Cypate‐Conjugated Porous Upconversion Nanocomposites for Programmed Delivery of Heat Shock Protein 70 Small Interfering RNA for Gene Silencing and Photothermal Ablation

Synergistic therapy is an accepted method of enhancing the efficacy of cancer therapies. In this study, cypate‐conjugated porous NaLuF₄ doped with Yb³⁺, Er³⁺, and Gd³⁺ is synthesized and its potential for upconversion luminescence/magnetic resonance dual‐modality molecular imaging for guiding oncotherapy is tested. Loading cypate‐conjugated upconversion nanoparticles (UCNP‐cy) with small interfering RNA gene against heat shock protein 70 (UCNP‐cy‐siRNA) enhances the cell damage. UCNP‐cy‐siRNA exhibits remarkable antitumor efficacy in vivo as a result of the synergistic effects of gene silencing and photothermal therapy, with low drug dose and minimal side effects. This result thus provides an explicit strategy for developing next‐generation multifunctional nanoplatforms for multimodal imaging‐guided synergistic oncotherapy.     

Reductively Dissociable siRNA‐Polymer Hybrid Nanogels for Efficient Targeted Gene Silencing

A highly efficient approach for target‐specific gene silencing based on a reductively dissociable nanogel incorporating small interfering RNA (siRNA) crosslinked with linear polyethylenimine (LPEI) via disulfide bonds is presented. Thiol‐terminated siRNA at both 3′‐ends is electrostatically complexed with thiol‐grafted LPEI. The prepared siRNA/LPEI complex contains inter‐ and intramolecular linkages, generating a mutually crosslinked siRNA/LPEI nanogel (MCN) that exhibits excellent structural stability against the addition of heparin but is readily disintegrated to biologically active, monomeric siRNA upon exposure to reductive conditions. Accordingly, the highly condensed, stable MCN shows greatly enhanced cellular uptake and gene silencing efficiency compared to the siRNA/LPEI complexes without crosslinks or with only LPEI‐mediated crosslinks.     

Soft Nanostructured Films for Actuated Surface‐Based siRNA Delivery

Substrate‐mediated gene delivery is an emerging technology that enables spatial control of gene expression and localized delivery. This is of particular interest for siRNA where surface‐based release can greatly impact the fields of stem‐cell reprograming, wound healing, and medical device coatings in general. However, reports on the use of siRNA for substrate‐mediated delivery are scarce and have suffered from low efficiency. Here, an alternative strategy is reported by designing self‐assembled substrates that experience stimuli‐responsive topological transformations. Specifically, a methodology is established to promote the molecular organization of lipid films having 3D‐bicontinuous cubic, 2D‐inverted hexagonal, or 1D‐lamellar nanostructures encapsulating siRNA. In response to a compositional, temperature, or humidity stimulus, the nanostructures evolve from 1D‐lamellar or 2D‐hexagonal to 3D‐cubic resulting in efficient siRNA release to human cell cultures. Grazing incidence X‐ray diffraction reveals that film nanostructures are highly ordered and preferentially aligned. The results indicate that film structure substantially affects siRNA delivery, with the supported 3D‐bicontinuous cubic phase yielding the most effective reduction of gene expression. Subsequent studies suggest this enhanced performance arises due to the ability of this phase to cross cell membranes, particularly those of endocytic compartments. This work underpins that nanostructure tuning is decisive to the performance of therapeutic films.     

Binary Targeting of siRNA to Hematologic Cancer Cells In Vivo Using Layer‐by‐Layer Nanoparticles

Using siRNA therapeutics to treat hematologic malignancies has been unsuccessful because blood cancer cells exhibit remarkable resistance to standard transfection methods. Herein, the successful delivery of siRNA therapeutics with a dual‐targeted, layer‐by‐layer nanoparticle (LbL‐NP) is reported. The LbL‐NP protects siRNA from nucleases in the bloodstream by embedding it within polyelectrolyte layers that coat a polymeric core. The outermost layer consists of hyaluronic acid (a CD44‐ligand) covalently conjugated to CD20 antibodies. The CD20/CD44 dual‐targeting outer layer provides precise binding to blood cancer cells, followed by receptor‐mediated endocytosis of the LbL‐NP. This siRNA delivery platform is used to silence B‐cell lymphoma 2 (BCL‐2), a pro‐survival protein, in vitro and in vivo. The dual‐targeting approach significantly enhances internalization of BCL‐2 siRNA in lymphoma and leukemia cells, which leads to significant downregulation of BCL‐2 expression. Systemic administration of the dual‐targeted, siRNA‐loaded nanoparticle induces apoptosis and hampers proliferation of blood cancer cells, both in cell culture and in orthotopic non‐Hodgkin's lymphoma animal models. These results provide the basis for approaches to targeting blood‐borne cancers and other diseases and suggest that LbL nanoassemblies are a promising approach for delivering therapeutic siRNA to hematopoetic cell types that are known to evade transfection by other means.     

