Jeffrey Perkel has been a scientific writer and editor since 2000. He holds a PhD in Cell and Molecular Biology from the University of Pennsylvania, and did postdoctoral work at the University of Pennsylvania and at Harvard Medical School.
Drug development studies almost always begin in the test tube—or more probably, the microtiter plate. But if a compound is ever to transition to the clinic, it must move from microplate well into animals. Such preclinical studies are a must in pharmaceutical development, as compounds that look promising in a cell culture dish may not work in animals or may produce unexpected and undesirable side effects.
There fundamentally are two approaches to animal studies. One is to administer a compound, sacrifice the animal after some period of time and examine its organs to see what happened. But that requires a lot of animals, not to mention hands-on time. Plus, it makes long-term (longitudinal) studies difficult if not impossible, as it requires comparisons between different animals sacrificed at different times.
The other in vivo approach circumvents those problems. In this case, animals are probed by noninvasive imaging rather than sacrificed, meaning individual animals can be studied over time. That cuts down on the number of animals required to complete a study, and the time required to analyze them.
Researchers have several options when it comes to in vivo imaging in terms of the kinds and numbers of modalities available, ease of clinical translation and price. If you find yourself in the market for an in vivo imaging system, or you are considering migrating your research into the small animal arena, read on. With the diversity of systems now available, you should have no problem identifying one that meets your needs.
In vivo imaging modalities
San Diego-based service provider BioLaurus Inc. offers its customers—mostly Big Pharma and big biotech companies, according to chief scientific officer, Mario Bourdon—access to the full range of imaging modalities. There’s optical imaging based on fluorescence and bioluminescence, as well as the more translatable modalities: SPECT, PET, CT, microCT and MRI.
SPECT, or “single-photon emission computed tomography,” images gamma particle emission from an injected radioisotope or radiolabeled compound. Images are collected at multiple angles to produce a three-dimensional reconstruction of the sample.
PET, or “positron emission tomography,” also relies on radioactive compounds (higher energy than in SPECT, but with a shorter lifetime generally). But with PET, the emitted positron degrades into two gamma particles that shoot out at 180 degrees relative to each other, making it relatively easy to map molecules in 3D. “The problem with PET is that you can imagine from the [release of the] initial positron to the degradation into two gamma particles; there is some time elapsed,” explains Mat Brevard, vice president of preclinical imaging for North America at Bruker BioSpin. As a result, “There’s some blurring of the point source.”
One PET source, fluorine-18 (F18), for instance, has a “sphere” of 1.1 mm, Brevard says. Thus, F18-based PET imaging (for instance, using F18-fluorodeoxyglucose, a marker of brain activity) has a built-in positional uncertainty of 1.1 mm.
CT, or “computed tomography,” is essentially a high-resolution 3D X-ray, and microCT is a higher-resolution version built especially for small animal work.
Finally, MRI, or “magnetic resonance imaging,” measures the movement of molecules in a strong magnetic field to map anatomic features, fluid motion and so on.
PET, SPECT, CT and MRI are all considered “translatable” modalities in that they all are already being used on human patients in the clinic. Thus, if a company develops a SPECT tracer for its compound for use in animals, that same compound could theoretically also be put into humans. A fluorescently tagged molecule for optical imaging, though, cannot be so easily translated, as the combination of such imaging agents and fluorophores is not yet approved for use in humans.
One exception is indocyanine green, which is FDA-approved but untargeted. The situation could be changing, however. LI-COR Biosciences’ IRDye 800CW, a bright near-infrared dye that emits at 800 nm, fused to a therapeutic antibody, is being investigated in a Dutch clinical trial of breast cancer patients.
At BioLaurus, says Bourdon, most studies look at biodistributions—determining where in the body a particular compound goes and when, and how quickly it is excreted. The company also does a fair bit of imaging toxicology testing, Bourdon says, which is a more direct indication of early compound toxicity. “It can range from looking at changes in myocardial function using ultrasound … to looking at changes for instance, in markers of renal function.”
Among the company’s clients, optical modalities—fluorescence and bioluminescence—are probably the least popular, says Bourdon, both because of the inability to directly translate results to the clinic and the fact that optical modalities are typically restricted to mice. That’s because the instruments are small, but also because the efficiency of light penetration and light emission drops with increasing tissue depth. That means a deeply embedded tumor is harder to see in, say, a dog than a mouse.
Most commonly, the company tends to use a combination of PET and CT or MRI. CT and MRI provide anatomic detail, and PET (or SPECT) pinpoints a tracer’s location and pharmacokinetics to a particular organ or even a subregion of the organ. “That makes a very powerful combination,” Bourdon says.
Imaging providers
In the past few years, the in vivo imaging arena has contracted as corporate mergers removed once-familiar names from the marketplace. Bruker BioSpin acquired the Preclinical imaging Division of Carestream Health (a provider of optical imaging equipment originally named Eastman Kodak’s Health Group) in 2012, as well as SkyScan, a microCT company. In 2006, Caliper Life Sciences purchased Xenogen, which developed the popular IVIS® line of in vivo imagers. In late 2010, Caliper acquired Cambridge Research & Instrumentation (CRI), another optical imaging company. Revvity, in turn, acquired Caliper in 2011, as well as (in 2010) VisEn, a company specializing in fluorescence tomographic (3D) imaging.
