Multimodal Precision Imaging of Pulmonary Nanoparticle Delivery in Mice: Dynamics of Application,Spatial Distribution,and Dosimetry

Targeted delivery of nanomedicine/nanoparticles (NM/NPs) to the site of disease (e.g., the tumor or lung injury) is of vital importance for improved therapeutic efficacy. Multimodal imaging platforms provide powerful tools for monitoring delivery and tissue distribution of drugs and NM/NPs. This study introduces a preclinical imaging platform combining X‐ray (two modes) and fluorescence imaging (three modes) techniques for time‐resolved in vivo and spatially resolved ex vivo visualization of mouse lungs during pulmonary NP delivery. Liquid mixtures of iodine (contrast agent for X‐ray) and/or (nano)particles (X‐ray absorbing and/or fluorescent) are delivered to different regions of the lung via intratracheal instillation, nasal aspiration, and ventilator‐assisted aerosol inhalation. It is demonstrated that in vivo propagation‐based phase‐contrast X‐ray imaging elucidates the dynamic process of pulmonary NP delivery, while ex vivo fluorescence imaging (e.g., tissue‐cleared light sheet fluorescence microscopy) reveals the quantitative 3D drug/particle distribution throughout the entire lung with cellular resolution. The novel and complementary information from this imaging platform unveils the dynamics and mechanisms of pulmonary NM/NP delivery and deposition for each of the delivery routes, which provides guidance on optimizing pulmonary delivery techniques and novel‐designed NM for targeting and efficacy.     

Materials Chemistry of Nanoultrasonic Biomedicine

As a special cross‐disciplinary research frontier, nanoultrasonic biomedicine refers to the design and synthesis of nanomaterials to solve some critical issues of ultrasound (US)‐based biomedicine. The concept of nanoultrasonic biomedicine can also overcome the drawbacks of traditional microbubbles and promote the generation of novel US‐based contrast agents or synergistic agents for US theranostics. Here, we discuss the recent developments of material chemistry in advancing the nanoultrasonic biomedicine for diverse US‐based bio‐applications. We initially introduce the design principles of novel nanoplatforms for serving the nanoultrasonic biomedicine, from the viewpoint of synthetic material chemistry. Based on these principles and diverse US‐based bio‐application backgrounds, the representative proof‐of‐concept paradigms on this topic are clarified in detail, including nanodroplet vaporization for intelligent/responsive US imaging, multifunctional nano‐contrast agents for US‐based multi‐modality imaging, activatable synergistic agents for US‐based therapy, US‐triggered on‐demand drug releasing, US‐enhanced gene transfection, US‐based synergistic therapy on combating the cancer and potential toxicity issue of screening various nanosystems suitable for nanoultrasonic biomedicine. It is highly expected that this novel nanoultrasonic biomedicine and corresponding high performance in US imaging and therapy can significantly promote the generation of new sub‐discipline of US‐based biomedicine by rationally integrating material chemistry and theranostic nanomedicine with clinical US‐based biomedicine.     

Linking proteins with anionic nanoparticles via protamine: ultrasmall protein-coupled probes for magnetic resonance imaging of apoptosis

Magnetic resonance imaging (MRI) of a target in vivo depends on the surface, size, and particle relaxivity of the target-specific nanoparticles for MRI. Here a new method for decorating very small iron oxide particles (VSOPs) with target-specific ligands is described. The method is based on the electrostatic attraction of the strongly positively charged peptide protamine to the anionic citrate shell of the electrostatically stabilized VSOPs. The protamine coat allows linkage chemistry and chimera technology to functionalize VSOPs or other negative charged surfaces with biologics. Annexin A5 (anxA5)-VSOP utilizing thiol chemistry was generated to couple biologically active anxA5 to VSOPs for in vivo MRI of apoptosis. Annexin A5-VSOP comprises five anxA5 molecules per iron oxide nanoparticle with a high R2 particle relaxivity of 180 000 mM(-1) s(-1) yet small hydrodynamic diameter of only 14.7+/-2.9 nm beneficial for in vivo MRI of extravascular targets.     

