19F MR Imaging Research Articles

Single-Step Synthesis of Highly Sensitive 19F MRI Tracers by Gradient Copolymerization-Induced Self-Assembly.

Amphiphilic gradient copolymers are promising alternatives to block copolymers for self-assembled nanomaterials due to their straightforward synthesis via statistical copolymerization of monomers with different reactivities and hydrophilicity. By carefully selecting monomers, nanoparticles can be synthesized in a single step through gradient copolymerization-induced self-assembly (gPISA). We synthesized highly sensitive 19F MRI nanotracers via aqueous dispersion gPISA of hydrophilic poly(ethylene glycol) methyl ether methacrylate (PEGMA) with core-forming N,N-(2,2,2-trifluoroethyl)acrylamide (TFEAM). The PPEGMA-grad-PTFEAM nanoparticles were optimized to achieve spherical morphology and exceptional 19F MRI performance. Noncytotoxicity was confirmed in Panc-1 cells. In vitro 19F MR relaxometry and imaging demonstrated their diagnostic imaging potential. Notably, these gradient copolymer nanotracers outperformed block copolymer analogs in 19F MRI performance due to their gradient architecture, enhancing 19F relaxivity. The synthetic versatility and superior 19F MRI performance of gradient copolymers highlight their potential in advanced diagnostic imaging applications.

Cationic fluorinated micelles for cell labeling and 19F-MR imaging

Magnetic resonance imaging (MRI) relies on appropriate contrast agents, especially for visualizing transplanted cells within host tissue. In recent years, compounds containing fluorine-19 have gained significant attention as MRI probe, particularly in dual 1H/19F-MR imaging. However, various factors affecting probe sensitivity, such as fluorine content and the equivalency of fluorine atoms, must be considered. In this study, we synthesized fluorinated micelles with adjustable surface positive charge density and investigated their physicochemical properties and MRI efficacy in phantoms and labeled cells. While the micelles exhibited clear signals in 19F-MR spectra and imaging, the concentrations required for MRI visualization of labeled cells were relatively high, adversely affecting cell viability. Despite their favourable physicochemical properties, achieving higher labeling rates without compromising cell viability during labeling remains a challenge for potential in vivo applications.

Self-Assembled Fluorinated Nanoparticles as Sensitive and Biocompatible Theranostic Platforms for 19F MRI.

Theranostics is a novel paradigm integrating therapy and diagnostics, thereby providing new prospects for overcoming the limitations of traditional treatments. In this context, perfluorocarbons (PFCs) are the most widely used tracers in preclinical fluorine-19 magnetic resonance (19F MR), primarily for their high fluorine content. However, PFCs are extremely hydrophobic, and their solutions often display reduced biocompatibility, relative instability, and subpar 19F MR relaxation times. This study aims to explore the potential of micellar 19F MR imaging (MRI) tracers, synthesized by polymerization-induced self-assembly (PISA), as alternative theranostic agents for simultaneous imaging and release of the non-steroidal antileprotic drug clofazimine. In vitro, under physiological conditions, these micelles demonstrate sustained drug release. In vivo, throughout the drug release process, they provide a highly specific and sensitive 19F MRI signal. Even after extended exposure, these fluoropolymer tracers show biocompatibility, as confirmed by the histological analysis. Moreover, the characteristics of these polymers can be broadly adjusted by design to meet the wide range of criteria for preclinical and clinical settings. Therefore, micellar 19F MRI tracers display physicochemical properties suitable for in vivo imaging, such as relaxation times and non-toxicity, and high performance as drug carriers, highlighting their potential as both diagnostic and therapeutic tools.

Design of multi-modal antenna arrays for microwave hyperthermia and 1H/1⁹F MRI monitoring of drug release.

