Perfluorocarbon Nanoparticles Research Articles

Physicochemical signatures of nanoparticle-dependent complement activation.

Nanoparticles are potentially powerful therapeutic tools that have the capacity to target drug payloads and imaging agents. However, some nanoparticles can activate complement, a branch of the innate immune system, and cause adverse side-effects. Recently, we employed an in vitro hemolysis assay to measure the serum complement activity of perfluorocarbon nanoparticles that differed by size, surface charge, and surface chemistry, quantifying the nanoparticle-dependent complement activity using a metric called Residual Hemolytic Activity (RHA). In the present work, we have used a decision tree learning algorithm to derive the rules for estimating nanoparticle-dependent complement response based on the data generated from the hemolytic assay studies. Our results indicate that physicochemical properties of nanoparticles, namely, size, polydispersity index, zeta potential, and mole percentage of the active surface ligand of a nanoparticle, can serve as good descriptors for prediction of nanoparticle-dependent complement activation in the decision tree modeling framework.

Perfluorocarbon nanoparticles: evolution of a multimodality and multifunctional imaging agent.

Perfluorocarbon nanoparticles offer a biologically inert, highly stable, and nontoxic platform that can be specifically designed to accomplish a range of molecular imaging and drug delivery functions in vivo. The particle surface can be decorated with targeting ligands to direct the agent to a variety of biomarkers that are associated with diseases such as cancer, cardiovascular disease, obesity, and thrombosis. The surface can also carry a high payload of imaging agents, ranging from paramagnetic metals for MRI, radionuclides for nuclear imaging, iodine for CT, and florescent tags for histology, allowing high sensitivity mapping of cellular receptors that may be expressed at very low levels in the body. In addition to these diagnostic imaging applications, the particles can be engineered to carry highly potent drugs and specifically deposit them into cell populations that display biosignatures of a variety of diseases. The highly flexible and robust nature of this combined molecular imaging and drug delivery vehicle has been exploited in a variety of animal models to demonstrate its potential impact on the care and treatment of patients suffering from some of the most debilitating diseases.

Atherosclerosis endothelial activation quantification in vivo with fluorine magnetic resonance imaging and spectroscopy

Background Endothelial activation is one of the necessary initial steps in the formation of atherosclerotic plaque that facilitates immune cell recruitment and retention. To image and quantify early markers of endothelial inflammation, we recently developed a multiplexing peptide-based targeting system for post-formulation functionalization of perfluorocarbon (19F) nanoparticles (PFC NP) that is intrinsically quantifiable at a binding site based on magnetic resonance spectroscopy. The approach is now demonstrated in vivo to detect and quantify VCAM-1 in ApoE-/- mouse aorta. Methods

Anti-angiogenesis therapy in the Vx2 rabbit cancer model with a lipase-cleavable Sn 2 taxane phospholipid prodrug using α(v)β₃-targeted theranostic nanoparticles.

In nanomedicine, the hydrophobic nature of paclitaxel has favored its incorporation into many nanoparticle formulations for anti-cancer chemotherapy. At lower doses taxanes are reported to elicit anti-angiogenic responses. In the present study, the facile synthesis, development and characterization of a new lipase-labile docetaxel prodrug is reported and shown to be an effective anti-angiogenic agent in vitro and in vivo. The Sn 2 phosphatidylcholine prodrug was stably incorporated into the lipid membrane of αvβ3-integrin targeted perfluorocarbon (PFC) nanoparticles (αvβ3-Dxtl-PD NP) and did not appreciably release during dissolution against PBS buffer or plasma over three days. Overnight exposure of αvβ3-Dxtl-PD NP to plasma spiked with phospholipase enzyme failed to liberate the taxane from the membrane until the nanoparticle integrity was compromised with alcohol. The bioactivity and efficacy of αvβ3-Dxtl-PD NP in endothelial cell culture was as effective as Taxol® or free docetaxel in methanol at equimolar doses over 96 hours. The anti-angiogenesis effectiveness of αvβ3-Dxtl-PD NP was demonstrated in the Vx2 rabbit model using MR imaging of angiogenesis with the same αvβ3-PFC nanoparticle platform. Nontargeted Dxtl-PD NP had a similar MR anti-angiogenesis response as the integrin-targeted agent, but microscopically measured decreases in tumor cell proliferation and increased apoptosis were detected only for the targeted drug. Equivalent dosages of Abraxane® given over the same treatment schedule had no effect on angiogenesis when compared to control rabbits receiving saline only. These data demonstrate that αvβ3-Dxtl-PD NP can reduce MR detectable angiogenesis and slow tumor progression in the Vx2 model, whereas equivalent systemic treatment with free taxane had no benefit.