In Situ Bioorthogonal Metabolic Labeling for Fluorescence Imaging of Virus Infection In Vivo

Optical fluorescence imaging is an important strategy to explore the mechanism of virus–host interaction. However, current fluorescent tag labeling strategies often dampen viral infectivity. The present study explores an in situ fluorescent labeling strategy in order to preserve viral infectivity and precisely monitor viral infection in vivo. In contrast to pre‐labeling strategy, mice are first intranasally infected with azide‐modified H5N1 pseudotype virus (N₃‐H5N1p), followed by injection of dibenzocyclooctyl (DBCO)‐functionalized fluorescence 6 h later. The results show that DBCO dye directly conjugated to N₃‐H5N1p in lung tissues through in vivo bioorthogonal chemistry with high specificity and efficacy. More remarkably, in situ labeling rather than conventional prelabeling strategy effectively preserves viral infectivity and immunogenicity both in vitro and in vivo. Hence, in situ bioorthogonal viral labeling is a promising and reliable strategy for imaging and tracking viral infection in vivo.     

Molecular Cancer Imaging in the Second Near‐Infrared Window Using a Renal‐Excreted NIR‐II Fluorophore‐Peptide Probe

In vivo molecular imaging of tumors targeting a specific cancer cell marker is a promising strategy for cancer diagnosis and imaging guided surgery and therapy. While targeted imaging often relies on antibody‐modified probes, peptides can afford targeting probes with small sizes, high penetrating ability, and rapid excretion. Recently, in vivo fluorescence imaging in the second near‐infrared window (NIR‐II, 1000–1700 nm) shows promise in reaching sub‐centimeter depth with microscale resolution. Here, a novel peptide (named CP) conjugated NIR‐II fluorescent probe is reported for molecular tumor imaging targeting a tumor stem cell biomarker CD133. The click chemistry derived peptide‐dye (CP‐IRT dye) probe afforded efficient in vivo tumor targeting in mice with a high tumor‐to‐normal tissue signal ratio (T/NT > 8). Importantly, the CP‐IRT probes are rapidly renal excreted (≈87% excretion within 6 h), in stark contrast to accumulation in the liver for typical antibody‐dye probes. Further, with NIR‐II emitting CP‐IRT probes, urethra of mice can be imaged fluorescently for the first time noninvasively through intact tissue. The NIR‐II fluorescent, CD133 targeting imaging probes are potentially useful for human use in the clinic for cancer diagnosis and therapy.     

Multifunctional Magnetic Gd3+‐Based Coordination Polymer Nanoparticles: Combination of Magnetic Resonance and Multispectral Optoacoustic Detections for Tumor‐Targeted Imaging in vivo

To overcome traditional barriers in optical imaging and microscopy, optoacoustic‐imaging has been changed to combine the accuracy of spectroscopy with the depth resolution of ultrasound, achieving a novel modality with powerful in vivo imaging. However, magnetic resonance imaging provides better spatial and anatomical resolution. Thus, a single hybrid nanoprobe that allows for simultaneous multimodal imaging is significant not only for cutting edge research in imaging science, but also for accurate clinical diagnosis. A core‐shell‐structured coordination polymer composite microsphere has been designed for in vivo multimodality imaging. It consists of a Fe₃O₄ nanocluster core, a carbon sandwiched layer, and a carbocyanine‐Gd^III (Cy‐Gd^III) coordination polymer outer shell (Fe₃O₄@C@Cy‐Gd^III). Folic acid‐conjugated poly(ethylene glycol) chains are embedded within the coordination polymer shell to achieve extended circulation and targeted delivery of probe particles in vivo. Control of Fe₃O₄ core grain sizes results in optimal r₂ relaxivity (224.5 × 10^–3 m ⁻¹ s^‐1) for T₂‐weighted magnetic resonance imaging. Cy‐Gd^III coordination polymers are also regulated to obtain a maximum 25.1% of Cy ligands and 5.2% of Gd^III ions for near‐infrared fluorescence and T₁‐weighted magnetic resonance imaging, respectively. The results demonstrate their impressive abilities for targeted, multimodal, and reliable imaging.     