Multifunctional Glyco‐Nanofibers: siRNA Induced Supermolecular Assembly for Codelivery In Vivo

Targeted codelivery and controlled release of drug/siRNA (small interfering RNA) in a safe and effective vehicle hold great promises for overcoming drug resistance and optimal efficacy in cancer treatment; however, rational design and preparation of such vehicles remain a critical challenge. Thus, glyco‐nanofibers (GNFs) are fabricated via supermolecular assembly of polyanionic siRNA and cationic vesicles to simultaneously deliver siRNA and doxorubicin hydrochloride (DOX) in vitro and in vivo. The vesicles are created through self‐assembly of a positively charged amphiphilic lactose derivative featuring a lactose moiety and a ferrocenium unit on either end of the molecule. The GNFs display excellent biocompatibility, enhanced cell‐penetrating ability, and hepatoma targetability. The high transport efficiency of siRNA, effective gene silencing ability, and enhanced cytotoxicity to HepG2 cells of GNFs loaded with DOX are observed in vitro. Furthermore, in vivo experiments show reduced systemic toxicity and enhanced therapeutic efficacy of DOX to both HepG2 and HepG2/ADR subcutaneous tumor‐bearing nude mice. This work proves the electrostatic self‐assembly between cationic carbohydrates and polyanionic siRNA to be a convenient and effective strategy to fabricate a single vehicle for safe and effective codelivery of drug/siRNA, which can be used to combine chemo‐ and gene‐therapy against cancers and other diseases.     

Single Molecule with Dual Function on Nanogold: Biofunctionalized Construct for In Vivo Photoacoustic Imaging and SERS Biosensing

Multimodal imaging provides complimentary information that is advantageous in studying both cellular and molecular mechanisms in vivo, which has tremendous potential in pre‐clinical research and clinical translational imaging. It is desirable to design probes for multimodal imaging that can be administered minimally but provides multifaceted information. Herein, we demonstrate the complementary dual functional ability of a nanoconstruct for molecular imaging in both photoacoustic (PA) and surface‐enhanced Raman scattering (SERS) biosensing simultaneously in tandem. To realize this, a group of NIR active organic molecules are designed and synthesized that possess both SERS and PA activity. Nanoconstructs realized by anchoring such molecules onto gold nanoparticles are demonstrated for targeting cancer biomarkers in vivo while providing complimentary information about biodistribution and targeting efficiency. In future, such nanoconstructs could play a major role in identifying surgical margins and also for disease monitoring in translational medicine.     

Engineered Proteinticles for Targeted Delivery of siRNA to Cancer Cells

Considering the problems of small interfering RNA (siRNA) delivery using traditional viral and nonviral vehicles, a new siRNA delivery system to enhance efficiency and safety needs to be developed. Here human ferritin‐based proteinticles are genetically engineered to simultaneously display various functional peptides on the surface of proteinticles: cationic peptide to capture siRNA, tumor cell targeting and penetrating peptides, and enzymatically cleaved peptide to release siRNA inside tumor cell. In the in vitro treatment of poly‐siRNA‐proteinticle complex, both of the tumor cell targeting and penetrating peptides are important for efficient delivery of siRNA, and the red fluorescent protein (RFP) expression in RFP‐expressing tumor cells is notably suppressed by the delivered siRNA with the complementary sequence to RFP mRNA. It seems that the human ferritin‐based proteinticle is an efficient, stable, and safe tool for siRNA delivery, having a great potential for application to in vivo cancer treatment. The unique feature of proteinticles is that multiple and functional peptides can be simultaneously and evenly placed and also easily switched on the proteinticle surface through a simple genetic modification, which is likely to make proteinticles appropriate for targeted delivery of siRNA to a wide range of cancer cells.     