Relaxometric studies of gadolinium‐functionalized perfluorocarbon nanoparticles for MR imaging

Fluorine MRI ((19) F MRI) is receiving an increasing attention as a viable alternative to proton-based MRI ((1) H MRI) for dedicated application in molecular imaging. The (19) F nucleus has a high gyromagnetic ratio, a 100% natural abundance and is furthermore hardly present in human tissues allowing for hot spot MR imaging. The applicability of (19) F MRI as a molecular and cellular imaging technique has been exploited, ranging from cell tracking to detection and imaging of tumors in preclinical studies. In addition to applications, developing new contrast materials with improved relaxation properties has also been a core research topic in the field, since the inherently low longitudinal relaxation rates of perfluorocarbon compounds result in relatively low imaging efficiency. Borrowed from (1) H MRI, the incorporation of lanthanides, specifically Gd(III) complexes, as signal modulating ingredients in the nanoparticle formulation has emerged as a promising approach to improvement of the fluorine signal. Three different perfluorocarbon emulsions were investigated at five different magnetic field strengths. Perfluoro-15-crown-5-ether was used as the core material and Gd(III)DOTA-DSPE, Gd(III)DOTA-C6-DSPE and Gd(III)DTPA-BSA as the relaxation altering components. While Gd(III)DOTA-DSPE and Gd(III)DOTA-C6-DSPE were favorable constructs for (1) H NMR, Gd(III)DTPA-BSA showed the strongest increase in (19F) R(1). These results show the potential of the use of paramagnetic lipids to increase (19F) R(1) at clinical field strengths (1.5-3 T). At higher field strengths (6.3-14 T), gadolinium does not lead to an increase in (19F) R(1) compared with emulsions without gadolinium, but leads to an significant increase in (19F) R(2). Our data therefore suggest that the most favorable situation for fluorine measurements is at high magnetic fields without the inclusion of gadolinium constructs.

Abstract 18726: Thrombin-Inhibiting Nanoparticles Prevent Growth of Fibrin Clots by Inhibiting and Adhering to Clot-Bound Thrombin

Introduction: Sustained and localized prevention of clotting via thrombin inhibition could provide efficient care without significant bleeding effects for clinical scenarios including angioplasty, myocardial infarction, microangiopathy, and acute kidney injury. Thrombin-inhibiting perfluorocarbon nanoparticles (PFC NPs) carrying the direct inhibitors bivalirudin (BVR) and PPACK have demonstrated minimal bleeding effects with inhibition of and binding at sites of acute thrombosis in mouse and rabbit models. Here, we demonstrate that thrombin-inhibiting PFC NPs inhibit clot-bound thrombin and seal clots against further growth when exposed to plasma. Methods and Results: In solution, BVR, PPACK, BVR NPs, and PPACK NPs demonstrated concentration-dependent inhibition of Fibrinopeptide A (FPA) production by thrombin, as evaluated by ELISA. FPA ELISA for human plasma exposed to human fibrin clots demonstrated that BVR, PPACK, BVR NPs, and PPACK NPs inhibited thrombin bound in fibrin clots to the same extent as in solution. Fibrin clots were incubated with non-targeted NPs, BVR NPs, or PPACK NPs. 19F MRI at 12T demonstrated that BVR and PPACK NPs specifically bind to clots (figure 1A). Clots treated with BVR, PPACK, BVR NPs, PPACK NPs, or saline, were submerged in human plasma and imaged sequentially over one hour in T2-weighted 1H MRI at 12T. Clots treated with saline, BVR, or PPACK grew in volume when exposed to plasma, adding loose fibrin gel. Clots treated with BVR or PPACK NPs did not grow significantly (figure 1B-E). Conclusions: Thrombin-inhibiting PFC NPs have previously been shown to specifically bind to sites of acute thrombotic injury. Data presented herein show that PPACK and BVR NPs inhibit activity of clot-bound thrombin and specifically bind to fibrin clots derived from human plasma. PPACK and BVR NPs bound on the surface of fibrin clots prevent further clot growth on exposure to plasma.