This simulation-based study presented a novel hybrid RF antenna array designed for neck cancer treatment within a 7T MRI system. The proposed design aimed to provide microwave hyperthermia to release 19F-labeled anticancer drugs from thermosensitive liposomes, facilitating drug concentration monitoring through 19F imaging and enabling 1H anatomical imaging and MR thermometry for temperature control. The design featured a bidirectional microstrip for generating the magnetic |B1|-fields required for 1H and 19F MR imaging, along with a patch antenna for localized RF heating. The bidirectional microstrip was operated at 300 MHz and 280 MHz through the placement of excitation ports at the ends of the antenna and an asymmetric structure along the antenna. Additionally, a patch antenna was positioned at the center. Based on this setup, an array of six antennas was designed. Simulation results using a tissue-mimicking simulation model confirmed the intensity and uniformity of |B1|-fields for both 19F and 1H nuclei, demonstrating the suitability of the design for clinical imaging. RF heating from the patch antennas was effectively localized at the center of the cancer model. In simulations with a human model, average |B1|-fields were 0.21 μT for 19F and 0.12 μT for 1H, with normalized-absolute-average-deviation values of 81.75% and 87.74%, respectively. Hyperthermia treatment was applied at 120 W for 600 s, achieving an average temperature of 40.22°C in the cancer model with a perfusion rate of 1 ml/min/kg. This study demonstrated the potential of a hybrid antenna array for integrating 1H MR, 19F drug monitoring, and hyperthermia.

Linsitinib, an IGF-1R inhibitor, attenuates disease development and progression in a model of thyroid eye disease.

Graves' disease (GD) is an autoimmune disorder caused by autoantibodies against the thyroid stimulating hormone receptor (TSHR) leading to overstimulation of the thyroid gland. Thyroid eye disease (TED) is the most common extra thyroidal manifestation of GD. Therapeutic options to treat TED are very limited and novel treatments need to be developed. In the present study we investigated the effect of linsitinib, a dual small-molecule kinase inhibitor of the insulin-like growth factor 1 receptor (IGF-1R) and the Insulin receptor (IR) on the disease outcome of GD and TED. Linsitinib was administered orally for four weeks with therapy initiating in either the early ("active") or the late ("chronic") phases of the disease. In the thyroid and the orbit, autoimmune hyperthyroidism and orbitopathy were analyzed serologically (total anti-TSHR binding antibodies, stimulating anti TSHR antibodies, total T4 levels), immunohistochemically (H&E-, CD3-, TNFa- and Sirius red staining) and with immunofluorescence (F4/80 staining). An MRI was performed to quantify in vivo tissue remodeling inside the orbit. Linsitinib prevented autoimmune hyperthyroidism in the early state of the disease, by reducing morphological changes indicative for hyperthyroidism and blocking T-cell infiltration, visualized by CD3 staining. In the late state of the disease linsitinib had its main effect in the orbit. Linsitinib reduced immune infiltration of T-cells (CD3 staining) and macrophages (F4/80 and TNFa staining) in the orbita in experimental GD suggesting an additional, direct effect of linsitinib on the autoimmune response. In addition, treatment with linsitinib normalized the amount of brown adipose tissue in both the early and late group. An in vivo MRI of the late group was performed and revealed a marked decrease of inflammation, visualized by 19F MR imaging, significant reduction of existing muscle edema and formation of brown adipose tissue. Here, we demonstrate that linsitinib effectively prevents development and progression of thyroid eye disease in an experimental murine model for Graves' disease. Linsitinib improved the total disease outcome, indicating the clinical significance of the findings and providing a path to therapeutic intervention of Graves' Disease. Our data support the use of linsitinib as a novel treatment for thyroid eye disease.

First in vivo fluorine-19 magnetic resonance imaging of the multiple sclerosis drug siponimod.