Effective PEGylation of Iron Oxide Nanoparticles for High Performance In Vivo Cancer Imaging

A practical and effective strategy for synthesizing PEGylated superparamagnetic iron oxide nanoparticles (SPIONs) is established. In this strategy, poly(acrylic acid) (PAA) is combined with SPIONs via multiple coordination between the carboxylic groups of PAA and SPIONs, which introduces abundant carboxylic groups, then, α,ω‐diamino PEG is linked to SPIONs via the amidation of the carboxylic groups. The synthesized PEGylated SPIONs exhibit no cytotoxicity and high resistance to phagocytosis by macrophages in vitro as well as low uptake by the liver and spleen in vivo, which makes the SPIONs highly efficient in tumor imaging by magnetic resonance imaging (MRI) at a relatively low dose of SPIONs. These outstanding properties are largely due to the significant shielding effect of the dense PEG coating as well as the net neutral surface of the PEGylated SPIONs in physiological conditions. In summary, the PEGylated SPIONs prepared by this strategy exhibit great application potential in tumor imaging as MRI contrast agents targeting through enhanced permeability and retention (EPR) effect.     

Biocompatible,Uniform, and Redispersible Mesoporous Silica Nanoparticles for Cancer‐Targeted Drug Delivery In Vivo

Engineering multifunctional nanocarriers for targeted drug delivery shows promising potentials to revolutionize the cancer chemotherapy. Simple methods to optimize physicochemical characteristics and surface composition of the drug nanocarriers need to be developed in order to tackle major challenges for smooth translation of suitable nanocarriers to clinical applications. Here, rational development and utilization of multifunctional mesoporous silica nanoparticles (MSNPs) for targeting MDA‐MB‐231 xenograft model breast cancer in vivo are reported. Uniform and redispersible poly(ethylene glycol)‐incorporated MSNPs with three different sizes (48, 72, 100 nm) are synthesized. They are then functionalized with amino‐β‐cyclodextrin bridged by cleavable disulfide bonds, where amino‐β‐cyclodextrin blocks drugs inside the mesopores. The incorporation of active folate targeting ligand onto 48 nm of multifunctional MSNPs (PEG‐MSNPs48‐CD‐PEG‐FA) leads to improved and selective uptake of the nanoparticles into tumor. Targeted drug delivery capability of PEG‐MSNPs48‐CD‐PEG‐FA is demonstrated by significant inhibition of the tumor growth in mice treated with doxorubicin‐loaded nanoparticles, where doxorubicin is released triggered by intracellular acidic pH and glutathione. Doxorubicin‐loaded PEG‐MSNPs48‐CD‐PEG‐FA exhibits better in vivo therapeutic efficacy as compared with free doxorubicin and non‐targeted nanoparticles. Current study presents successful utilization of multifunctional MSNP‐based drug nanocarriers for targeted cancer therapy in vivo.     

PEGylated Micelle Nanoparticles Encapsulating a Non‐Fluorescent Near‐Infrared Organic Dye as a Safe and Highly‐Effective Photothermal Agent for In Vivo Cancer Therapy

Photothermal therapy (PTT), as a minimally invasive and highly effective cancer treatment approach, has received widespread attention in recent years. Tremendous effort has been devoted to explore various types of photothermal agents with high near‐infrared (NIR) absorbance for PTT cancer treatment. Despite many exciting progresses in the area, effective yet safe photothermal agents with good biocompatibility and biodegradability are still highly desired. In this work, a new organic PTT agent based on polyethylene glycol (PEG) coated micelle nanoparticles encapsulating a heptamethine indocyanine dye IR825 is developed, showing a strong NIR absorption band and a rather low quantum yield, for in vivo photothermal treatment of cancer. It is found that the IR825–PEG nanoparticles show ultra‐high in vivo tumor uptake after intravenous injection, and appear to be an excellent PTT agent for tumor ablation under a low‐power laser irradiation, without rendering any appreciable toxicity to the treated animals. Compared with inorganic nanomaterials and conjugated polymers being explored in PTT, the NIR‐absorbing micelle nanoparticles presented here may have the least safety concern while showing excellent treatment efficacy, and thus may be a new photothermal agent potentially useful in clinical applications.     