Preclinical Development Of a Nanomedicne Approach For Multiple Myeloma Targeting The Myc Oncoprotein

PurposeThis study investigated alpha 4 beta 1/ Very Late Antigen-4 (α4β1/ VLA-4)-integrin targeted nanotherapy approach to deliver a new lipase-labile prodrug. Experimental DesignA phospholipid-based MYC-MAX inhibitor prodrug (MI1-PD) was synthesized, and its inherent anti-proliferate potency was compared to the lipid-free compound (MI1) using mouse multiple myeloma (MM) cell line (5TGM1). VLA-4-targeted perfluorocarbon (PFC) nanoparticles binding to 5TGM1 cells was measured and compared to biomarker expression assessed with flow cytometry using antibodies. The efficacy of MI1-PD incorporated into non-targeted and VLA-4-targeted PFC NP exposed to 5TGM1 cells was assessed with MTT assays, Annexin V and cell cycle analysis. ResultsMI1-PD was more potent by several orders of magnitude than its free drug counterpart in culture. Targeted NP binding correlated well with biomarker expression assessment by flow cytometry in 5TGM1 cells. Non-targeted NPs had no appreciable binding to 5TGM1 cells. High anti-MM potency of MI1-PD was noted in VLA-4-targeted NPs compared to the non-targeted NPs demonstrating that the efficacy was dependent on expression of the targeted biomarker to afford particle-to-cell drug delivery. ConclusionsThese results suggest the feasibility of an improved integrin VLA-4- targeted nanotherapy approach to deliver a lipase- labile prodrug construct, MI1-PD. Disclosures:No relevant conflicts of interest to declare.

Application of a hemolysis assay for analysis of complement activation by perfluorocarbon nanoparticles

Nanoparticles offer new options for medical diagnosis and therapeutics with their capacity to specifically target cells and tissues with imaging agents and/or drug payloads. The unique physical aspects of nanoparticles present new challenges for this promising technology. Studies indicate that nanoparticles often elicit moderate to severe complement activation. Using human in vitro assays that corroborated the mouse in vivo results we previously presented mechanistic studies that define the pathway and key components involved in modulating complement interactions with several gadolinium-functionalized perfluorocarbon nanoparticles (PFOB). Here we employ a modified in vitro hemolysis-based assay developed in conjunction with the mouse in vivo model to broaden our analysis to include PFOBs of varying size, charge and surface chemistry and examine the variations in nanoparticle-mediated complement activity between individuals. This approach may provide the tools for an in-depth structure-activity relationship study that will guide the eventual development of biocompatible nanoparticles. From the Clinical EditorUnique physical aspects of nanoparticles may lead to moderate to severe complement activation in vivo, which represents a challenge to clinical applicability. In order to guide the eventual development of biocompatible nanoparticles, this team of authors report a modified in vitro hemolysis-based assay developed in conjunction with their previously presented mouse model to enable in-depth structure-activity relationship studies.

Assessing intrarenal nonperfusion and vascular leakage in acute kidney injury with multinuclear 1H/19F MRI and perfluorocarbon nanoparticles

We sought to develop a unique sensor-reporter approach for functional kidney imaging that employs circulating perfluorocarbon nanoparticles and multinuclear (1) H/(19) F MRI. (19) F spin density weighted and T1 weighted images were used to generate quantitative functional mappings of both healthy and ischemia-reperfusion (acute kidney injury) injured mouse kidneys. (1) H blood-oxygenation-level-dependent (BOLD) MRI was also employed as a supplementary approach to facilitate the comprehensive analysis of renal circulation and its pathological changes in acute kidney injury. Heterogeneous blood volume distributions and intrarenal oxygenation gradients were confirmed in healthy kidneys by (19) F MRI. In a mouse model of acute kidney injury, (19) F MRI, in conjunction with blood-oxygenation-level-dependent MRI, sensitively delineated renal vascular damage and recovery. In the cortico-medullary junction region, we observed 25% lower (19) F signal (P < 0.05) and 70% longer (1) H T2* (P < 0.01) in injured kidneys compared with contralateral kidneys at 24 h after initial ischemia-reperfusion injury. We also detected 71% higher (19) F signal (P < 0.01) and 40% lower (1) H T2* (P < 0.05) in the renal medulla region of injured kidneys compared with contralateral uninjured kidneys. Integrated (1) H/(19) F MRI using perfluorocarbon nanoparticles provides a multiparametric readout of regional perfusion defects in acutely injured kidneys.

