Chapter 19 TARGETED NANOPARTICLES FOR MOLECULAR IMAGING AND THERAPY

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Chapter 19

TARGETED NANOPARTICLES FOR MOLECULAR IMAGING AND THERAPY

A Multi-Modality Approach to Molecular Medicine Shelton D. Caruthers ^1,2 , Samuel A. Wickline ¹ , Gregory M. Lanza ¹

1 Washington University, St. Louis, MO, USA; ² Philips Medical Systems, Cleveland, OH, USA

Abstract: The emerging era of personalized medicine offers the potential for early diagnosis and, therefore, treatment of pathology close to onset. Molecular imaging will allow noninvasive phenotypic characterization of pathologies leading to segmentation of patients for custom-tailored therapy. Additionally, the application and efficacy of this targeted therapy will be monitored non- invasively via molecular imaging. Nanoparticulate agents are being intensively researched as formulation platforms for various targeted clinical applications.

As exemplified by perfluorocarbon nanoparticles, these new agents in combination with the rapid innovations in imaging hardware and software will allow the emergence of new diagnostic and therapeutic paradigms.

Keywords: Perfluorocarbon nanoparticles, multimodality imaging, MRI, ultrasound, applications in cardiovascular and oncology, monitoring therapy effectiveness

1. INTRODUCTION

Recent advances in the interrelated fields of genomics, proteomics, molecular imaging and targeted drug delivery have prompted a paradigm shift in medical diagnosis and therapy from a ‘one-size-fits-all’ strategy to an approach tailored to the individual ^1,2 . Requiring novel contrast agents, the general tactic of molecular imaging is to recognize and characterize presymptomatic, early disease, which otherwise would be difficult or

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© 2006 Springer. Printed in the Netherlands.

impossible to detect using routine imaging techniques. In essence, the intent of biochemical imaging agents is to provide non-invasive assessments of pathology analogous to the use of immuno-histochemistry. However, such characterization is quite complex, requiring the simultaneous detection and

G. Spekowius and T. Wendler (Eds.), Advances in Healthcare Technology,

quantification of multiple epitopes, and thus far, molecular imaging agents have been primarily focused on detecting single biomarkers as a pathognomonic signature of disease. Nonetheless, techniques for cellular and molecular imaging have been developed, and are continually being improved, for virtually every imaging modality including nuclear ^3,4 , optical ^4-6 , CT ^7,8 , ultrasound ⁹ , and MRI ^10,11 .

With specific examples of current and imminent applications, this chapter addresses molecular imaging using a liquid perfluorocarbon (PFC) nanoparticle platform that can be detected and quantified by multiple modalities and, through specific targeting, offers great potential for diagnosis, therapy, and monitoring.

2. GENERALIZED PLATFORM FOR TARGETED MOLECULAR IMAGING

Success with molecular imaging contrast agents requires not only reaching and binding the specific target, but also being able to detect the agent once bound ¹² . While optical ^13,14 and nuclear ^15,16 techniques are exquisitely sensitive to their agents, MRI, for instance, is not as sensitive.

With MR molecular imaging agents (see Figure 19-1), a common theme is to deliver to important (but sparse) biochemical epitopes a large ‘payload’ of paramagnetic metal to overcome the scarcity. One example, dendrimers, can bear a vast number of metal atoms to a target site ^17,18 , but as yet, limited utility as a targeted agent in vivo has been established. Nanoparticles, due to their inherently great surface area, have been shown to be a more effective platform, with predominately two classes: liposomes ^19-21 and emulsions ^22,23 .

2.1 Paramagnetic liposomes

Paramagnetic liposomes, initially instituted as an instrument to overcome the partial volume dilution problem, are vesicles with a lipid bilayer membrane that enclose hefty payloads of gadolinium in an aqueous volume.

Unfortunately, for MRI applications, the limited water diffusivity across the

liposomal membranes drastically reduces the efficacy of the paramagnetic

chelates entrapped within ^24,25 . More effectively, amphipathic paramagnetic

chelates have been incorporated directly into lipid membranes ^26,27 , which

improves interactions between the relaxation agent and surrounding water

thus augmenting r 1 relaxivity.

Figure 19-1. Generalized paradigm for targeted nanoparticle contrast agents. The core provides a skeleton onto which volumes of the targeting system, imaging ‘payload’ and even therapeutic agents can be placed. Such amplification strategies permit molecular imaging to be feasible when targeting sparse epitopes. (Reproduced by courtesy of Medicamundi ²⁸ ).