Advanced Photoacoustic Imaging Applications of Near‐Infrared Absorbing Organic Nanoparticles

Progress of nanotechnology in recent years has stimulated fast development of nanoparticles in biomedical research. Photoacoustic (PA) imaging as an emerging non‐invasive technique in molecular imaging has improved imaging depth relative to conventional optical imaging, demonstrating great potential in clinical applications. The convergence of nanotechnology and PA imaging has enabled a broad spectrum of new opportunities in fundamental biology and translation medicine. This review focuses on the recent advances of organic nanoparticles in PA imaging applications. Near‐infrared absorbing organic nanoparticles are classified and discussed according to their different imaging applications, which include tumor imaging, gastrointestinal imaging, sentinel lymph node imaging, disease microenvironment imaging and real‐time drug imaging. The chemistry and PA properties of organic nanoparticles are discussed in details to highlight their own merits, and their challenges and perspectives in PA imaging are also discussed.     

Activatable Semiconducting Theranostics: Simultaneous Generation and Ratiometric Photoacoustic Imaging of Reactive Oxygen Species In Vivo

Enhancing the generation of reactive oxygen species (ROS) is an effective anticancer strategy. However, it is a great challenge to control the production and to image ROS in vivo, both of which are vital for improving the efficacy and accuracy of cancer therapy. Herein, an activatable semiconducting theranostic nanoparticle (NP) platform is developed that can simultaneously enhance ROS generation while self‐monitoring its levels through ratiometric photoacoustic (PA) imaging. The NP platform can further guide in vivo therapeutic effect in tumors. The theranostic NP platform is composed of: (i) cisplatin prodrug and ferric ion catalyst for ROS generation, a part of combination cancer therapy; and (ii) a ratiometric PA imaging nanoprobe consisting of inert semiconducting perylene‐diimide (PDI) and ROS activatable near‐infrared dye (IR790s), used in ratiometric PA imaging of ROS during cancer treatment. Ratiometric PA signals are measured at two near‐infrared excitation wavelengths: 680 and 790 nm for PDI and IR790s, respectively. The measurements show highly accurate visualization of ^?OH generation in vivo. This novel ROS responsive organic theranostic NP allows not only synergistic cancer chemotherapy but also real‐time monitoring of the therapeutic effect through ratiometric PA imaging.     

Targetable gold nanorods for epithelial cancer therapy guided by near-IR absorption imaging

Well-designed nanoparticle-mediated, image-guided cancer therapy has attracted interest for increasing the efficacy of cancer treatment. A new class of smart theragnostic nanoprobes employing cetuximab (CET)-conjugated polyethylene glycol (PEG)ylated gold nanorods (CET-PGNRs) is presented; these nanoprobes target epithelial cancer cells using near-infrared light. The cetyltrimethylammonium bromide bilayer on GNRs is replaced with heterobifunctional PEG (COOH-PEG-SH) to serve as a biocompatible stabilizer and to increase specificity. The carboxylated GNRs are further functionalized with CET using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC-NHS) chemistry. To assess the potential of such GNRs, their optical properties, biocompatibility, colloidal stability, in vitro/in vivo binding affinities for cancer cells, absorption imaging, and photothermal therapy effects are investigated. CET-PGNRs exhibit excellent tumor targeting ability and strong potential for simultaneous absorption imaging and photothermal ablation of epithelial cancer cells.     

Nanoagents Based on Poly(ethylene glycol)‐b‐Poly(l‐thyroxine) Block Copolypeptide for Enhanced Dual‐Modality Imaging and Targeted Tumor Radiotherapy

