance to nuclease digestion, making them ideal candidates for drug delivery and cancer therapy.
Researchers are developing many types of nanoscale “magic bullets” that can specifically deliver drugs into target cells or tissues.
Such advances in targeting are now making it possible to deliver combinations of drugs and ensure that they reach target cells simultaneously. Paula Hammond and Michael Yaffe of MIT recently reported a liposome-based combination chemotherapy delivery system that can simultaneously deliver two synergistic chemotherapeutic drugs, erlotinib and doxorubicin, for enhanced tumor killing.10Erlotinib, an inhibitor of epidermal growth factor receptor (EGFR), promotes the dynamic rewiring of apoptotic pathways, which then sensitizes cancer cells to subsequent exposure to the DNA-damaging agent doxorubicin. By incorporating erlotinib, a hydrophobic molecule, into the lipid bilayer shell while packaging the hydrophilic doxorubicin inside of the liposomes, the researchers achieved the desired time sequence of drug release—first erlotinib, then doxorubicin—in a one-two punch against the cancer. They also demonstrated that the efficiency of drug delivery to cancer cells was enhanced by coating the liposomes with folate.
Scientists are also engineering “smart” nanoparticles, which activate only in the disease microenvironment. For example, George Church of Harvard Medical School and the Wyss Institute and colleagues invented a logic-gated DNA nanocapsule that they programmed to deliver drugs inside cells only when a specific panel of disease biomarkers is overexpressed on the cell surface.11And Donald Ingber’s group, also at Harvard Medical School and the Wyss Institute, developed microscale aggregates of thrombolytic-drug-coated nanoparticles that break apart under the abnormally high fluid shear stress of narrowed blood vessels and then bind and dissolve the problematic clot.12
With these and other nanoplatforms for targeted drug delivery being tested in animal models, medicine is now approaching the prototypic magic bullet, sparing healthy tissue while exterminating disease.
Nano-fighters
In addition to serving as mere drug carriers that deliver the toxic payload to target cells, nanomaterials can themselves function as therapeutics. For example, thermal energy is emerging as an important means of therapy, and many gold nanomaterials can convert photons into thermal energy for targeted photothermal therapy. Taking advantage of these properties, we conjugated aptamers onto the surfaces of gold-silver nanorods, which efficiently absorb near-infrared light and convert energy from photons to heat. These aptamer-conjugated nanorods were capable of selectively binding to target cells in culture and inducing dramatic cytotoxicity by converting laser light to heat.13
Magnetic nanoparticles are also attractive for their ability to mediate heat induction. Jinwoo Cheon of Yonsei University in Korea developed core–shell magnetic nanoparticles, which efficiently generated thermal energy by a magnetization-reversal process as these nanoparticles returned to their relaxed states under an external, alternating-current magnetic field.14 Using this technology, Cheon and his colleagues saw dramatic tumor regression in a mouse model.
A third type of nanosize therapeutic involves cytotoxic polymers. For example, we synthesized a nucleotide-like molecule called an acrydite with an attached DNA aptamer that specifically binds to and enters target cancer cells.15 The acrydite molecules in the resultant acrydite-aptamer conjugates polymerized with each other to form an aptamer-decorated molecular string that led to cytotoxicity in target cancer cells, including those exhibiting multidrug resistance, a common challenge in cancer chemotherapy.
A third type of nanosize therapeutic involves cytotoxic polymers. For example, we synthesized a nucleotide-like molecule called an acrydite with an attached DNA aptamer that specifically binds to and enters target cancer cells.15 The acrydite molecules in the resultant acrydite-aptamer conjugates polymerized with each other to form an aptamer-decorated molecular string that led to cytotoxicity in target cancer cells, including those exhibiting multidrug resistance, a common challenge in cancer chemotherapy.
Many other subfields have been advanced by recent developments in nanomedicine, including tissue engineering and regenerative medicine, medical devices, and vaccines. We must proceed with caution until these different technologies prove safe in patients, but nanomedicine is now poised to make a tremendous impact on health care and the practice of clinical medicine.
Guizhi Zhu is a postdoctoral associate in the Department of Chemistry and at the Health Cancer Center of the University of Florida. Weihong Tan is a professor and associate director of the Center for Research at the Bio/Nano Interface at the University of Florida. He also serves as the director of the Molecular Science and Biomedicine Laboratory at Hunan University in China, where Lei Mei is a graduate student.
Guizhi Zhu is a postdoctoral associate in the Department of Chemistry and at the Health Cancer Center of the University of Florida. Weihong Tan is a professor and associate director of the Center for Research at the Bio/Nano Interface at the University of Florida. He also serves as the director of the Molecular Science and Biomedicine Laboratory at Hunan University in China, where Lei Mei is a graduate student.
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