In 1998, Sipkins et al. ²⁹ first reported a paramagnetic polymerized liposome targeted to α ν β 3 for the detection of angiogenesis in a cancer tumor model. This biotinylated liposome system, complexed through avidin to a pre-targeted biotinylated antibody, provided early in vivo examples that sparse pathologic biomarkers were detectable via MR molecular imaging with ‘ultraparamagnetic’ particles. Later, Hood et al. ³⁰ , in a mouse melanoma model, reported a therapeutic application using a similar construct, again, targeted to the α ν β 3 -integrins of angiogenic blood vessels, to deliver a mutant Raf gene and thereby induce tumor regression.

2.2 Paramagnetic perfluorocarbon emulsions

The second major class of paramagnetic nanoparticles is emulsions, the most notable being a liquid perfluorocarbon-based emulsion that has been targeted to multiple cardiovascular and oncological markers ^22,31-36 .

PFC nanoparticles (see Figure 19-1), in contradistinction to bubbles and liposomes, consist of a liquid PFC core encapsulated by a phospholipid monolayer and can be functionalized for targeted molecular imaging by attaching homing ligands into this monolayer ^31,35-37 . With typical nominal diameters of 200 to 300 nm, PFC nanoparticles are endowed with an enormous surface area to transport, and concentrate, payloads such as radioactive and/or paramagnetic metals to important vascular biomarker sites.

This translates into, e.g. for the case of paramagnetic agents with ~100,000

gadolinium atoms per nanoparticle, particulate r 1 relaxivities greater than

2,000,000 (mM

s) ^-1 at 1.5T ³⁸ .

While the surface area available for exploitation is an invaluable attribute, the core is also key. Owing to the properties of the perfluorocarbons in their core, PFC nanoparticles, unique from other oil-based emulsions, exhibit robust stability against handling, pressure, heat and shear. The liquid core, representing 98% by volume of the nanoparticle, can be any one of a number of PFCs (e.g., perfluorooctylbromide (PFOB), perfluoro-15-crown-5 ether (CE), perfluorodichlorooctane), with PFOB being the most commonly used.

Thus, within the core of each PFOB nanoparticle, there is a concentration of roughly 100M of ¹⁹ F.

The carbon-fluorine bond is not only chemically and thermally stable, but also essentially biologically inert. Having been used for liquid ventilation, oxygen delivery and imaging, the biocompatibility of most liquid fluorocarbons has been well documented to be, even at extremely generous doses, innocuous and physiologically inactive ^39,40 . Regarding pure fluoro- carbons within the range of 460 – 520 MW, no toxicity, carcinogenicity, mutagenicity or teratogenic effects have been reported. A reversible increase in pulmonary residual volumes has been reported for very large doses of PFC emulsions (i.e., complete blood transfusion), in macaque, swine and rabbits, but this increase was not seen in mouse, dog or human ^41-43 . PFC nanoparticle distribution and clearance data fit well into a biexponential function with a circulatory half-life in excess of one hour (300 min ± 110 min, Lanza et al., unpublished). The tissue half-life residencies range from 4 days for PFOB to as much as 65 days for perfluorotripropylamine. The fate of fluorocarbons is not metabolism, but rather gradual reintroduction by lipid carriers into the circulation, in dissolved form, where they are ultimately released in the lungs and exhaled ³⁹ .

3. LIQUID PERFLUOROCARBON NANO- PARTICLES FOR TARGETED IMAGING AGENTS

3.1 Fibrin targeted nanoparticles

The first in vivo demonstration of adapting PFC nanoparticles for site-

targeted molecular imaging was in the mid-1990’s wherein the nanoparticles

were homed to fibrin via an antifbrin monoclonal antibody and a three-step

avidin-biotin technique ²² (see Figure 19-2). The contrast agent, bound to

intravascular thrombus in canines, was detected with ultrasound imaging

based on the inherent acoustic properties of liquid PFC nanoparticles ^44-46 .

Figure 19-2. Ultrasound imaging of femoral artery thrombus before (a) and after (b) exposure to targeted PFC nanoparticles. The acute thrombus is poorly visualized with a 7.5-MHz linear-array, focused transducer. (a) The transmural electrode (Anode) and the wall boundaries of the femoral artery are clearly delineated. (b) After exposure to the targeted emulsion, the thrombus is easily visualized. (Reprinted with permission from Circulation ²² ).