Future healthcare requires development of novel theranostic agents that are capable of not only enhancing diagnosis and monitoring therapeutic responses but also augmenting therapeutic outcomes. Here, a versatile and stable nanoagent is reported based on poly(ethylene glycol)‐b‐poly(l ‐thyroxine) (PEG‐PThy) block copolypeptide for enhanced single photon emission computed tomography/computed tomography (SPECT/CT) dual‐modality imaging and targeted tumor radiotherapy in vivo. PEG‐PThy acquired by polymerization of l ‐thyroxine‐N‐carboxyanhydride (Thy‐NCA) displays a controlled M_n, high iodine content of ≈49.2 wt%, and can spontaneously form 65 nm‐sized nanoparticles (PThyN). In contrast to clinically used contrast agents like iohexol and iodixanol, PThyN reveals iso‐osmolality, low viscosity, and long circulation time. While PThyN exhibits comparable in vitro CT attenuation efficacy to iohexol, it greatly enhances in vivo CT imaging of vascular systems and soft tissues. PThyN allows for surface decoration with the cRGD peptide achieving enhanced CT imaging of subcutaneous B16F10 melanoma and orthotopic A549 lung tumor. Taking advantages of a facile iodine exchange reaction, ¹²⁵I‐labeled PThyN enables SPECT/CT imaging of tumors and monitoring of PThyN biodistribution in vivo. Besides, ¹³¹I‐labeled and cRGD‐functionalized PThyN displays remarkable growth inhibition of the B16F10 tumor in mice (tumor inhibition rate > 89%). These poly(l ‐thyroxine) nanoparticles provide a unique and versatile theranostic platform for varying diseases.     

Gold Nanoshelled Liquid Perfluorocarbon Nanocapsules for Combined Dual Modal Ultrasound/CT Imaging and Photothermal Therapy of Cancer

The integration of multimodal contrast‐enhanced diagnostic imaging and therapeutic capabilities could utilize imaging guided therapy to plan the treatment strategy based on the diagnostic results and to guide/monitor the therapeutic procedures. Herein, gold nanoshelled perfluorooctylbromide (PFOB) nanocapsules with PEGylation (PGsP NCs) are constructed by oil‐in‐water emulsion method to form polymeric PFOB nanocapsules, followed by the formation of PEGylated gold nanoshell on the surface. PGsP NCs could not only provide excellent contrast enhancement for dual modal ultrasound and CT imaging in vitro and in vivo, but also serve as efficient photoabsorbers for photothermal ablation of tumors on xenografted nude mouse model. To our best knowledge, this is the first report of gold nanoshell serving as both CT contrast agents and photoabsorbers for photothermal therapy. The novel multifunctional nanomedicine would be of great value to offer more comprehensive diagnostic information to guide more accurate and effective cancer therapy.     

Ultrasmall Near‐Infrared Non‐cadmium Quantum Dots for in vivo Tumor Imaging

The high tumor uptake of ultrasmall near‐infrared quantum dots (QDs) attributed to the enhanced permeability and retention effect is reported. InAs/InP/ZnSe QDs coated by mercaptopropionic acid (MPA) exhibit an emission wavelength of about 800 nm (QD800‐MPA) with very small hydrodynamic diameter (<10 nm). Using 22B and LS174T tumor xenograft models, in vivo and ex vivo imaging studies show that QD800‐MPA is highly accumulated in the tumor area, which is very promising for tumor detection in living mice. The ex vivo elemental analysis (Indium) using inductively coupled plasma (ICP) spectrometry confirm the tumor uptake of QDs. The ICP data are consistent with the in vivo and ex vivo fluorescence imaging. Human serum albumin (HSA)‐coated QD800‐MPA nanoparticles (QD800‐MPA‐HSA) show reduced localization in mononuclear phagocytic system‐related organs over QD800‐MPA plausibly due to the low uptake of QD800‐MPA‐HSA in macrophage cells. QD800‐MPA‐HSA may have great potential for in vivo fluorescence imaging.     