Theranostic imaging methods could greatly enhance our understanding of the distribution of CNS-acting drugs in individual patients. Fluorine-19 magnetic resonance imaging (19F MRI) offers the opportunity to localize and quantify fluorinated drugs non-invasively, without modifications and without the application of ionizing or other harmful radiation. Here we investigated siponimod, a sphingosine 1-phosphate (S1P) receptor antagonist indicated for secondary progressive multiple sclerosis (SPMS), to determine the feasibility of in vivo 19F MR imaging of a disease modifying drug. Methods: The 19F MR properties of siponimod were characterized using spectroscopic techniques. Four MRI methods were investigated to determine which was the most sensitive for 19F MR imaging of siponimod under biological conditions. We subsequently administered siponimod orally to 6 mice and acquired 19F MR spectra and images in vivo directly after administration, and in ex vivo tissues. Results: The 19F transverse relaxation time of siponimod was 381 ms when dissolved in dimethyl sulfoxide, and substantially reduced to 5 ms when combined with serum, and to 20 ms in ex vivo liver tissue. Ultrashort echo time (UTE) imaging was determined to be the most sensitive MRI technique for imaging siponimod in a biological context and was used to map the drug in vivo in the stomach and liver. Ex vivo images in the liver and brain showed an inhomogeneous distribution of siponimod in both organs. In the brain, siponimod accumulated predominantly in the cerebrum but not the cerebellum. No secondary 19F signals were detected from metabolites. From a translational perspective, we found that acquisitions done on a 3.0 T clinical MR scanner were 2.75 times more sensitive than acquisitions performed on a preclinical 9.4 T MR setup when taking changes in brain size across species into consideration and using equivalent relative spatial resolution. Conclusion: Siponimod can be imaged non-invasively using 19F UTE MRI in the form administered to MS patients, without modification. This study lays the groundwork for more extensive preclinical and clinical investigations. With the necessary technical development, 19F MRI has the potential to become a powerful theranostic tool for studying the time-course and distribution of CNS-acting drugs within the brain, especially during pathology.

Pivotal role of the protein corona in the cell uptake of fluorinated nanoparticles with increased sensitivity for 19F-MR imaging.

In vivo cell tracking by non-invasive imaging technologies is needed to accelerate the clinical translation of innovative cell-based therapies. In this regard, 19F-MRI has recently gained increased attention for unbiased localization of labeled cells over time. To push forward the use of 19F-MRI for cell tracking, the development of highly performant 19F-probes is required. PLGA-based NPs containing PERFECTA, a multibranched superfluorinated molecule with an optimal MRI profile thanks to its 36 magnetically equivalent fluorine atoms, are promising 19F-MRI probes. In this work we demonstrate the importance of the surface functionalization of these NPs in relation to their interaction with the biological environment, stressing the pivotal role of the formation of the protein corona (PC) in their cellular labelling efficacy. In particular, our studies showed that the formation of PC NPs strongly promotes the cellular internalization of these NPs in microglia cells. We advocate that the formation of PC NPs in the culture medium can be a key element to be used for the optimization of cell labelling with a considerable increase of the detection sensitivity by 19F-MRI.

Ventilation Study of the Human Lungs by 19F MRI at 0.5 Tesla

To show the feasibility of functional lung assessment by 19F MRI using low field (0.5 T) MRI scanner. One healthy volunteer participated in the studies. As a contrast for 19F pulmonary MRI, the gas mixture of 70% octafluorocyclobutane (OFCB) and 30% oxygen was used. 19F MR images of human lungs were obtained using 2D and 3D FSE methods. MRI data were used for volume reconstruction and for calculation of wash-in/-out and single-breath dynamics measurements. 19F 3D imaging provided information about gas distribution and lung volume assessment. The measured volume of the left and right parts of lungs were ≈1.7L and ≈1.8L, respectively. The wash-in/-out dynamics measurements determined that the effective time of gas washing in was 30 ± 5 s and washing out was 19 ± 4 s. Fractional ventilation was 29 ± 3% and 18 ± 2% for wash-in and wash-out processes, respectively. Dynamics of gas distribution during one breath cycle was analyzed. The calculated inspiration and expiration maps gave normalized effective times [rel. un.] for these stages- 0.95 ± 0.18 and 0.84 ± 0.15, respectively. Different 19F pulmonary MRI methods were implemented: 3D imaging, wash-in/-out dynamics and single respiratory cycle imaging. The results are agreed with known data and demonstrates possibility of ventilation assessment of the lungs at 0.5 Tesla.

