Fabrication and characterization of PLGA nanoparticles encapsulating large CRISPR-Cas9 plasmid.

BACKGROUND: The clustered regularly interspaced short palindromic repeats (CRISPR) and Cas9 protein system is a revolutionary tool for gene therapy. Despite promising reports of the utility of CRISPR-Cas9 for in vivo gene editing, a principal problem in implementing this new process is delivery of high molecular weight DNA into cells. RESULTS: Using poly(lactic-co-glycolic acid) (PLGA), a nanoparticle carrier was designed to deliver a model CRISPR-Cas9 plasmid into primary bone marrow derived macrophages. The engineered PLGA-based carriers were approximately 160 nm and fluorescently labeled by encapsulation of the fluorophore 6,13-bis(triisopropylsilylethynyl) pentacene (TIPS pentacene). An amine-end capped PLGA encapsulated 1.6 wt% DNA, with an encapsulation efficiency of 80%. Release studies revealed that most of the DNA was released within the first 24 h and corresponded to ~ 2-3 plasmid copies released per nanoparticle. In vitro experiments conducted with murine bone marrow derived macrophages demonstrated that after 24 h of treatment with the PLGA-encapsulated CRISPR plasmids, the majority of cells were positive for TIPS pentacene and the protein Cas9 was detectable within the cells. CONCLUSIONS: In this work, plasmids for the CRISPR-Cas9 system were encapsulated in nanoparticles comprised of PLGA and were shown to induce expression of bacterial Cas9 in murine bone marrow derived macrophages in vitro. These results suggest that this nanoparticle-based plasmid delivery method can be effective for future in vivo applications of the CRISPR-Cas9 system.

Synthesis and Evaluation of Doxorubicin-Loaded Gold Nanoparticles for Tumor-Targeted Drug Delivery.

Doxorubicin is an effective and widely used cancer chemotherapeutic agent, but its application is greatly compromised by its cumulative dose-dependent side effect of cardiotoxicity. A gold nanoparticle-based drug delivery system has been designed to overcome this limitation. Five novel thiolated doxorubicin analogs were synthesized and their biological activities evaluated. Two of these analogs and PEG stabilizing ligands were then conjugated to gold nanoparticles, and the resulting Au-Dox constructs were evaluated. The results show that release of native drug can be achieved by the action of reducing agents such as glutathione or under acidic conditions, but reductive drug release gave the cleanest drug release. Gold nanoparticles (Au-Dox) were prepared with different loadings of PEG and doxorubicin, and one formulation was evaluated for mammalian stability and toxicity. Plasma levels of doxorubicin in mice treated with Au-Dox were significantly lower than in mice treated with the same amount of doxorubicin, indicating that the construct is stable under physiological conditions. Treatment of mice with Au-Dox gave no histopathologically observable differences from mice treated with saline, while mice treated with an equivalent dose of doxorubicin showed significant histopathologically observable lesions.

Design and Fabrication of Streptavidin-Functionalized, Fluorescently Labeled Polymeric Nanocarriers.

Targeted drug delivery has great potential for improving therapeutic outcomes for many diseases. Polymeric nanocarriers can improve the targeted delivery of insoluble and toxic drugs but, to achieve this, it is important to tailor the particle properties. In this study, nanoparticles comprised of poly(ethylene oxide)- b-poly(d,l-lactic acid) (PEO- b-PDLLA) were made by flash nanoprecipitation while varying the compositions of the additives poly(l-lactic acid) (PLLA), a fluorophore 6,13-bis(triisopropylsylylethynyl) (TIPS) pentacene, and poly(acrylic acid)- b-poly(d,l-lactic acid) (PAA- b-PDLLA) to characterize their effects on size, ζ potential, fluorescence, and surface functionalization. The particle size was readily increased by addition of PLLA homopolymer up to ∼40 wt % without significant change to the ζ potential. The maximum nanoparticle fluorescence was at 0.5 wt % TIPS based on the PDLLA core and exhibited quenching that could be described by Förster resonant energy transfer. The cores of the particles were coupled with streptavidin through 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide coupling chemistry. Even without the added carboxylate groups from the PAA, the base PEO- b-PDLLA nanoparticles were conjugated with streptavidin at comparable levels while retaining the functionality of streptavidin for further biotinylated ligand binding.