Smart Magnetic Fluorescent Nanoparticle Imaging Probes to Monitor MicroRNAs

An imaging system that can be used to evaluate the expression levels of microRNAs during neuronal development can provide noninvasive information for investigating a variety of biological phenomena related to microRNAs (miRNAs, miRs). Herein, the development of a novel imaging platform to monitor intracellular miR124a during neuronal differentiation is reported using rhodamine‐coated cobalt ferrite magnetic fluorescent (MF) nanoparticles linked to a quenching molecular system containing an miR124a binding sequence (MF‐miR124a beacon). During neuronal differentiation, in vitro fluorescence signals of the MF‐miR124a beacon are significantly increased under conditions where miR124a is highly expressed, and dramatically return to the original quenched fluorescence after anti‐miR124a treatment. In vivo fluorescence images show enhanced fluorescence signals in mice with P19 cells within a poly‐L ‐lactic acid scaffold after induction of neuronal differentiation. In addition, magnetic resonance (MR) images provide in vivo tracking of cells containing the MF‐miR124a beacon. These studies represent the first step toward the use of nanotechnological imaging of mature miRNA, and this technique could be used for cellular tracking with a MR imaging system as well as for simultaneous monitoring of the miRNA expression pattern in vivo.     

Targeting Peptide‐Based Probes for Molecular Imaging and Diagnosis

A series of novel peptide‐based molecular probes for different biomarkers is highlighted herein. These probes can provide targeted recognition with high affinity, high specificity, high penetration, and rapid excretion ability. These sensitive peptides can achieve rapid and specific detection when they are conjugated with imaging moieties or are formed into nanoprobes, which can be adapted for in vivo molecular imaging in targeted diagnosis and therapy.     

Precisely Defined Conjugated Oligoelectrolytes for Biosensing and Therapeutics

Conjugated oligoelectrolytes (COEs) are a relatively new class of synthetic organic molecules with, as of yet, untapped potential for use in organic optoelectronic devices and bioelectronic systems. COEs also offer a novel molecular approach to biosensing, bioimaging, and disease therapy. Substantial progress has been made in the past decade at the intersection of chemistry, materials science, and the biological sciences developing COEs and their polymer analogues, namely, conjugated polyelectrolytes (CPEs), into synthetic systems with biological and biomedical utility. CPEs have traditionally attracted more attention in arenas of sensing, imaging, and therapy. However, the precisely defined molecular structures and interactions of COEs offer potential key advantages over CPEs, including higher reliability and fluorescence quantum efficiency, larger diversity of subcellular targeting strategies, and improved selectivity to biomolecules. Here, the unique—and sometimes overlooked—properties of COEs are discussed and the noticeable progress in their use for biological sensing, imaging, and therapy is reviewed.     

Multifunctional core/satellite polydopamine@Nd<Superscript>3+</Superscript>-sensitized upconversion nanocomposite: A single 808 nm near-infrared light-triggered theranostic platform for <Emphasis Type="Italic">in vivo</Emphasis> imaging-guided photothermal therapy

Significant attenuation and overheating,caused by the absorption of the excitation band (980 nm) in water,are the major obstacles in the in vivo application of lanthanide-doped upconversion nanopartides (UCNPs).Therefore,appropriately-structured Nd3+-doped UCNPs with 808 nm excitation could be a promising alternative.Herein,we developed core-shell-shell structured Nd3+-sensitized UCNPs as imaging agents,and decorated them onto the surface of polydopamine (PDA) to construct a novel multifunctional core/satellite nanotheranostic (PDA@UCNPs) for in vivo imaging guidance photothermal therapy using single 808 nm laser irradiation.The core-shell-shell structured design enabled outstanding upconversion luminescence properties and strong X-ray attenuation,thereby making the nanocomposites potential candidates for excellent upconversion luminescence/computed tomography dual modal imaging.In addition,the PDA core not only provides high photothermal conversion efficiency and outstanding antitumor effect,but also endows the platform with robust biocompatibility owing to its natural features.Therefore,this multifunctional nanocomposite could be a promising theranostic in future oncotherapy,with high therapeutic effectiveness but low side effects.This study would stimulate interest in designing bioapplication-compatible multifunctional nanocomposites,especially for cancer diagnosis and treatment in vivo.     