Later, this acoustic agent was further modified for MR molecular imaging by attaching gadolinium chelates onto the outer surface ⁴⁷ , and again applied not only toward the non-invasive in vivo detection of thrombi in canine models (see Figure 19-3) but also in vitro detection of microthrombi deposits in human endarterectomy specimens ³⁷ . Although fibrin clots offer an abundance of binding sites that results in relatively easy detection on T1- weighted MR images, these ‘ultraparamagnetic’ nanoparticles are designed to have ample payloads (50,000 - 100,000 Gd-chelate molecules per nano- particle) ³⁸ such that minute concentrations (i.e., picomolar) of targeted agent can be conspicuously visualized ⁴⁸ .

Figure 19-3. A scanning electron micrograph (a) shows the fibrin tendrils of a clot. Targeted nanoparticles densely decorate the fibrin (b), bringing >50,000 Gd chelates per binding site.

The effect, in this canine model (c) is a marked enhancement (arrow), as compared to the

control clot in the contralateral vein. (Reprinted with permission from Circulation ³⁷ ).

Figure 19-4. CT images of human plasma clots treated with either iodinated oil or PFOB nanoparticles targeted to fibrin. Radio-opaque particles were mixed with nanoparticles of safflower oil for graduation of targeted contrast effects. (Reprinted with permission from Acad. Radiol. ⁵⁰ ).

More recently, the fibrin-targeted nanoparticles have been imaged using CT. While PFOB, due mostly to the bromine, is radio-opaque, other emulsions have also been designed to be more conspicuous on CT. The first results of a fibrin-targeted nanoparticle molecular imaging agent were presented in 2003 ^49,50 wherein the CT agent provided a four-fold increase in contrast-to-noise between targeted and non-targeted fibrin clots in vitro. In Figure 19-4 ⁵⁰ the CT images of ~3mm-diameter fibrin clots demonstrate the graduation of attenuation as the amount of fibrin-targeted, iodinated nanoparticles is varied.

Regardless of modality, the ability to detect and localize non-invasively the exposed fibrin associated with fissured atherosclerotic plaque is of great value in discerning the difference between plaques that are restricting the lumen but stable versus those at greater risk for catastrophic rupture ^51-53 . Current standards call for therapeutic intervention when the luminal narrowing exceeds 50%, but ironically, it is often those plaques of more moderate grade that rupture leading to stroke or heart attack and frequently sudden death ^54-56 . Furthermore, while the detection of lumenally-exposed fibrin may prove useful as a sine qua non for unstable plaque, others ^55,57 have shown that angiogenesis within the vasa vasorum is also a requirement for growth and development of atherosclerotic plaques, but at a far earlier stage than rupture. Thus, molecular imaging of angiogenesis offers a portal to observe early atherosclerosis, in addition to other important pathologies.

3.2 Molecular imaging of angiogenesis

Just as physicians and scientists interested in cardiovascular diseases are

keen to detect and manage angiogenesis, so are many others in fields such as

oncology and rheumatology. Angiogenesis is a complex process with many

biomarkers, but one molecular signature, the α ν β ³ integrin, has attracted

significant attention as a target ^58-63 . The α ν β ³ integrin, which plays a critical part during the formation of new blood vessels ⁶⁴ , is expressed on activated endothelial cells but not on mature quiescent cells of established vessels ⁵⁸ . Thus, α ν β ³ is a popular recipient for angiogenesis-targeted applications.

The utility of paramagnetic PFC nanoparticles to image the expression of α ν β 3 on angiogenic vessels associated with not only early atherosclerosis but also nascent tumors 58-61,63,65,66

has been ascertained in vivo 28,34-36,67

.

As an example, atherosclerotic rabbits fed high cholesterol diets for eighty days develop early expansion of the vasa vasorum in the adventia of coronaries and aortic arteries, which continues to fuel plaque progression as the diffusional limits from the arterial lumen are exceeded. If these rabbits are then given intravenous injections of α ν β ³ -targeted paramagnetic nanoparticles, significant MRI signal enhancement can be detected (heterogeneously distributed) along the aorta (see Figure 19-6); whereas little signal enhancement can be seen in the aortas of animals receiving a control diet ³⁶ . These findings were corroborated with histology.

Importantly, the specificity of nanoparticle binding was verified through competitive blockade experiments wherein high-avidity α ν β ³ -targeted non- paramagnetic nanoparticles (i.e., invisible on MRI) were injected in excess into hypercholesterolemic rabbits to saturate the binding sites. When α ν β ³ - targeted paramagnetic nanoparticles were later injected, no significant enhancement was detected ³⁶ (see Figure 19-5)– thus confirming that T1- weighted MRI signal enhancement was due to the specific binding of paramagnetic nanoparticles to the α ν β ³ marker of angiogenesis resulting from early-stage atherosclerotic disease.