TIPS pentacene loaded PEO-PDLLA core-shell nanoparticles have similar cellular uptake dynamics in M1 and M2 macrophages and in corresponding in vivo microenvironments.

Nanoparticle based drug delivery platforms have the potential to transform disease treatment paradigms and therapeutic strategies, especially in the context of pulmonary medicine. Once administered, nanoparticles disperse throughout the lung and many are phagocytosed by macrophages. However, there is a paucity of knowledge regarding cellular up-take dynamics of nanoparticles due largely to macrophage heterogeneity. To address this issue, we sought to better define nanoparticle up-take using polarized M1 and M2 macrophages and novel TIPS-pentacene loaded PEO-PDLLA nanoparticles. Our data reveal that primary macrophages polarized to either M1 or M2 phenotypes have similar levels of nanoparticle phagocytosis. Similarly, M1 and M2 polarized macrophages isolated from the lungs of mice following either acute (Th1) or allergic (Th2) airway inflammation also demonstrated equivalent levels of nanoparticle up-take. Together, these studies provide critical benchmark information pertaining to cellular up-take dynamics and biodistribution of nanoparticles in the context of clinically relevant inflammatory microenvironments.

A New Interleukin-13 Amino-Coated Gadolinium Metallofullerene Nanoparticle for Targeted MRI Detection of Glioblastoma Tumor Cells.

The development of new nanoparticles as next-generation diagnostic and therapeutic ("theranostic") drug platforms is an active area of both chemistry and cancer research. Although numerous gadolinium (Gd) containing metallofullerenes as diagnostic magnetic resonance imaging (MRI) contrast agents have been reported, the metallofullerene cage surface, in most cases, consists of negatively charged carboxyl or hydroxyl groups that limit attractive forces with the cellular surface. It has been reported that nanoparticles with a positive charge will bind more efficiently to negatively charged phospholipid bilayer cellular surfaces, and will more readily undergo endocytosis. In this paper, we report the preparation of a new functionalized trimetallic nitride template endohedral metallofullerene (TNT EMF), Gd3N@C80(OH)x(NH2)y, with a cage surface bearing positively charged amino groups (-NH3(+)) and directly compare it with a similar carboxyl and hydroxyl functionalized derivative. This new nanoparticle was characterized by X-ray photoelectron spectroscopy (XPS), dynamic light scattering (DLS), and infrared spectroscopy. It exhibits excellent (1)H MR relaxivity. Previous studies have clearly demonstrated that the cytokine interleukin-13 (IL-13) effectively targets glioblastoma multiforme (GBM) cells, which are known to overexpress IL-13Rα2. We also report that this amino-coated Gd-nanoplatform, when subsequently conjugated with interleukin-13 peptide IL-13-Gd3N@C80(OH)x(NH2)y, exhibits enhanced targeting of U-251 GBM cell lines and can be effectively delivered intravenously in an orthotopic GBM mouse model.

Short-term patient-centered nutrition counseling impacts weight and nutrient intake in patients with systemic lupus erythematosus.

