Preclinical Imaging: The Rapidly Evolving Role of Nanotechnology

	Three scientists from the RatCAP (Rat Conscious Animal PET) team at Brookhaven National Laboratory (L to R: Sean Stoll, Bosky Ravindranath and Paul Vaska) tend to the scanner that scans freely moving animals—and could some day be modified to image humans.

Researchers at Memorial Sloan-Kettering Cancer Center, along with collaborators at Cornell University and Hybrid Silica Technologies, have received approval for their first Investigational New Drug Application (IND) from the U.S. Food and Drug Administration (FDA) for an ultrasmall inorganic (silica) nanoparticle platform for tumor targeting and for the treatment of cancers in the future.

Fluorescent Cornell dots or C dots, which are cleared through the kidney, have been modified with radiolabels on their surface to create dual-modality probes for optical and PET imaging. These tiny silica spheres contain a fluorescent dye, Cy5, and peptide ligands for targeting integrin receptors on melanoma and other integrin-expressing tumors. "The first clinical trial using this silica nanoparticle [started in May] and will be performed in five metastatic melanoma patients," says Michelle S. Bradbury, MD, PhD, a clinician-scientist at Memorial Sloan-Kettering Cancer Center in New York City.  

The purpose of this clinical trial is to evaluate the behavior of this inorganic particle, which is the first of its class and properties, in humans. Using PET imaging, the biodistribution/pharmacokinetics, safety, dosimetry, and uptake of the multimodal particles in tumor and other major tissues/organs will be investigated. "We want to first make sure that in humans the particle behaves in a way that is similar to that observed in our preclinical models. With the success of this trial, we would perform tumor targeting studies in a larger cohort of patients as part of a Phase 1/2 clinical trial. We estimate that it would take about three to four years to enter the clinic," predicts Bradbury.

"One advantage of using these probes is that diseases can be targeted and treated using a single platform," says Bradbury. "Nanoparticles also offer additional benefits, such as improving the toxicity profile and/or altering the kinetic behavior of certain classes of drugs.  For instance, by attaching a drug to a tumor-targeting nanoparticle, the kinetics may be altered from that of the free drug, which typically distributes throughout the body. Particle-bound drugs are larger agents that reside for a longer time in the circulation, leading to preferential accumulation within tumor tissue. Finally, small modifications to the C dot platform, such as the switch of the iodine radiolabel from one that detects (Iodine-124) to one that treats tumors (Iodine-131), may be exploited for treating certain radiosensitive cancers (i.e., thyroid cancers), she adds.

"Nanoparticles are not going to simply replace all other probes in use. But I do think patients will benefit from a different type of targeting and/or drug delivery vehicle under certain circumstances," says Bradbury. For example, if there is a small molecule inhibitor for melanoma which cannot be used because of toxicity to normal tissues, binding the drug to a nanoparticle may redirect the drug to where the particle goes—to the kidney—and not where the drug wants to go (i.e. to the liver), she explains.

Nanotechnology also may open a new door for the treatment of liver cancer. Penn State College of Medicine researchers have evaluated the use of molecular-sized bubbles filled with C6-ceramide, called cerasomes, as an anti-cancer agent. Ceramide is a lipid molecule naturally present in the cell's plasma membrane that controls cell functions, including cell aging, or senescence.

"Cerasomes reduce the size of hepatocellular carcinoma—about 90 percent in the last three weeks of treatment," says Mark Kester, PhD, the G. Thomas Passanati professor of pharmacology at Penn State Milton S. Hershey Medical Center in Hershey, Pa. He also is the co-founder and chief medical officer of Keystone Nano, which is currently working to complete the preclinical package for cerasomes in human studies in patients with liver cancer. The same formulation also works in other solid tumors such as breast cancer and melanoma models as well as in non-solid tumors like large granular lymphocytic leukemia, adds Kester.