Figure 19-5. Analysis of angiogenesis from data of Figure 19-6 as MRI signal enhancement

from the entire aorta (left) and skeletal muscle (right) after treatment with either α ν β 3 -targeted

(squares and diamonds) or nontargeted (circles) paramagnetic nanoparticles in cholesterol-fed

or control diet groups. In atherosclerotic rabbits, the enhancement at two hours is significantly

greater (P<0.05) than all other groups, which remained virtually unchanged. Importantly, if

α ν β 3 -integrins are pre-saturated with non-paramagnetic nanoparticles, the signal enhancement

is competitively blocked (triangles). (Reprinted with permission from Circulation ³⁶ ).

Besides atherosclerosis, cancer is an important disease in which angiogenesis plays a vital role and is widely targeted for molecular imaging.

As a model of breast cancer, Vx-2 tumors were implanted into the hindlimb of New Zealand white rabbits and the induced neovasculature was imaged

ν 3

magnetic nanoparticles ³⁵ . Using a clinical 1.5T MRI and dedicated surface receive coils, high-resolution 3D T1-weighted images were acquired dyna- mically to track the accumulation of nanoparticles within the tumor versus the surrounding musculature. For the two hours imaged, the signal intensity increased as a function of time with the final targeted contrast enhancement of the neovasculature having increased 126% over baseline, twice the change in MR signal due to passive extravascular leakage alone. The neovascular- related signal enhancement was heterogeneous and typically located asymmetrically around the tumor capsule, at interfaces between tumor and muscle, and in neighboring vasculature (see Figure 19-7). Again, in vivo competitive binding experiments, wherein the α ν β 3 -integrins were pre- saturated with targeted non-paramagnetic nanoparticles then followed by α ν β 3 - targeted paramagnetic nanoparticles, demonstrated receptor specificity of the targeted agent.

Further substantiating these findings are similar studies, with equivalent results and histological corroboration, in a tumor model of human melanoma (C-32) implanted into athymic mice ³⁴ . The Vx-2 model has also been repeated with α ν β ³ -integrin-targeted nanoparticles adapted for nuclear imaging with dramatic increases in detection sensitivity. Using single pinhole, planar nuclear imaging, Figure 19-8 presents an image of a Vx-2 tumor following α ν β ³ -targeted ^99m Tc nanoparticles. Note the prominent contrast enhancement of the tumor as well as enhancement of the testis and epiphyseal heads of long bones in the leg. In these young rabbits, angiogenesis is a prominent and natural feature of the maturing epididymis (which is highly vascular) and the growth plates of the long bones. All sites are specifically targeted and identified by the expression of α ν β ³ -integrin, which is not seen in the mature vasculature of the muscle and surrounding tissues. α ν β ³ -Integrin targeted ¹¹¹ In nanoparticles have also shown significant promise with similar in vivo findings.

with MRI twelve days later with intravenously −injected α β -targeted para-

Figure 19-6. In vivo MR images of hyperlipidemic rabbit aorta. (Top) From renal artery to diaphragm, the vessel wall enhances heterogeneously due to the α ν β 3 targeted nanoparticles.

(Bottom) Transverse images before (Pre) and after (Post) nanoparticles, after segmentation of the vessel wall (Segmented), and the final image (Enhancement) with color encoding of the percent enhancement resulting from the contrast agent bound to the marker of angiogenesis in this model of early atherosclerosis. (Reprinted with permission from Circulation ³⁶ ).

Figure 19-7. In this Vx-2 tumor model, the T1-weighted MR signal enhancement from the α ν β 3 -targeted paramagnetic nanoparticle is overlaid on the baseline image (left). The integrin is expressed heterogeneously around the 3mm tumor capsule and other nearby areas of angiogenesis. The histology (right) corroborates the heterogeneous distribution of α ν β 3 - integrin (dark stain). (Reproduced by courtesy of Medicamundi ²⁸ ).

Figure 19-8. Planar nuclear image of Vx-2 tumor in the hind limb of a rabbit following α ν β 3 -

targeted ^99m Tc nanoparticles. Note the prominent contrast enhancement of the tumor as well

as other areas of natural angiogenesis in this young, maturing rabbit.