BACKGROUND: Patients with systemic lupus erythematosus (SLE) are at a high risk for cardiovascular disease (CVD) due to increased prevalence of traditional and nontraditional CVD risks factors. OBJECTIVE: To evaluate the effect of patient-centered nutrition counseling methods on changes in select nutrient, anthropometric, and clinical outcomes in SLE patients enrolled in a CVD prevention counseling program (CVD PCP). METHODS: From March 2009 to June 2011 a subgroup of SLE patients enrolled in our CVD PCP were referred to a registered dietitian for individualized nutrition counseling. Outcomes of interest were assessed at baseline and six months. A primary analysis evaluated the six-month changes in nutrient intake, weight, body mass index (BMI), waist circumference, and lipid levels. A secondary analysis compared the same measurements between the nutrition counseling patients and the group that was referred but did not attend. RESULTS: Of 71 referrals, 41 (58%) attended nutrition counseling (female: 89%, African American/Hispanic: 72%, mean age of 39.7 ± 12.82 years, and a mean disease duration of 11.49 ± 8.02 years). Over a six-month period, patients attending nutritional counseling: (a) reduced their intake of sodium (p = 0.006), total calories (p = 0.07), and percent calories from fat (p = 0.011) and saturated fat (p = 0.068); (b) had decreased weight (-1.64 kg, p = 0.025); and (c) were more likely to report increases in eating a diet rich in fruits and vegetables (p < 0.001), a high fiber diet (p = 0.011), ≥two servings of fish/week (p = 0.002), and a low cholesterol diet (p = 0.034). There were no significant changes observed over the six months in BMI and clinical outcomes among nutrition counseling patients. When comparing nutrition counseling patients to those who were referred but did not attend, we found at six months a higher percentage of nutrition counseling patients reportedly followed a high-fiber diet (p = 0.03), consumed two or more servings of fish per week (p = 0.01), followed a low-cholesterol diet (p = 0.03), and achieved a greater weight loss (p = 0.04) compared to the group that did not attend. CONCLUSION: At six months we found that nutrition counseling using patient-centered methods appears to be an effective method for promoting changes in nutrient intake, diet habits, and, possibly, anthropometric measures in SLE patients. However, the counseling did not show a significant improvement in lipid levels, possibly due to short follow-up and/or SLE related factors.

Gd3N@C84(OH)x: a new egg-shaped metallofullerene magnetic resonance imaging contrast agent.

Water-soluble derivatives of gadolinium-containing metallofullerenes have been considered to be excellent candidates for new magnetic resonance imaging (MRI) contrast agents because of their high relaxivity and characteristic encapsulation of the lanthanide ions (Gd(3+)), preventing their release into the bioenvironment. The trimetallic nitride template endohedral metallofullerenes (TNT EMFs) have further advantages of high stability, high relative yield, and encapsulation of three Gd(3+) ions per molecule as illustrated by the previously reported nearly spherical, Gd3N@I(h)-C80. In this study, we report the preparation and functionalization of a lower-symmetry EMF, Gd3N@C(s)-C84, with a pentalene (fused pentagons) motif and an egg-shaped structure. The Gd3N@C84 derivative exhibits a higher (1)H MR relaxivity compared to that of the Gd3N@C80 derivative synthesized the same way, at low (0.47 T), medium (1.4 T), and high (9.4 T) magnetic fields. The Gd3N@C(s)-C84 derivative exhibits a higher hydroxyl content and aggregate size, as confirmed by X-ray photoelectron spectroscopy (XPS) and dynamic light scattering (DLS) experiments, which could be the main reasons for the higher relaxivity.

Toward design of magnetic nanoparticle clusters stabilized by biocompatible diblock copolymers for T2-weighted MRI contrast.

We report the fabrication of magnetic particles comprised of clusters of iron oxide nanoparticles, 7.4 nm mean diameter, stabilized by a biocompatible, amphiphilic diblock copolymer, poly(ethylene oxide-b-D,L-lactide). Particles with quantitative incorporation of up to 40 wt % iron oxide and hydrodynamic sizes in the range of 80-170 nm were prepared. The particles consist of hydrophobically modified iron oxide nanoparticles within the core-forming polylactide block with the poly(ethylene oxide) forming a corona to afford aqueous dispersibility. The transverse relaxivities (r2) increased with average particle size and exceeded 200 s(-1) mM Fe(-1) at 1.4 T and 37 °C for iron oxide loadings above 30 wt %. These experimental relaxivities typically agreed to within 15% with the values predicted using analytical models of transverse relaxivity and cluster (particle core) size distributions derived from cryo-TEM measurements. Our results show that the theoretical models can be used for the rational design of biocompatible MRI contrast agents with tailored compositions and size distributions.

Improved microchip design and application for in situ transmission electron microscopy of macromolecules.