Ceramide nanoliposome also can extend the lifespan of hepatocellular carcinoma patients by targeted delivery of chemotherapeutic antineoplastic drugs such as Sorafenib. When patients are diagnosed with hepatocellular carcinoma, they have about seven months to live and Sorafenib adds an additional six to 10 weeks, says Kester. The first line of therapy will be to use ceramide nanoliposomes themselves and the second line would be to combine them with drugs, he adds. Liquid chromatography–mass spectrometry (LC-MS) technology and magnetic resonance imaging (MRI) would be used in humans to see how ceramide accumulates in the tumor. "It would take six to nine months to get into a human trial and then somewhere within the next two years we will be able to see how it works in hepatocellular carcinoma patients," he says.

Molecularly targeted ultrasound assesses tumor growth

PET scans of a rat’s brain made with the RatCAP scanner (horizontal view superimposed on a rat brain atlas figure, top, and a coronal slice, bottom). The rainbow scale (red = high, violet = low) indicates the level of a radiotracer that binds to receptors for dopamine, which are concentrated in the striatum, a brain region involved in reward and motivation. Source: Brookhaven National Laboratory

Ultrasound also in playing a key role in tumor assessment. Recent research published this month in Radiology, shows that contrast-enhanced ultrasound imaging with molecularly targeted microbubbles can be used for monitoring angiogenic marker expression during tumor growth. In the study, ultrasonographic microbubbles targeted with one of several antibodies—anti-integrin, anti-endoglin, or anti-vascular endothelial growth factor receptor 2 were injected into mice with implanted breast, ovarian, or pancreatic tumors. Changes in the relative uptake of microbubbles were observed as the tumors grew, suggesting that in vivo molecular profiling of tumor angiogenesis using microbubbles could be used as a diagnostic tool.

"The results provide further insights into the biology of tumor angiogenesis and may help in defining promising imaging targets for both early cancer detection and treatment monitoring using targeted contrast-enhanced ultrasonography imaging," said Jürgen K. Willmann, MD, assistant professor of radiology, Molecular Imaging Program at School of Medicine, Stanford University in Stanford, Calif., who led the study. This also could personalize therapy as microbubbles can be combined with any human antibody.

But is clinical translation feasible? "Before labeled microbubbles are introduced into the clinic, they must be shown to be safe and effective. For reimbursement, labeled microbubbles must not only impact patient care but also alter management in a cost-neutral or cost-negative manner. This will require extensive clinical testing in phase II trials," predicts Peter L. Choyke, MD, program director of Molecular Imaging Program, National Cancer Institute in Bethesda, Md., in an accompanying editorial in the same issue of Radiology.

Miniature 'wearable' PET scanner

Scientists from the U.S. Department of Energy's (DOE) Brookhaven National Laboratory, Stony Brook University and collaborators have developed a device that will give neuroscientists a new tool for simultaneously studying brain function and behavior in fully awake, moving animals. The researchers have described the tool, RatCAP—Rat Conscious Animal PET and validation studies—in the April issue of Nature Methods.

"It allows the animal to move freely while being scanned with the PET to see brain function. In other small animal scanners, the rat must be immobilized and this usually means that it must be anesthetized. Our scanner has similar spatial resolution to the highest resolution existing commercial small animal scanners, but allows the rat to be awake and moving freely during the PET scan," says David J. Schlyer, PhD, senior scientist at the medical department, Brookhaven National Laboratory in Upton, N.Y.

"Humans can be scanned now while they are awake and lying down in the PET scanner, but with a modification of our device, they could be walking or performing other tasks at the same time we are looking at brain activity and function. Such a human device does not exist at present, although the modular technology we are using could easily be expanded to a human-sized PET helmet," predicts Schlyer.

Many questions remain, such as how long the translation into clinics could take. "It would depend on many factors, including interest from the medical community and the resources available in terms of money for parts and time for working out the details of fabrication," says Schlyer.