4. THERAPEUTIC PERFLUOROCARBON NANOPARTICLES

As briefly presented above, the capability to target and image specific molecular biomarkers with perfluorocarbon nanoparticles provides the unique opportunity to 1) segment patient populations for the presence and severity of disease, 2) deliver potent individualized therapy directly to disease sites, and 3) monitor the treatment response through noninvasive imaging ^33,68 . This emerging paradigm of personalized medicine has been demonstrated in several animal models using targeted therapeutic nanoparticles ^33,69 , which are laced with therapeutic agents either by dissolving lipophilic drugs into the phospholipid monolayer or by fixing lipophobic drugs onto the outer surface with a lipophilic anchor. The drugs, then, are transferred to targeted cells via ‘contact facilitated drug delivery,’

wherein the transfer of lipids and associated drugs between the nanoparticle monolayer and the cell membrane is enabled by ligand-directed binding ^10,33,70 (see Figure 19-9). In contrast, nanoparticles that do not bind (i.e., non-targeted) retain greater than 90% of their apportioned drug. Lanza et al. ³³ elucidated this concept first in vitro demonstrating that targeted therapeutic nanoparticles (delivering, e.g., paclitaxel) had a significant effect on smooth muscle cell proliferation, whereas non-targeted therapeuctic nanoparticles had no effect.

Figure 19-9. Schematic representation of contact-facilitated drug delivery. Phospholipids and drug within the perfluorocarbon nanoparticle surfactant exchange with lipids of the target membrane through a convection process rather than diffusion, as is common among other targeted systems. Without the stable membrane contact, enabled through specific binding, drug is not released from the particle. (Reprinted with permission from J Nucl Cardiol ⁷⁰ ).

An in vivo example of targeted nanoparticle drug delivery is that

performed by Winter et al. ⁶⁹ in atherosclerotic rabbits, which, when fed high

cholesterol diets, develop early neovascular expansion of the vasa vasorum,

which fuels plaque progression ^71,72 . Delivered intravenously, α ν β ³ -targeted

paramagnetic nanoparticles laden with fumagillin, a potent antiangiogenic drug also known in its water soluble form as TNP-470 ⁷³ , markedly pruned the nascent vessels from the vasa vasorum when measured one week later (using drug-free α ν β ³ -targeted paramagnetic nanoparticles as described in the previous section) (See Figure 19-10). Hyperlipidemic rabbits given

‘control’ nanoparticles (i.e., α ν β ³ -targeted paramagnetic nanoparticles without drug) at baseline had no change in angiogenic vessel density when imaged one week after treatment. Moreover, nontargeted fumagillin nanoparticles also had no significant effect on the neovasculature – again, substantiating that without ‘contact facilitated drug delivery’ little drug is released from the nanoparticles into the circulation. By way of corroboration, histological assessments showed that the level of angiogenesis in those animals receiving targeted drug-carrying nanoparticles was significantly reduced in comparison with the vasa vasorum expansion noted in rabbits receiving non-targeted or targeted drug-free nanoparticles. It is interesting to note that the total injected drug dosage of fumagillan associated with these α ν β ³ -targeted therapeutic nanoparticles was about 50,000 times lower than the total cumulative dose given orally in previous studies ⁷³ with similar outcomes. This mechanism of locally concentrating drug via site-targeted delivery, coupled with the virtual absence of drug release unless bound, provides therapeutic nanoparticles with enormous potential for achieving results while reducing side effects.

Figure 19-10. Using α ν β 3 -targeted paramagnetic nanoparticles allows both the detection of angiogenesis and the delivery of anti-angiogenic therapy. At the time of treatment (baseline) the level of angiogenesis is the same in both groups of cholesterol-fed atherosclerotic rabbits.

One week after the single treatment, the effect of the drug is clear, but animals receiving drug- free nanoparticles exhibit no therapeutic effects. (Reprinted with permission from ⁷⁴ ).

Angiogenesis-Related MRI Enhancement (%)

5. QUANTIFICATION OF PERFLUOROCARBON NANOPARTICLES

The examples above have shown that not only can specific biomarkers be explicitly visualized through the use of targeted nanoparticles, but they can also be the sole recipients of locally-intensified drug delivery. Moreover, the follow-up monitoring of therapeutic efficacy has been exemplified. But with this new archetype of particulate drug delivery, where serum levels of drug are meaningless, new models of measuring and predicting therapeutic effects will be required ^10,75 . Perhaps the best method will be simply to quantify non- invasively the amount of drug amassed at binding sites. Thus, the unique ability to image quantitatively these therapeutic nanoparticles would be of incalculable benefit for determining local drug concentrations and developing new pharmacokinetic models of drug transport and personalized response. Furthermore, given the complexity of pathology, the ability to independently interrogate multiple biomarkers would greatly facilitate the phenotyping of disease and allow better prediction and monitoring of individualized response to therapy. Depending on the imaging modality, PFC nanoparticles can be quantified in a variety of manners.