According to Anna Christensen, Revvity’s director of marketing for in vivo imaging products, the IVIS imagers enable fluorescence, bioluminescence and radioisotopic Cerenkov imaging, as well as (in some models) microCT, X-ray and DyCE (dynamic contrast enhancement), a technique that enables spectral unmixing for biodistribution studies, Christensen says.
The IVIS line includes the IVIS Kinetic, IVIS Lumina II, IVIS Lumina XR, IVIS Spectrum and the IVIS SpectrumCT. The IVIS Lumina XR, for instance, combines planar (2D) bioluminescence, fluorescence and X-ray imaging; the Spectrum allows for tomographic imaging; and the IVIS SpectrumCT adds microCT capabilities. Cerenkov imaging uses the exquisite sensitivity of bioluminescence detectors to capture the weak photon emission of radioactive materials—the effect that causes some radioactive materials to literally “glow”—thereby allowing researchers to image PET or SPECT tracers optically.
“We can detect a wide range of PET and SPECT compounds in the animal, and that’s enormously enabling when looking at toxicology applications,” Christensen says.
Revvity’s FMT® line (FMT 1000, FMT 2000 and FMT 4000) enables 3D fluorescent tomography (as opposed to 2D planar imaging), and its Quantum FX microCT provides rapid, high-resolution, 3D X-ray imaging.
The Quantum, says Christensen, is a “game changer” in the microCT world, as it is both fast and relatively low-dose. “Instead of doing just five mice a day, you can do 100 at 17 seconds per scan, at doses of about 12 milliGray”—low enough for long-term imaging studies.
Multimodal imagers like the IVIS SpectrumCT simplify studies by enabling researchers to probe multiple events or datasets simultaneously. For instance, they might monitor the impact of a radiolabeled drug on a fluorescent protein-expressing tumor, while at the same time collecting anatomic detail via microCT. They then obtain a unified view of the biology of the system by overlaying the different datasets, a process called co-registration.
But researchers don’t necessarily need to use multimodal imagers to co-register their data, Christensen says. As long as the datasets are all tomographic, they can be overlaid in silico, for instance using Revvity’s imaging software suite. The company does help simplify matters, though, by providing a transferrable animal immobilization “cassette” for the IVIS or FMT imagers that also can snap into the Quantum FX microCT.
Bruker BioSpin also offers its customers a range of imaging modalities. The company has four preclinical magnetic resonance imagers, the BioSpec, ClinScan, PharmaScan and ICON. It also offers (via the Carestream acquisition) optical imagers like the MS FX PRO and Xtreme, which include planar X-ray and radioisotopic imaging; a pair of microCT instruments (SkyScan 1176 and 1178); and a combination PET/SPECT/CT imager called Albira.
According to Brevard, Bruker’s MR imagers all offer slightly different capabilities and features. The ICON, for instance, is a 1-Tesla benchtop instrument that, unlike clinical imagers, uses a permanent magnet rather than a superconducting magnet, meaning it requires very little maintenance.
The PharmaScan and BioSpec MRs feature stronger magnetic fields—the PharmaScan is available at 4.7T or 7T, and the BioSpec is available from 4.7T to 17.2T—but they also use superconducting magnets, which must be cooled with liquid nitrogen and liquid helium. (MRI resolution increases with magnetic field strength, but so does price. A small-animal 7T imager costs under $1 million, Brevard estimates, whereas an 11.7T instrument might cost over $3 million.)
The ClinScan is specifically intended for translational studies, as it uses a Siemens MR interface. “It’s a translational tool for hospitals that are very used to running a Siemens scanner,” Brevard explains.
Bruker also enables dataset co-registration. MR and CT datasets may be directly overlaid, as both are 3D. To add optical datasets, the company offers an accessory called MARS (multimodal animal rotation system), which rotates the animal inside the optical imager to provide tomographic capabilities.
Also active in the in vivo imaging field is LI-COR Biosciences. The company’s Pearl® Impulse imager provides fluorescence imaging only, specifically at 680 nm and 800 nm (near-infrared).
The Pearl Impulse uses laser excitation to rapidly excite and image these fluors. According to Jeff Harford, senior product marketing manager at LI-COR Biosciences, it takes just 20 to 30 seconds to collect a white light, 680 nm and 800 nm image of a single animal. The Pearl’s wide dynamic range of six orders of magnitude means researchers can image an animal over time using the same settings without saturating the detector.
According to Harford, the instrument is used widely in oncology research, and also for biodistribution studies of infrared imaging agents. At the moment, however, the Pearl Impulse is used mostly by academics, he says, as pharmaceutical companies wait to see the utility of fluorescent tags before embracing that technology.
Before you buy
In vivo imaging systems cost anywhere from tens of thousands to hundreds of thousands or even millions of dollars. LI-COR’s Pearl Impulse costs about $50,000, Harford says, whereas MRIs can run into the millions. According to Christensen, users can spend anywhere from $100,000 to $600,000 on a Revvity in vivo imaging system, depending on the instrument and its features.
As a result, imagers often are found at large corporations and in shared facilities, such as core facilities. For a one-off experiment, or if money is limiting, researchers should consider outsourcing their imaging work to a company like BioLaurus. That’s especially true if they also are unfamiliar with the different modalities, their strengths and weaknesses, and how to process and interpret the data—not to mention the chemical expertise needed to develop custom probes.
“The specialized instruments, people and [imaging/analytic] software is something biopharma clients can plug in and out without investing the millions to set it up themselves,” Bourdon says.
The image at the top of the page is from Revvity.