Understanding the fundamental properties of macromolecules has enhanced the development of emerging technologies used to improve biomedical research. Currently, there is a critical need for innovative platforms that can illuminate the function of biomedical reagents in a native environment. To address this need, we have developed an in situ approach to visualize the dynamic behavior of biomedically relevant macromolecules at the nanoscale. Newly designed silicon nitride devices containing integrated "microwells" were used to enclose active macromolecular specimens in liquid for transmission electron microscopy imaging purposes.We were able to successfully examine novel magnetic resonance imaging contrast reagents, micelle suspensions, liposome carrier vehicles, and transcribing viral assemblies. With each specimen tested, the integrated microwells adequately maintained macromolecules in discrete local environments while enabling thin liquid layers to be produced.

Interplay of degradation, dissolution and stabilization of clarithromycin and its amorphous solid dispersions.

Clarithromycin (CLA) is an aminomacrolide antibiotic whose physical properties are fascinating and challenging. It has very poor solubility at neutral intestinal pH, but much better solubility under acidic conditions due to amine protonation. The improved solubility in an acid environment is confounded by the poor chemical stability of clarithromycin that is quite labile toward acid-catalyzed degradation. This creates a complex system under gastrointestinal (GI) conditions: dissolution in the stomach, degradation, potential for precipitation in the small intestine, and interplay with the formulation components. We report herein a study of amorphous solid dispersion (ASD) of CLA with carboxyl-containing cellulose derivatives, which have recently been shown to be excellent ASD matrices for maximizing oral bioavailability. This approach was intended to improve CLA solubility in neutral media while minimizing release in an acid environment, and thereby increase its uptake from the small intestine. Amorphous polymer/CLA nanoparticles were also prepared by high-shear mixing in a multi-inlet vortex mixer (MIVM). Different extents of release were observed at low pH from the various formulations. Thus the solubility increase from nanosizing was deleterious to the concentration of intact CLA obtained upon reaching small intestine conditions; the high extent of release at gastric pH led to complete degradation of CLA. Using pH-switch experiments, it was possible to separate the effects of loss of CLA from solution by crystallization vs. that from chemical degradation. It was found that the hydrophobic cellulose derivative cellulose acetate adipate propionate (CAAdP) was effective at protecting CLA from dissolution in the stomach, and preventing CLA decomposition at low pH; 54% of CLA in CAADP ASD was released intact, vs. 0% and 6% from HPMCAS and CMCAB ASDs, respectively. We conclude that protection against degradation is central to enhancing overall release of intact CLA from ASD formulations; the formulations studied herein have great promise for simultaneous CLA solubility enhancement and protection from loss to chemical degradation, thereby reducing dose requirements and potentially decreasing colonic exposure to CLA (reduced colonic exposure is expected to minimize killing of beneficial colonic bacteria by CLA).

Encapsulation of PI3K Inhibitor LY294002 within Polymer Nanoparticles Using Ion Pairing Flash Nanoprecipitation.

Flash nanoprecipitation (FNP) is a turbulent mixing process capable of reproducibly producing polymer nanoparticles loaded with active pharmaceutical ingredients (APIs). The nanoparticles produced with this method consist of a hydrophobic core surrounded by a hydrophilic corona. FNP produces nanoparticles with very high loading levels of nonionic hydrophobic APIs. However, hydrophobic compounds with ionizable groups are not as efficiently incorporated. To overcome this, ion pairing agents (IPs) can be incorporated into the FNP formulation to produce highly hydrophobic drug salts that efficiently precipitate during mixing. We demonstrate the encapsulation of the PI3K inhibitor, LY294002, within poly(ethylene glycol)-b-poly(D,L lactic acid) nanoparticles. We investigated how incorporating two hydrophobic IPs (palmitic acid (PA) and hexadecylphosphonic acid (HDPA)) during the FNP process affected the LY294002 loading and size of the resulting nanoparticles. The effect of organic solvent choice on the synthesis process was also examined. While the presence of either hydrophobic IP effectively increased the encapsulation of LY294002 during FNP, HDPA resulted in well-defined colloidally stable particles, while the PA resulted in ill-defined aggregates. The incorporation of hydrophobic IPs with FNP opens the door for the intravenous administration of APIs that were previously deemed unusable due to their hydrophobic nature.