Some modalities are inherently quantitative. The detection of nanoparticles labeled with radioisotopes, for example, gives an absolute number of counts per minute. MRI, on the other hand, gives images with relative signal intensity simultaneously weighted by a multitude of factors.

Even so, Winter et al., in the hyperlipidemic rabbit study ⁶⁹ , were able to observe a relationship between initial T1-weighted signal enhancement at the time of treatment with targeted paramagnetic therapeutic nanoparticles and the resultant reduction in angiogenesis.

More precise, however, would be traditional techniques rooted in

quantitative MR imaging such as T1 mapping ⁷⁶ , etc. In typical MRI, contrast

agents interact with their immediate milieu resulting in changes in both T1

and T2 ^* relaxation phenomena of the hydrogen nuclei. For well-behaved

systems, these effects can be modeled to predict signal intensity as a function

of contrast agent concentration. This approach has been described by

Morawski et al. ⁴⁸ for predicting the minimum concentration of targeted

paramagnetic nanoparticles with known relaxivities that must be bound

within an imaging voxel to produce conspicuous enhancement. Importantly,

this group also used T1 measurement techniques in combination with these

models to calculate the number of tissue-factor-targeted nanoparticles bound

to smooth muscle cells in culture. This algorithm, while more precise than

quantifying from signal intensity alone, remains limited by assumptions

about relaxivities, water exchange rates, molecular geometries, and other

confounding effects.

An alternative to modeling or measuring signal change as a function of contrast agent concentration would be to measure the agent directly, i.e., detecting a unique signal that otherwise would not be present. In the case of PFC nanoparticles, that alternative exists with the nuclei of ¹⁹ F. Actually, fluorine is a strong candidate for MR spectroscopy and imaging because ¹⁹ F, with a spin ½ nucleus and a natural abundance of 100%, has a gyromagnetic ratio close to that of ¹ H and a relative sensitivity 83% that of ¹ H. With essentially no ¹⁹ F in the human body, there is no background signal.

Additionally, owing to its nine electrons, ¹⁹ F has a greater range and sensitivity to its environment via chemical shift than hydrogen does. This is the foremost reason ¹⁹ F MRI has been used in a wide variety of applications such as the study of tumor metabolism ^77-79 , the mapping of physiologic pO 2

tension ^80-82 , and the characterization of liquid ventilation ^83,84 .

Figure 19-11. ¹⁹ F MR spectroscopy and imaging of liquid PFC nanoparticles bound to fibrin clots. The four ¹⁹ F spectra (left) acquired from the corresponding four clots (right) demonstrate the changing concentrations of the two nanoparticles applied. The three ¹⁹ F images (right), which have no proton background, represent three ‘weightings’ of the same four clots. Without selectivity (NS), all clots have high signal. Employing a species-selective excitation allows independent visualization of the bound CE or PFOB nanoparticles.

After exploring the minimum detection limits of PFC nanoparticles with

19 F spectroscopy ⁴⁸ , Morawski et al. ⁸⁵ went on to employ ¹⁹ F spectroscopy

and imaging at 4.7T to quantify the varying amounts of PFC nanoparticles

bound to in vitro clots and in human endarterectomy samples. Extending

for the first time this application to clinical systems and utilizing rapid

steady-state imaging techniques ⁸⁶ , ¹⁹ F image-based quantification of fibrin-

bound nanoparticles was demonstrated. Further, this group demonstrated

simultaneous independent detection and imaging of multiple unique species

of PFC nanoparticles in varying volumes (see Figure 19-11).

6. CONCLUSION

The synergistic combination of targeted drug delivery and molecular imaging has revolutionary potential. PFC nanoparticles, homed to specific biomarkers, provide the non-invasive tools to characterize pathology at the molecular level. With the promise of extending to a multi-spectral palette, this technique offers the potential to characterize phenotypically the biochemical nature or therapeutic sensitivity of disease. Furthermore, image- based confirmation and measurement of localized drug delivery and serial monitoring of biomarker response permits tailored treatment. Coupling the rapid developments in nanotechnology, genomics, proteomics, molecular biology, and imaging systems has catalyzed the multi-disciplinary field of molecular imaging, thus helping to usher in the era of personalized medicine.

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