Characterization of early neovascular response to acute lung ischemia using simultaneous 19F/1H MR molecular imaging

Angiogenesis is an important constituent of many inflammatory pulmonary diseases, which has been unappreciated until recently. Early neovascular expansion in the lungs in preclinical models and patients is very difficult to assess noninvasively, particularly quantitatively. The present study demonstrated that (19)F/(1)H MR molecular imaging with αvβ3-targeted perfluorocarbon nanoparticles can be used to directly measure neovascularity in a rat left pulmonary artery ligation (LPAL) model, which was employed to create pulmonary ischemia and induce angiogenesis. In rats 3days after LPAL, simultaneous (19)F/(1)H MR imaging at 3T revealed a marked (19)F signal in animals 2h following αvβ3-targeted perfluorocarbon nanoparticles [(19)F signal (normalized to background)=0.80±0.2] that was greater (p=0.007) than the non-targeted (0.30±0.04) and the sham-operated (0.07±0.09) control groups. Almost no (19)F signal was found in control right lung with any treatment. Competitive blockade of the integrin-targeted particles greatly decreased the (19)F signal (p=0.002) and was equivalent to the non-targeted control group. Fluorescent and light microscopy illustrated heavy decorating of vessel walls in and around large bronchi and large pulmonary vessels. Focal segmental regions of neovessel expansion were also noted in the lung periphery. Our results demonstrate that (19)F/(1)H MR molecular imaging with αvβ3-targeted perfluorocarbon nanoparticles provides a means to assess the extent of systemic neovascularization in the lung.

Molecular MR Imaging of Neovascular Progression in the Vx2 Tumor with αvβ3-Targeted Paramagnetic Nanoparticles

To assess the dependence of neovascular molecular magnetic resonance (MR) imaging on relaxivity (r1) of αvβ3-targeted paramagnetic perfluorocarbon (PFC) nanoparticles and to delineate the temporal-spatial consistency of angiogenesis assessments for individual animals. Animal protocols were approved by the Washington University Animal Studies Committee. Proton longitudinal and transverse relaxation rates of αvβ3-targeted and nontargeted PFC nanoparticles incorporating gadolinium diethylenetrianime pentaacedic acid (Gd-DTPA) bisoleate (BOA) or gadolinium tetraazacyclododecane tetraacetic acid (Gd-DOTA) phosphatidylethanolamine (PE) into the surfactant were measured at 3.0 T. These paramagnetic nanoparticles were compared in 30 New Zealand White rabbits (four to six rabbits per group) 14 days after implantation of a Vx2 tumor. Subsequently, serial MR (3.0 T) neovascular maps were developed 8, 14, and 16 days after tumor implantation by using αvβ3-targeted Gd-DOTA-PE nanoparticles (n = 4) or nontargeted Gd-DOTA-PE nanoparticles (n = 4). Data were analyzed with analysis of variance and nonparametric statistics. At 3.0 T, Gd-DTPA-BOA nanoparticles had an ionic r1 of 10.3 L · mmol(-1) · sec(-1) and a particulate r1 of 927000 L · mmol(-1) · sec(-1). Gd-DOTA-PE nanoparticles had an ionic r1 of 13.3 L · mmol(-1) · sec(-1) and a particulate r1 of 1 197000 L · mmol(-1) · sec(-1). Neovascular contrast enhancement in Vx2 tumors (at 14 days) was 5.4% ± 1.06 of the surface volume with αvβ3-targeted Gd-DOTA-PE nanoparticles and 3.0% ± 0.3 with αvβ3-targeted Gd-DTPA-BOA nanoparticles (P = .03). MR neovascular contrast maps of tumors 8, 14, and 16 days after implantation revealed temporally consistent and progressive surface enhancement (1.0% ± 0.3, 4.5% ± 0.9, and 9.3% ± 1.4, respectively; P = .0008), with similar time-dependent changes observed among individual animals. Temporal-spatial patterns of angiogenesis for individual animals were followed to monitor longitudinal tumor progression. Neovasculature enhancement was dependent on the relaxivity of the targeted agent.