LED-Based Photo-CIDNP Hyperpolarization Enables 19F MR Imaging and 19F NMR Spectroscopy of 3-Fluoro-DL-tyrosine at 0.6 T

Although 19F has high potential to serve as a background-free molecular marker in bioimaging, the molar amount of marker substance is often too small to enable 19F MR imaging or 19F NMR spectroscopy with a sufficiently high signal-to-noise ratio (SNR). Hyperpolarization methods such as parahydrogen-based hyperpolarization or dynamic nuclear polarization (DNP) can significantly improve the SNR, but require expensive and complex sample preparation and the removal of toxic catalysts and solvents. Therefore, we used the biologically compatible model of the fluorinated amino acid 3-Fluoro-DL-tyrosine with riboflavin 5ʹ-monophosphate (FMN) as a chromophore dissolved in D2O with 3.4% H2Odest., allowing to transform light energy into hyperpolarization of the 19F nucleus via photo-chemically induced dynamic nuclear polarization (photo-CIDNP). We used a low-cost high-power blue LED to illuminate the sample replacing traditionally used laser excitation, which is both potentially harmful and costly. For the first time, we present results of hyperpolarized 19F MRI and 19F NMR performed with a low-cost 0.6 T benchtop MRI system. The device allowed simultaneous dual-channel 1H/19F NMR. 19F imaging was performed with a (0.94 mm)2 in-plane resolution. This enabled the spatial resolution of different degrees of hyperpolarization within the sample. We estimated the photo-CIDNP-based 19F signal enhancement at 0.6 T to be approximately 465. FMN did not bleach out even after multiple excitations, so that the signal-to-noise ratio could be further improved by averaging hyperpolarized signals. The results show that the easy-to-use experimental setup has a high potential to serve as an efficient preclinical tool for hyperpolarization studies in bioimaging.

Design, Characterization and Molecular Modeling of New Fluorinated Paramagnetic Contrast Agents for Dual 1H/19F MRI

One major goal in medical imaging is the elaboration of more efficient contrast agents (CAs). Those agents need to be optimized for the detection of affected tissues such as cancers or tumors while decreasing the injected quantity of agents. The paramagnetic contrast agents containing fluorine atoms can be used for both proton and fluorine magnetic resonance imaging (MRI), and they open the possibility of simultaneously mapping the anatomy using 1H MRI and accurately locating the agents using 19F MRI. One of the challenges in this domain is to synthesize molecules containing several chemically equivalent fluorine atoms with relatively short relaxation times to allow the recording of 19F MR images in good conditions. With that aim, we propose to prepare a CA containing a paramagnetic center and nine chemically equivalent fluorine atoms using a cycloaddition reaction between two building blocks. These fluorinated contrast agents are characterized by 19F NMR, showing differences in the fluorine relaxation times T1 and T2 depending on the lanthanide ion. To complement the experimental results, molecular dynamics simulations are performed to shed light on the 3D-structure of the molecules in order to estimate the distance between the lanthanide ion and the fluorine atoms.

Supramolecular Fluorine Magnetic Resonance Spectroscopy Probe Polymer Based on Passerini Bifunctional Monomer.

A water-soluble fluorine magnetic resonance spectroscopy host-guest probe, P(HPA-co-AdamCF3A), was successfully constructed from the facile synthesis of a bifunctional monomer via a quantitative Passerini reaction. Supramolecular complexation with (2-hydroxypropyl)-β-cyclodextrin promoted a change in the chemical environment, leading to modulation of both the relaxation properties as well as chemical shift of the fluorine moieties. This change was used to probe the supramolecular interaction by 19F MRI spectroscopy and give insight into fluorine probe formulation. This work provides a fundamental basis for an 19F MR imaging tracer capable of assessing host-guest inclusion and a potential model to follow the fate of a drug delivery system in vivo.

Furin‐Controlled Fe3O4 Nanoparticle Aggregation and 19F Signal “Turn‐On” for Precise MR Imaging of Tumors

AbstractFor cancer diagnosis, 1H magnetic resonance imaging (MRI) is advantageous in sensitivity but lacks selectivity. Endogenous 19F MRI signal in humans is barely detectable and thus 19F MRI has very high selectivity. A combination of 1H and 19F MRI is ideal for precise tumor imaging but a protease‐controlled strategy of simultaneous T2 1H MRI enhancement and 19F MRI “Turn‐On” has not been reported. Here, used is a click condensation reaction to rationally project a dual‐functional fluorine probe 4‐(trifluoromethyl)benzoic acid (TFMB)‐Arg‐Val‐Arg‐Arg‐Cys(StBu)‐Lys‐CBT (1), which is further utilized to functionalize Fe3O4 nanoparticle (IONP) to achieve IONP@1. As such, the IONP aggregation can be activated by furin addition, thereby enhancing the T2 1H MRI signal and switching the 19F NMR/MRI signal “On”. Using this strategy, IONP@1 is successfully applied to detect the activity of the furin enzyme with “Turn‐On” 19F NMR/MRI and T2 1H MRI signals are enhanced. Moreover, IONP@1 is also applied for precise dual‐mode (1H and 19F) MR imaging of tumors in zebrafish under 14.1 T. The current approach, therefore, provides a feasible and robust means to reconcile the dilemma between selectivity and sensitivity of conventional MRI probes. More importantly, it is envisioned that, by substituting the TFMB moiety in 1 with a perfluorinated compound, this “smart” method could be of potential use for precise 1H MR and 19F MR imaging of tumor in mouse or in bigger rodents in near future.