Probability of Cavitation in a Custom Iron-Based Coupling Medium for Transcranial Magnetic Resonance-Guided Focused Ultrasound Procedures.

OBJECTIVE: A coupling bath of circulating, chilled, degassed water is essential to safe and precise acoustic transmittance during transcranial magnetic resonance-guided focused ultrasound (tMRgFUS) procedures, but the circulating water impairs the critical real-time magnetic resonance imaging (MRI). An iron-based coupling medium (IBCM) using iron oxide nanoparticles previously developed by our group increased the relaxivity of the coupling bath such that it appears to be invisible on MRI compared with degassed water. However, the nanoparticles also reduced the pressure threshold for cavitation. To address this concern for prefocal cavitation, our group recently developed an IBCM of electrosterically stabilized and aggregation-resistant poly(methacrylic acid)-coated iron oxide nanoparticles (PMAA-FeOX) with a similar capability to reduce the MR signal of degassed water. This study examines the effect of the PMAA-FeOX IBCM on the cavitation threshold. METHODS: Increasing concentrations of PMAA-FeOX nanoparticles in degassed, deionized water were placed at the focus of two different transducers to assess low and high duty-cycle pulsing parameters which are representative of two modes of focused ultrasound being investigated for tMRgFUS. Passive cavitation detection and high-speed optical imaging were used to measure cavitation threshold pressures. RESULTS: The mean cavitation threshold was determined in both cases to be indistinguishable from the degassed water control, between 6-8 MPa for high duty-cycle pulsing (CW) and between 25.5-26.5 MPa for very low duty-cycle pulsing. CONCLUSION: The findings of this study indicate that an IBCM of PMAA-FeOX nanoparticles is a possible solution to reducing MRI interference from the coupling bath without increasing the risk of prefocal cavitation.

Iron-based coupling media for MRI-guided ultrasound surgery.

PURPOSE: In this study, we examine the effects of a recently developed, iron-based coupling medium (IBCM) on guidance magnetic resonance (MR) scans during transcranial, magnetic-resonance-guided, focused ultrasound surgery (tMRgFUS) procedures. More specifically, this study tests the hypotheses that the use of the IBCM will (a) not adversely affect image quality, (b) remove aliasing from small field-of-view scans, and (c) decouple image quality from the motion state of the coupling fluid. METHODS: An IBCM, whose chemical synthesis and characterization are reported elsewhere, was used as a coupling medium during tMRgFUS procedures on gel phantoms. Guidance magnetization-prepared rapid-gradient-echo (MP-RAGE), TSE, and GRE scans were acquired with fields of view of 28 and 18 cm. Experiments were repeated with the IBCM in several distinct flow states. GRE scans were used to estimate temperature time courses as a gel target was insonated. IBCM performance was measured by computing (i) the root mean square difference (RMSD) of TSE and GRE pixel values acquired using water and the IBCM, relative to the use of water; (ii) through-time temperature uncertainty for GRE scans; and (iii) Bland-Altman analysis of the temperature time courses. Finally, guidance TSE and GRE scans of a human volunteer were acquired during a separate sham tMRgFUS procedure. As a control, all experiments were repeated using a water coupling medium. RESULTS: Use of the IBCM reduced RMSD in TSE scans by a factor of 4 or more for all fields of view and nonstationary motion states, but did not reduce RMSD estimates in MP-RAGE scans. With the coupling media in a stationary state, the IBCM altered estimates of temperature uncertainty relative to the use of water by less than 0.2°C. However, with a high flow state, the IBCM reduced temperature uncertainties by the statistically significant amounts (at the 0.01 level) of 0.5°C (28 cm field of view) and 5°C (18 cm field of view). Bland-Altman analyses found a 0.1°C ± 0.5°C difference between temperature estimates acquired using water and the IBCM as coupling media. Finally, scans of a human volunteer using the IBCM indicate more conspicuous grey/white matter contrast, a reduction in aliasing, and a less than 0.2°C change in temperature uncertainty. CONCLUSIONS: The use of an IBCM during tMRgFUS procedures does not adversely affect image quality for TSE and GRE scans, can decouple image quality from the motion state of the coupling fluid, and can remove aliasing from scans where the field of view is set to be much smaller than the spatial extent of the coupling fluid.