Multifunctional nanoparticles for detection, quantification, and treatment of endothelial erosions in experimental atherosclerosis

Acute atherothrombotic occlusion in heart attack and stroke implies disruption of the vascular endothelial barrier that exposes a highly procoagulant intimal milieu. However, the evolution, severity, and pathophysiological consequences of vascular barrier damage in advanced atherosclerotic plaque remain unknown, in part because quantifiable methods and experimental models are lacking for its in vivo assessment. Here, we report a multifunctional perfluorocarbon (PFC) nanoparticle (NP) for delineation of endothelial erosions as well as targeted antiinflammatory drug delivery to the erosion site. Detection and quantification (MRS and fluorescence imaging, shown in Figure) of endothelial erosions were evaluated in ApoE−/− mice fed a high fat diet for 4 – 7 months. Endothelial barrier disruption was apparent from massive and rapid passive permeation and intimal trapping of PFC nanoparticles (~250nm diameter) that normally do not penetrate intact endothelial barriers after in vivo circulation for 2 hours. RT‐PCR (CCL2, Npy) and immunohistological (I‐kB protein)results suggested that treatment (6 doses, i.v. every other day) PFC nanoparticles loaded with NF‐kB inhibitors (NEMO binding domain peptides) in 4 month fed ApoE−/− mice downregulated inflammation in aortas. Accordingly, PFC NP may be useful for defining disease stage and for following therapy to heal unstable plaques.

Quantitative molecular imaging of angiogenesis-targeted fluorinated nanoparticles: new approaches for B1-mapping compensation for 19F-MRI

Background Quantitative MR molecular imaging allows for the detection of targeted contrast agents to diagnose disease states and monitor response to therapy, such as anti-angiogenic therapy in atherosclerosis and cancer with aνb3-integrin targeted perfluorocarbon (PFC) nanoparticles. Recently, 19 FM R using a 19 F/ 1 H dual-tuned RF coil has been utilized to directly image and quantify the fluorinated core of these PFC nanoparticle (NP) emulsions. However, low concentrations of these fluorine agents in the body, in conjunction with varying RF coil sensitivity profiles (B1field inhomogeneities) raise obstacles to accurate quantification. This study presents a strategy to more accurately quantify the sparse 19 Fs ignal from PFC NP emulsions with a 1 H image-based Actual Flip-angle Imaging (AFI) B1-mapping correction to the 19 Fa nd 1 Hi mages.

Programmable nanoparticle functionalization forin vivotargeting

The emerging demand for programmable functionalization of existing base nanocarriers necessitates development of an efficient approach for cargo loading that avoids nanoparticle redesign for each individual application. Herein, we demonstrate in vivo a postformulation strategy for lipidic nanocarrier functionalization with the use of a linker peptide, which rapidly and stably integrates cargos into lipidic membranes of nanocarriers after simple mixing through a self-assembling process. We exemplified this strategy by generating a VCAM-1-targeted perfluorocarbon nanoparticle for in vivo targeting in atherosclerosis (ApoE-deficient) and breast cancer (STAT-1-deficient) models. In the atherosclerotic model, a 4.1-fold augmentation in binding to affected aortas was observed for targeted vs. nontargeted nanoparticles (P<0.0298). Likewise, in the breast cancer model, a 4.9-fold increase in the nanoparticle signal from tumor vasculature was observed for targeted vs. nontargeted nanoparticles (P<0.0216). In each case, the nanoparticle was registered with fluorine ((19)F) magnetic resonance spectroscopy of the nanoparticle perfluorocarbon core, yielding a quantitative estimate of the number of tissue-bound nanoparticles. Because other common nanocarriers with lipid coatings (e.g., liposomes, micelles, etc.) can employ this strategy, this peptide linker postformulation approach is applicable to more than half of the available nanosystems currently in clinical trials or clinical uses.