A systematic optimization of 19F MR image acquisition to detect macrophage invasion into an ECM hydrogel implanted in the stroke-damaged brain

19F-MR imaging of perfluorocarbon (PFC)-labeled macrophages can provide a unique insight into their participation and spatio-temporal dynamics of inflammatory events, such as the biodegradation of an extracellular matrix (ECM) hydrogel implanted into a stroke cavity. To determine the most efficient acquisition strategy for 19F-MR imaging, five commonly used sequences were optimized using a design of experiment (DoE) approach and compared based on their signal-to-noise ratio (SNR). The fast imaging with steady-state precession (FISP) sequence produced the most efficient detection of a 19F signal followed by the rapid acquisition with relaxation enhancement (RARE) sequence. The multi-slice multi-echo (MSME), fast low angle shot (FLASH), and zero echo time (ZTE) sequences were significantly less efficient. Imaging parameters (matrix/voxel size; slice thickness, number of averages) determined the accuracy (i.e. trueness and precision) of object identification by reducing partial volume effects, as determined by analysis of the point spread function (PSF). A 96 × 96 matrix size (0.35 mm3) produced the lowest limit of detection (LOD) for RARE (2.85 mM PFPE; 119 mM 19F) and FISP (0.43 mM PFPE; 18.1 mM 19F), with an SNR of 2 as the detection threshold. Imaging of a brain phantom with PFC-labeled macrophages invading an ECM hydrogel further illustrated the impact of these parameter changes. The systematic optimization of sequence and imaging parameters provides the framework for an accurate visualization of 19F-labeled macrophage distribution and density in the brain. This will enhance our understanding of the contribution of periphery-derived macrophages in bioscaffold degradation and its role in brain tissue regeneration.

Importance of Thermally Induced Aggregation on 19F Magnetic Resonance Imaging of Perfluoropolyether-Based Comb-Shaped Poly(2-oxazoline)s.

An understanding of thermally induced aggregation and consequent 19F magnetic resonance imaging (MRI) performance is essential for improved design of thermoresponsive 19F MRI contrast agents. Herein we describe a series of novel thermoresponsive perfluoropolyether (PFPE)-based comb-shaped poly(2-oxazoline)s (POxs) with different side-chain structures (2-methyl- (MeOx), 2-ethyl- (EtOx), and 2-( n-propyl)-2-oxazoline (nPrOx)). The comb polymers were prepared through reversible addition-fragmentation chain transfer (RAFT) polymerization of the respective oligo(2-oxazoline)acrylates using a perfluoropolyether macro-RAFT agent. The fluoropolyether chain end drives aggregation of the polymers, with small aggregates forming at 300 K for both poly(OMeOx5A)9-PFPE and poly(OEtOx4A)9-PFPE. The aggregates decrease in size and display increases in 19F MRI intensity with temperature, and at 350 K the MeOx polymers are in the form of unimers in solution, similar to the oligoethylene glycol (OEG)-based PFPE polymer. Above the TCP of poly(OEtOx4A)9-PFPE, the polymer forms large aggregates, and the 19F MR imaging performance is degraded. Likewise, poly(OnPrOx4A)-PFPE is above the LCST at all temperatures studied (300-350 K), and so weak imaging intensity is obtained. This report of novel thermoresponsive POx-based PFPE polymers highlights the importance of understanding self-association of polymers in solution and provides important insights for the development of "smart" thermoresponsive 19F MRI contrast agents.