Design of stable polyether-magnetite complexes in aqueous media: effects of the anchor group, molecular weight, and chain density.

The colloidal stability of polymer-stabilized nanoparticles is critical for therapeutic use. However, phosphates in physiological media can induce polymer desorption and consequently flocculation. Colloidal characteristics of PEO-magnetite nanoparticles with different anchors for attaching PEO to magnetite were examined in PBS. The effects of the number of anchors, PEO molecular weight, and chain density were examined. It was observed that ammonium phosphonates anchored PEO to magnetite effectively in phosphate-containing solutions because of interactions between the phosphonates and magnetite. Additionally, a method to estimate the magnetite surface coverage was developed and was found to be critical to the prediction of colloidal stability. This is key to understanding how functionalized surfaces interact with their environment.

The effect of polymer coatings on proton transverse relaxivities of aqueous suspensions of magnetic nanoparticles.

Iron oxide magnetic nanoparticles are good candidates for magnetic resonance imaging (MRI) contrast agents due to their high magnetic susceptibilities. Here we investigate 19 polyether-coated magnetite nanoparticle systems comprising three series. All systems were synthesized from the same batch of magnetite nanoparticles. A different polyether was used for each series. Each series comprised systems with systematically varied polyether loadings per particle. A highly significant (p < 0.0001) linear correlation (r = 0.956) was found between the proton relaxivity and the intensity-weighted average diameter measured by dynamic light scattering in the 19 particle systems studied. The intensity-weighted average diameter measured by dynamic light scattering is sensitive to small number fractions of larger particles/aggregates. We conclude that the primary effect leading to differences in proton relaxivity between systems arises from the small degree of aggregation within the samples, which appears to be determined by the nature of the polymer and, for one system, the degree of polymer loading of the particles. For the polyether coatings used in this study, any changes in relaxivity from differences in water exclusion or diffusion rates caused by the polymer are minor in comparison with the changes in relaxivity resulting from variations in the degree of aggregation.

Correction to "Luteinizing Hormone Releasing Hormone-Targeted Cisplatin-Loaded Magnetite Nanoclusters for Simultaneous MR Imaging and Chemotherapy of Ovarian Cancer".

[This corrects the article PMC10317193.].

Biocompatible, detachable, and free-standing polyelectrolyte multilayer films.

Self-assembled polyelectrolyte multilayers have gained tremendous popularity over the past decade and have been incorporated in diverse applications. However, the fabrication of detachable and free-standing polyelectrolyte multilayers (PEMs) has proven to be difficult. We report the design of detachable, free-standing, and biocompatible PEMs comprised of hyaluronic acid (anionic PE) and chitosan (cationic PE). These PEMs can be detached from an underlying inert substrate without any postprocessing steps. Our approach enables the fabrication of detachable PEMs from a wide range of polyelectrolytes. Cross-linked PEMs exhibited greater than 95% weight retention when maintained in phosphate buffered saline at 37 °C over a seven day period. The PEM thickness was approximately 3 µm for dried films and increased 2-fold under hydration. A unique feature of the detachable, free-standing PEMs is their optical transparency in the 400-900 nm range under hydrated conditions. The Young's modulus of the cross-linked films ranged from 300-400 MPa, rendering these detachable free-standing multilayers ideal for biomaterial applications. BALB/c 3T3 fibroblasts adhered on the PEMs and colonized the entire surface over a six day period. The cellular responses, as well as the physical properties, demonstrate that the detachable PEM films exhibit tremendous potential for applications in biomaterials and tissue engineering.