Rapid quantification of oxygen tension in blood flow with a fluorine nanoparticle reporter and a novel blood flow-enhanced-saturation-recovery sequence

We present a novel blood flow-enhanced-saturation-recovery (BESR) sequence, which allows rapid in vivo T1 measurement of blood for both (1)H and (19)F nuclei. BESR sequence is achieved by combining homogeneous spin preparation and time-of-flight image acquisition and therefore preserves high time efficiency and signal-to-noise ratio for (19)F imaging of circulating perfluorocarbon nanoparticles comprising a perfluoro-15-crown-5-ether core and a lipid monolayer (nominal size = 250 nm). The consistency and accuracy of the BESR sequence for measuring T1 of blood was validated experimentally. With a confirmed linear response feature of (19)F R1 with oxygen tension in both salt solution and blood sample, we demonstrated the feasibility of the BESR sequence to quantitatively determine the oxygen tension within mouse left and right ventricles under both normoxia and hyperoxia conditions. Thus, (19)F BESR MRI of circulating perfluorocarbon nanoparticles represents a new approach to noninvasively evaluate intravascular oxygen tension.

Bivalirudin Emulsions Demonstrate Efficacy of a Nanoparticle Strategy for Inhibition and Imaging of Thrombosis

A thrombin‐inhibiting perfluorocarbon nanoparticle (PFC NP), functionalized by PPACK (Phe(D)‐Pro‐Arg‐Chloromethylketone), was recently presented as a prototype for a novel class of targeted antithrombotic. Here, a NP functionalized with Bivalirudin (BVR), was compared to BVR and the PPACK NP.PPACK or BVR were covalently attached to PFC NPs. Optical assay verified that PPACK and BVR selectivity and activity against thrombin was not diminished on the NPs. In vivo activity was assessed for PPACK NPs, PPACK, BVR, BVR NPs, heparin, non‐functionalized NPs, or saline in C57BL6 mice subjected to laser injury of the carotid artery. Time to thrombotic occlusion of the injured artery was assessed via Doppler flow measurement. Selected arteries were excised to assess NP retention via 19F magnetic resonance (MR). 3T MRI of thrombosis following administration of PPACK NPs was also demonstrated in cholesterol‐fed rabbits.Previously, PPACK NPs outperformed both heparin (p=.001) and PPACK (p=.0006) in delaying occlusion of the carotid artery. PPACK or non‐functionalized NPs failed to delay occlusion of the carotid artery. BVR NPs significantly delayed occlusion (p=.02) whereas an equivalent dose of free BVR (120 mg/kg) did not (figure 1a). 19F MR captured thrombin‐specific PFC NP retention in occluded arteries of mice (figure 1b) and rabbits (figure 2).

Magnetic resonance chemical exchange saturation transfer imaging and nanotechnology

Chemical exchange saturation transfer (CEST) agents and paramagnetic CEST (PARACEST) agents display bound water signals that exchange protons with the bulk water. CEST magnetic resonance imaging (MRI) relies on exchangeable protons that resonate at a chemical shift that is distinguishable from the bulk water signal. In some cases, paramagnetic chelates are utilized to shift the bound water frequency further away from the bulk water. Radiofrequency prepulses applied at the appropriate frequency can saturate the exchangeable protons, which transfer into the bulk water pool and lead to reduced equilibrium magnetization. Therefore, CEST and PARACEST agents allow the image contrast to be switched 'on' and 'off' by simply changing the pulse sequence parameters. One of the main limitations with this approach is the inherent insensitivity of MRI to CEST and PARACEST agents. Nanoscale carriers have been developed to improve the limit of detection for these agents, demonstrating the feasibility of in vivo molecular or cellular MRI based on CEST or PARACEST contrast. These carriers have been based on a number of different nanoparticle constructs, such as liposomes, dendrimers, polymers, adenovirus particles, and perfluorocarbon nanoparticles. The unique MRI properties of CEST and PARACEST nanoparticle systems have spawned research into an array of potential medical applications.

Visualization of inflammation using 19F‐magnetic resonance imaging and perfluorocarbons