Multinuclear Applications on 0.5 T Magnetic Resonance Scanner

The article describes experiments on the detection of nuclear magnetic resonance (NMR) signals from nuclei other than protons on low field (0.5 T) clinical MR scanner. The possibility of obtaining NMR spectra and MR imaging (MRI) from nuclei with a high natural abundance (19F, 31P, 11B, 23Na), as well as isotope-enriched samples (13C and 2H), is presented. The possibilities of recording NMR spectra for low-sensitivity nuclei (13C, 29Si, 2H, 17O, 14N) are shown. Special attention is paid to the detection of NMR signal from perfluorocarbons (PFC). The examples of 19F MRI of PFC obtained in the in vivo study of both animals and human are presented. The 19F MRI of fluorinated gas, which can be used as a contrast agent for lung imaging, was also obtained. The opportunity of increasing 13C NMR signal from PFC by polarization transfer from fluorine nuclei is demonstrated. It was shown that 23Na can be detected in NMR spectroscopy and MRI of animal and human. The prospects of 17O NMR for technology applications are noticed.

Study of kinetics of 19F-MRI using a fluorinated imaging agent (19FIT) on a 3T clinical MRI system.

To use 19F imaging tracer (19FIT-27) to evaluate kinetics in major organs. Kinetics studies using proton MRI are difficult because of low concentration of 19FIT-27 protons relative to background water protons. Because there is no background source of 19F NMR in a biological body, 19F may be an ideal nucleus to directly trace 19FIT-27. However, there are several challenges for reliable 19F MR imaging and spectroscopy, particularly with clinical whole-body MRI systems, which include low concentrations and long 19F T1. We performed a dynamic 19F MRI study on mice at a 3T whole-body MRI system using a homemade transmit/receive (Tx/Rx) switch and a Tx/Rx volume RF coil. We used a newly developed fluorine imaging agent, which has 27 identical fluorine atoms with identical chemical shift, a relatively short T1, and high hydrophilicity. Basic kinetics parameters were estimated from the 19F signal-time curve. Resultant fluorine images show fairly high spatial (3 × 3 × 3mm3) and temporal resolutions. Biodistribution and kinetics of 19FIT-27 are obtained via 19F images for major uptake organs. Whole-body dynamic 19F MRI of newly developed 19FIT-27 in mice was obtained with fairly high spatial and temporal resolutions on a 3T clinical MRI system. The present study demonstrates the feasibility of 19F MRI using our newly developed compound to investigate major organ kinetics.

Elucidating the Impact of Molecular Structure on the 19F NMR Dynamics and MRI Performance of Fluorinated Oligomers.

To understand molecular factors that impact the performance of polymeric 19F magnetic resonance imaging (MRI) agents, a series of discrete fluorinated oligoacrylates with precisely defined structure were prepared through the combination of controlled polymerization and chromatographic separation techniques. These discrete oligomers enabled thorough elucidation of the dependence of 19F NMR and MRI properties on molecular structure, for example, the chain length. Importantly, the oligomer size and dispersity strongly influence NMR dynamics (T1 and T2 relaxation times) and MR imaging properties with higher signal-to-noise ratio (SNR) observed for oligomers with longer chain length and shorter T1. Our approach enables an effective pathway and thus opportunities to rationally design effective polymeric 19F MR imaging agents with optimized molecular structure and NMR relaxivity.

Low-cost LED-based Photo-CIDNP Enables Biocompatible Hyperpolarization of 19 F for NMR and MRI at 7 T and 4.7 T.

Substrates containing 19F can serve as background‐free reporter molecules for NMR and MRI. However, in vivo applications are still limited due to the lower signal‐to‐noise ratio (SNR) when compared with 1H NMR. Although hyperpolarization can increase the SNR, to date, only photo‐chemically induced dynamic nuclear polarization (photo‐CIDNP) allows for hyperpolarization without harmful metal catalysts. Photo‐CIDNP was shown to significantly enhance 19F NMR signals of 3‐fluoro‐DL‐tyrosine in aqueous solution using flavins as photosensitizers. However, lasers were used for photoexcitation, which is expensive and requires appropriate protection procedures in a medical or lab environment. Herein, we report 19F MR hyperpolarization at 4.7 T and 7 T with a biocompatible system using a low‐cost and easy‐to‐handle LED‐based set‐up. First hyperpolarized 19F MR images could be acquired, because photo‐CIDNP enabled repetitive hyperpolarization without adding new substrates.