Bioengineered Extracellular Membranous Nanovesicles for Efficient Small‐Interfering RNA Delivery: Versatile Platforms for Stem Cell Engineering and In Vivo Delivery

Naturally derived nanovesicles secreted from various cell types and found in body fluids can provide effective platforms for the delivery of various cargoes because of their intrinsic ability to be internalized for intercellular signal transmission and membrane recycling. In this study, the versatility of bioengineered extracellular membranous nanovesicles as potent carriers of small‐interfering RNAs (siRNAs) for stem cell engineering and in vivo delivery has been explored. Here, exosomes have been engineered, one of the cell‐derived vesicle types, to overexpress exosomal proteins fused with cell‐adhesion or cell‐penetrating peptides for enhanced intracellular gene transfer. To devise a more effective delivery system with potential for mass production, a new siRNA delivery system has also been developed by artificially inducing the outward budding of plasma membrane nanovesicles. Those nanovesicles have been engineered by overexpressing E‐cadherin to facilitate siRNA delivery to human stem cells with resistance to intracellular gene transfer. Both types of engineered nanovesicles deliver siRNAs to human stem cells for lineage specification with negligible cytotoxicity. The nanovesicles are efficient in delivering siRNA in vivo, suggesting feasibility for gene therapy. Cell‐derived, bioengineered nanovesicles used for siRNA delivery can provide functional platforms enabling effective stem cell therapeutics and in vivo gene therapy.     

Properties of Native High‐Density Lipoproteins Inspire Synthesis of Actively Targeted In Vivo siRNA Delivery Vehicles

Efficient systemic administration of therapeutic short interfering RNA (siRNA) is challenging. High‐density lipoproteins (HDLs) are natural in vivo RNA delivery vehicles. Specifically, native HDLs: 1) load single‐stranded RNA; 2) are anionic, which requires charge reconciliation between the RNA and HDL, and 3) actively target scavenger receptor type B‐1 (SR‐B1) to deliver RNA. Emphasizing these particular parameters, templated lipoprotein particles (TLP), mimics of spherical HDLs, are employed and are self‐assembled with single‐stranded complements of, presumably, any highly unmodified siRNA duplex pair after formulation with a cationic lipid. Resulting siRNA templated lipoprotein particles (siRNA‐TLP) are anionic and tunable with regard to RNA assembly and function. Data demonstrate that the siRNA‐TLPs actively target SR‐B1 to potently reduce androgen receptor and enhancer of zeste homolog 2 proteins in multiple cancer cell lines. Systemic administration of siRNA‐TLPs demonstrated no off‐target toxicity and significantly reduced the growth of prostate cancer xenografts. Thus, native HDLs inspired the synthesis of a hybrid siRNA delivery vehicle that can modularly load single‐stranded RNA complements after charge reconciliation with a cationic lipid, and that function due to active targeting of SR‐B1.     

In Situ Generated Gold Nanoparticle Hybrid Polymersomes for Water‐Soluble Chemotherapeutics: Inhibited Leakage and pH‐Responsive Intracellular Release

Drug leakage in blood circulation is generally a serious concern to polymersomes when loading water‐soluble chemotherapeutics. If packing density of polymersome membrane is strengthened, premature drug release will be inhibited. Therefore, synthesis of a series of amphiphilic polyphosphazenes (PNPs) with 2‐diethylaminoethyl 4‐aminobenzoate (DEAB) as hydrophobic side groups and amino‐terminal poly(ethylene glycol) (NH₂‐PEG₂₀₀₀) as hydrophilic chains is presented. By controlling the ratio of DEAB to NH₂‐PEG₂₀₀₀, the optimal PNP‐3 is screened to ensure polymersome formation and high loading of doxorubicin hydrochloride (DOX·HCl). In situ generation method is initially employed to introduce gold nanoparticles (AuNPs) into vesicles' lamella, which can homogeneously distribute among DEAB sides via coordination interaction and act as inorganic cross‐linkers to aggregate polymer chains. Drug leakage of resultant AuNP hybrid PNP‐3 polymersome (IAuPNP‐3) at pH 7.4 is effectively alleviated and the systemic circulation time of DOX·HCl in mice is obviously prolonged. Besides, pH‐responsive drug release, due to the protonation of tertiary amine in DEAB, contributes to fast intracellular action. Based on the cooperation of these functions, DOX·HCl‐loaded IAuPNP‐3 finally achieves the highest in vivo antitumor efficacy compared with free DOX·HCl, drug‐loaded PNP, or EAuPNP prepared by prepreparation AuNPs method.     