Inflammation plays a central pathophysiological role in a large number of diseases. While conventional magnetic resonance imaging (MRI) can depict gross tissue alterations due to proton changes, specific visualization of inflammation is an unmet task in clinical medicine. (19) F/(1) H MRI is a novel technology that allows tracking of stem and immune cells in experimental disease models after labelling with perfluorocarbon (PFC) emulsions. (19) F markers such as PFC compounds provide a unique signal in vivo due to the negligible (19) F background signal of the body. Concomitant acquisition of (1) H images places the labelled cells into their anatomical context. This novel imaging technique has been applied to monitor immune cell responses in myocardial infarction, pneumonia, bacterial abscess formation, peripheral nerve injury, and rejection of donor organs after transplantation. Upon systemic application PFC nanoparticles are preferentially phagozytosed by circulating monocytes/macrophages and, thus, the fluorine signal in inflamed organs mainly reflects macrophage infiltration. Moreover, attenuation of the inflammatory response after immunosuppressive or antibiotic treatments could be depicted based on (19) F/(1) H-MRI. Compared to other organ systems (19) F-MRI of neuroinflammation is still challenging, mainly because of lack in sensitivity. In focal cerebral ischemia early application of PFCs revealed ongoing thrombotic vessel occlusion rather than cell migration indicating that timing of contrast agent application is critical. Current restrictions of (19) F/(1) H-MRI in sensitivity may be overcome by improved imaging hardware, imaging sequences and reconstruction techniques, as well as improved label development and cell labelling procedures in the future.

PPACK and Bivalirudin nanoparticles enable simultaneous imaging and potent inhibition of acute clotting

Summary Perfluorocarbon nanoparticles functionalized for thombin inhibition with Bivalirudin or PPACK were tested as an inhibitor of thrombin in vitro and in acute thrombosis models. The particles significantly inhibited occlusive arterial thrombi. The particles also manifested binding providing magnetic resonance contrast highlighting thrombi.

Probing atherosclerotic angiogenesis with new manganese-based nanocolloid for T1-weighted MRI

Summary This study describes a novel T1-weighted MR molecular imaging approach for sparse epitopes, such as the avb3integrin receptor in atherosclerotic angiogenesis, utilizing a soft Mn-based nanoparticle with high-relaxivity augmented by minute amounts of surface gadolinium. This new agent utilizes about 1/30th the amount of lanthanide used for PFC paramagnetic particles and reduces gadolinium exposure by 300-fold compared with clinical single dose Gd-DTPA. Background We have previously reported in hyperlipidemic rabbit models that integrin-targeted gadolinium perfluorocarbon nanoparticles can effectively assess plaque angiogenesis using MRI. However, the concern for Nephrogenic Systemic Fibrosis (NSF) has motivated the development of alternative technologies that dramatically lower Gd use. We have developed a manganese oleate based nanoparticle with high longitudinal relaxivity for fibrin-specific imaging of thrombus. However, for sparse receptors, such as neovascular integrins, preliminary studies revealed that paramagnetic nanoparticles with greater T1w effects were needed. The objective of this research was to evaluate a manganese-gadolinium nanocolloid (MnGd NC) with gadolinium levels of less than 300X current clinical dosage levels, while providing high levels of T1-weighted MR imaging of sparse angiogenic avb3-integrin expression in vivo. Methods A new nanocolloid comprised of a bivalent manganese oleate (0.49±0.02 mg Mn/ml) /polysorbate core encapsulated with phospholipid surfactant enriched with 1.25 mole% Gd-DOTA-cholesterol and 0.3 mole% of quinolone-derived peptidomimetic avb3-integrin antagonistcoupled to phosphatidylethanolamine through a PEG (2000) spacer was produced (Diam., 134±2 nm.; Zeta, -25±02mv; PDI, 0.13±0.03). Hyperlipidemic New Zealand White rabbits (n=8), fed a 0.25% cholesterol diet (egg-derived) for 12 months, received avb3-integrinMnGd NC or nontargeted MnGd NC; avb3-IntegrinMnGd NC was also administered to rabbits fed a normal chow (n=4). Dynamic 3T MR imaging of the descending thoracic aorta was performed over 2 hours post-injection. Molecular imaging results were corroborated microscopically using fluoresence microscopy. Results MR signal enhancement at 2 hours was increased 18.7 ±1.95% when averaged over all slices and voxels of the aortic wall in hyperlipidemic rabbits treated with avb3integrin-MnGd NC. (Fig) Signal enhancement due to nonspecific neovascular accumulation of nontargeted MnGd NC was significantly less, (2.5±1.17%, p<0.05). Rabbits fed a normal diet and treated with avb3-integrin-MnGd NC had the signal increase, 3.17±1.94%, which did not differ from the nontargeted control. The MRI pattern observed was spatially heterogeneous along both transverse and longitudinal planes of the descending aorta, and predominantly localized between the left subclavian and the diaphragm. Immunohistochemistry corroborated the prominent, neointimal proliferation among cholesterol-fed, atherosclerotic rabbits and the