Local Intratracheal Delivery of Perfluorocarbon Nanoparticles to Lung Cancer Demonstrated with Magnetic Resonance Multimodal Imaging.

Eighty percent of lung cancers originate as subtle premalignant changes in the airway mucosal epithelial layer of bronchi and alveoli, which evolve and penetrate deeper into the parenchyma. Liquid-ventilation, with perfluorocarbons (PFC) was first demonstrated in rodents in 1966 then subsequently applied as lipid-encapsulated PFC emulsions to improve pulmonary function in neonatal infants suffering with respiratory distress syndrome in 1996. Subsequently, PFC nanoparticles (NP) were extensively studied as intravenous (IV) vascular-constrained nanotechnologies for diagnostic imaging and targeted drug delivery applications.Methods: This proof-of-concept study compared intratumoral localization of fluorescent paramagnetic (M) PFC NP in the Vx2 rabbit model using proton (1H) and fluorine (19F) magnetic resonance (MR) imaging (3T) following intratracheal (IT) or IV administration. MRI results were corroborated by fluorescence microscopy.Results: Dynamic 1H-MR and 19F-MR images (3T) obtained over 72 h demonstrated marked and progressive accumulation of M-PFC NP within primary lung Vx2 tumors during the first 12 h post IT administration. Marked 1H and 19F MR signal persisted for over 72 h. In contradistinction, IV M-PFC NP produced a modest transient signal during the initial 2 h post-injection that was consistent circumferential blood pool tumor enhancement. Fluorescence microscopy of excised tumors corroborated the MR results and revealed enormous intratumor NP deposition on day 3 after IT but not IV treatment. Rhodamine-phospholipid incorporated into the PFC nanoparticle surfactant was distributed widely within the tumor on day 3, which is consistent with a hemifusion-based contact drug delivery mechanism previously reported. Fluorescence microscopy also revealed similar high concentrations of M-PFC NP given IT for metastatic Vx2 lung tumors. Biodistribution studies in mice revealed that M-PFC NP given IV distributed into the reticuloendothelial organs, whereas, the same dosage given IT was basically not detected beyond the lung itself. PFC NP given IT did not impact rabbit behavior or impair respiratory function. PFC NP effects on cells in culture were negligible and when given IV or IT no changes in rabbit hematology nor serum clinical chemistry parameters were measured.Conclusion: IT delivery of PFC NP offered unique opportunity to locally deliver PFC NP in high concentrations into lung cancers with minimal extratumor systemic exposure.

Clinically-Applicable Perfluorocarbon-Loaded Nanoparticles For In vivo Photoacoustic, 19F Magnetic Resonance And Fluorescent Imaging.

Photoacoustic imaging (PAI) is an emerging biomedical imaging technique that is now coming to the clinic. It has a penetration depth of a few centimeters and generates useful endogenous contrast, particularly from melanin and oxy-/deoxyhemoglobin. Indocyanine green (ICG) is a Food and Drug Administration-approved contrast agents for human applications, which can be also used in PAI. It is a small molecule dye with limited applications due to its fast clearance, rapid protein binding, and bleaching effect.Methods: Here, we entrap ICG in a poly(lactic-co-glycolic acid) nanoparticles together with a perfluorocarbon (PFC) using single emulsion method. These nanoparticles and nanoparticle-loaded dendritic cells were imaged with PA, 19F MR, and fluorescence imaging in vitro and in vivo.Results: We formulated particles with an average diameter of 200 nm. The encapsulation of ICG within nanoparticles decreased its photobleaching and increased the retention of the signal within cells, making it available for applications such as cell imaging. As little as 0.1x106 cells could be detected in vivo with PAI using automated spectral unmixing. Furthermore, we observed the accumulation of ICG signal in the lymph node after subcutaneous injection of nanoparticles.Conclusion: We show that we can label primary human dendritic cells with the nanoparticles and image them in vitro and in vivo, in a multimodal manner. This work demonstrates the potential of combining PAI and 19F MRI for cell imaging and lymph node detection using nanoparticles that are currently produced at GMP-grade for clinical use.