Ribonucleoprotein: A Biomimetic Platform for Targeted siRNA Delivery

Many small interfering RNA (siRNA) carriers have been developed over the years, mostly based on cationic lipids and polymers that can condense siRNA into nanoparticle complexes and aid in endosomal escape. In comparison, development of charge‐neutral siRNA carriers to avoid or reduce nonspecific binding, aggregation, and toxicity is limited, due to a lack of mechanisms for carrier‐siRNA association beyond electrostatic interaction. Here, a unique, charge‐neutral, biomimetic platform is reported, mimicking the natural state of functional RNA, ribonucleoprotein (RNP). This RNP architecture is simple to make, precise in assembly stoichiometry, stable in serum, and biocompatible. Gene knockdown is achieved in vitro and in vivo, demonstrating excellent potential for translation.     

In Vivo Self‐Powered Wireless Transmission Using Biocompatible Flexible Energy Harvesters

Additional surgeries for implantable biomedical devices are inevitable to replace discharged batteries, but repeated surgeries can be a risk to patients, causing bleeding, inflammation, and infection. Therefore, developing self‐powered implantable devices is essential to reduce the patient's physical/psychological pain and financial burden. Although wireless communication plays a critical role in implantable biomedical devices that contain the function of data transmitting, it has never been integrated with in vivo piezoelectric self‐powered system due to its high‐level power consumption (microwatt‐scale). Here, wireless communication, which is essential for a ubiquitous healthcare system, is successfully driven with in vivo energy harvesting enabled by high‐performance single‐crystalline (1 ? x )Pb(Mg_1/3Nb_2/3)O₃?(x )Pb(Zr,Ti)O₃ (PMN‐PZT). The PMN‐PZT energy harvester generates an open‐circuit voltage of 17.8 V and a short‐circuit current of 1.74 µA from porcine heartbeats, which are greater by a factor of 4.45 and 17.5 than those of previously reported in vivo piezoelectric energy harvesting. The energy harvester exhibits excellent biocompatibility, which implies the possibility for applying the device to biomedical applications.     

A Fluorinated Bola‐Amphiphilic Dendrimer for On‐Demand Delivery of siRNA,via Specific Response to Reactive Oxygen Species

Functional materials capable of responding to stimuli intrinsic to diseases are extremely important for specific drug delivery at the disease site. However, developing on‐demand stimulus‐responsive vectors for targeted delivery is highly challenging. Here, a stimulus‐responsive fluorinated bola‐amphiphilic dendrimer is reported for on‐demand delivery of small interfering RNA (siRNA) in response to the characteristic high level of reactive oxygen species (ROS) in cancer cells. This dendrimer bears a ROS‐sensitive thioacetal in the hydrophobic core and positively charged poly(amidoamine) dendrons at the terminals, capable of interacting and compacting the negatively charged siRNA into nanoparticles to protect the siRNA and promote cellular uptake. The ROS‐sensitive feature of this dendrimer boosts specific and efficient disassembly of the siRNA/vector complexes in ROS‐rich cancer cells for effective siRNA delivery and gene silencing. Moreover, the fluorine tags in the vector enable ¹⁹F‐NMR analysis of the ROS‐responsive delivery process. In addition, this ingenious and distinct bola‐amphiphilic dendrimer is also able to combine the advantageous delivery features of both lipid and dendrimer vectors. Therefore, it represents an innovative on‐demand stimulus‐responsive delivery platform.     

Bioresorbable Wireless Sensors as Temporary Implants for In Vivo Measurements of Pressure

Pressures at targeted locations inside the human body serve as critically important diagnostic parameters for monitoring various types of serious or even potentially fatal medical conditions including intracranial, intra‐abdominal, and pulmonary hypertension, as well as compartment syndromes. Implantable commercial sensors provide satisfactory accuracy and stability in measurements of pressure, yet surgical removal is required after recovery of the patient to avoid infections and other risks associated with long‐term implantation. Sensors that dissolve in biofluids (or, equivalently, bioabsorb or bioresorb) avoid the need for such surgeries, yet current designs involve either hard‐wired connections and/or fail to provide quantitative measurements over clinically relevant lifetimes. Here, a bioresorbable, wireless pressure sensor based on passive inductor‐capacitor resonance circuits in layouts and with sets of materials that overcome these drawbacks is reported. Specifically, optimized designs offer sensitivity as high as ≈200 kHz mmHg^?1 and resolution as low as 1 mmHg. Encapsulation approaches that use membranes of Si₃N₄ and edge seals of natural wax support stable operation in vivo for up to 4 days. The bioresorbable pressure sensing technology reported here may serve as an important solution to temporary, real‐time monitoring of internal pressure for various medical conditions.