Novel acoustic coupling bath using magnetite nanoparticles for MR-guided transcranial focused ultrasound surgery.

PURPOSE: Acoustic coupling baths, nominally composed of degassed water, play important roles during transcranial focused ultrasound surgery. However, this large water bolus also degrades the quality of intraoperative magnetic resonance (MR) guidance imaging. In this study, we test the feasibility of using dilute, aqueous magnetite nanoparticle suspensions to suppress these image degradations while preserving acoustic compatibility. We examine the effects of these suspensions on metrics of image quality and acoustic compatibility for two types of transcranial focused ultrasound insonation regimes: low-duty cycle histotripsy procedures and high-duty cycle thermal ablation procedures. METHODS: Magnetic resonance guidance imaging was used to monitor thermal ablations of in vitro gel targets using a coupling bath composed of various concentrations of aqueous, suspended, magnetite nanoparticles in a clinical transcranial transducer under stationary and flowing conditions. Thermal deposition was monitored using MR thermometry simultaneous to insonation. Then, using normal degassed water as a coupling bath, various concentrations of aqueous, suspended, magnetite nanoparticles were placed at the center of this same transducer and insonated using high-duty cycle pulsing parameters. Passive cavitation detectors recorded cavitation emissions, which were then used to estimate the relative number of cavitation events per insonation (cavitation duty cycle) and the cavitation dose estimates of each nanoparticle concentration. Finally, the nanoparticle mixtures were exposed to low-duty cycle, histotripsy pulses. Passive cavitation detectors monitored cavitation emissions, which were used to estimate cavitation threshold pressures. RESULTS: The nanoparticles reduced the MR signal of the coupling bath by 90% in T2- and T2*-weighted images and also removed almost all imaging artifacts caused by coupling bath motion. The coupling baths caused <5% changes in peak temperature change achieved during sonication, as observed via MR thermometry. At low duty cycle insonations, the nanoparticles decreased the cavitation threshold pressure by about 15 ± 7% in a manner uncorrelated with nanoparticle concentration. At high duty cycle insonations, the 0.5 cavitation duty cycle acoustic power threshold varied linearly with nanoparticle concentration. CONCLUSIONS: Dilute aqueous magnetite nanoparticle suspensions effectively reduced MR imaging artifacts caused by the acoustic coupling bath. They also attenuated acoustic power deposition by <5%. For low duty cycle insonation regimes, the nanoparticles decreased the cavitation threshold by 15 ± 7%. However, for high-duty cycle regimes, the nanoparticles decreased the threshold for cavitation in proportion to nanoparticle concentration.

Nanoscale Bacteria-Enabled Autonomous Drug Delivery System (NanoBEADS) Enhances Intratumoral Transport of Nanomedicine.

Cancer drug delivery remains a formidable challenge due to systemic toxicity and inadequate extravascular transport of nanotherapeutics to cells distal from blood vessels. It is hypothesized that, in absence of an external driving force, the Salmonella enterica serovar Typhimurium could be exploited for autonomous targeted delivery of nanotherapeutics to currently unreachable sites. To test the hypothesis, a nanoscale bacteria-enabled autonomous drug delivery system (NanoBEADS) is developed in which the functional capabilities of the tumor-targeting S. Typhimurium VNP20009 are interfaced with poly(lactic-co-glycolic acid) nanoparticles. The impact of nanoparticle conjugation is evaluated on NanoBEADS' invasion of cancer cells and intratumoral transport in 3D tumor spheroids in vitro, and biodistribution in a mammary tumor model in vivo. It is found that intercellular (between cells) self-replication and translocation are the dominant mechanisms of bacteria intratumoral penetration and that nanoparticle conjugation does not impede bacteria's intratumoral transport performance. Through the development of new transport metrics, it is demonstrated that NanoBEADS enhance nanoparticle retention and distribution in solid tumors by up to a remarkable 100-fold without requiring any externally applied driving force or control input. Such autonomous biohybrid systems could unlock a powerful new paradigm in cancer treatment by improving the therapeutic index of chemotherapeutic drugs and minimizing systemic side effects.