Continuous Coaxial Electrohydrodynamic Atomization System for Water‐Stable Wrapping of Magnetic Nanoparticles

An electrohydrodynamic atomization (EHDA) system that generates an electrospray can achieve particle formation and encapsulation by accumulating an electric charge on liquid flowing out from the nozzle. A novel coaxial EHDA system for continuous fabrication of water‐stable magnetic nanoparticles (MNPs) is established, based on a cone‐jet mode of electrospraying. Systemic variables, such as flow rates from dual nozzles and inducing voltages, are controlled to enable the preparation of water‐soluble MNPs coated by polysorbate 80. The PEGylated MNPs exhibit water stability. The magnetic resonance imaging potential of these MNPs is confirmed by in vivo imaging using a gastric cancer xenograft mouse model. Thus, this advanced coaxial EHDA system demonstrates remarkable capabilities for the continuous encapsulation of MNPs to render them water‐stable while preserving their properties as imaging agents.     

Towards Single Biomolecule Imaging via Optical Nanoscale Magnetic Resonance Imaging

Nuclear magnetic resonance (NMR) spectroscopy is a physical marvel in which electromagnetic radiation is charged and discharged by nuclei in a magnetic field. In conventional NMR, the specific nuclei resonance frequency depends on the strength of the magnetic field and the magnetic properties of the isotope of the atoms. NMR is routinely utilized in clinical tests by converting nuclear spectroscopy in magnetic resonance imaging (MRI) and providing 3D, noninvasive biological imaging. While this technique has revolutionized biomedical science, measuring the magnetic resonance spectrum of single biomolecules is still an intangible aspiration, due to MRI resolution being limited to tens of micrometers. MRI and NMR have, however, recently greatly advanced, with many breakthroughs in nano‐NMR and nano‐MRI spurred by using spin sensors based on an atomic impurities in diamond. These techniques rely on magnetic dipole–dipole interactions rather than inductive detection. Here, novel nano‐MRI methods based on nitrogen vacancy centers in diamond are highlighted, that provide a solution to the imaging of single biomolecules with nanoscale resolution in‐vivo and in ambient conditions.     

Recent Advances in Nanotechnology for Autophagy Detection

Autophagy is closely related to various diseases, and is a diagnostic and therapeutic target for some diseases. In recent years, tremendous efforts have been made to develop excellent probes for detection of autophagy. Nanostructure‐based probes are interesting and promising approaches for in vivo biological imaging due to their unique structural and functional characteristics, e.g., modulating pharmacokinetics property by biocompatible coatings, multimodality capacity by delivering multiple imaging agents and highly specific targeting by antibody ligands. In this Review, we first introduce recent advancements in the development of nanostructure‐based probes for detection of autophagy, including inorganic hybrid nanomaterials and self‐assembled peptide polymeric nanoparticles. Meanwhile, a nanoprobe based on a “in vivo self‐assembly” strategy is highlighted. The “in vivo self‐assembly” endows nanoprobes with higher accumulation, and longer and better signal stability for in vivo detection of autophagy. Furthermore, this novel strategy could be widely used for biomedical imaging/diagnostics and therapeutics, which would attract more attention to this research area.     

Biofunctionalized,Phosphonate‐Grafted,Ultrasmall Iron Oxide Nanoparticles for Combined Targeted Cancer Therapy and Multimodal Imaging

A novel, inexpensive biofunctionalization approach is adopted to develop a multimodal and theranostic nanoagent, which combines cancer‐targeted magnetic resonance/optical imaging and pH‐sensitive drug release into one system. This multifunctional nanosystem, based on an ultrasmall superparamagnetic iron oxide (USPIO) nanocore, is modified with a hydrophilic, biocompatible, and biodegradable coating of N‐phosphonomethyl iminodiacetic acid (PMIDA). Using appropriate spacers, functional molecules, such as rhodamine B isothiocyanate, folic acid, and methotrexate, are coupled to the amine‐derivatized USPIO–PMIDA support with the aim of endowing simultaneous targeting, imaging, and intracellular drug‐delivering capability. For the first time, phosphonic acid chemistry is successfully exploited to develop a stealth, multifunctional nanoprobe that can selectively target, detect, and kill cancer cells overexpressing the folate receptor, while allowing real‐time monitoring of tumor response to drug treatment through dual‐modal fluorescence and magnetic resonance imaging.