High sensitivity detection of nanoparticles permeation through polymer membranes: A physico-chemical and nuclear imaging measurement approach.

Diffusion cells are devices made of donor and acceptor compartments (DC and AC), separated by a membrane. They are widely used in pharmaceutical, cosmetic, toxicology, and protective equipment tests (e.g., gloves) to measure the kinetics of permeants (molecules and nanoparticles) across biological membranes as the skin. However, rarely is the concentration of permeants in the AC measured in continuous or in real-time, and this limitation leads to significant discrepancies in the calculations of kinetic parameters that define the permeation mechanisms. In this study, a diffusion cell compatible with positron emission tomography was used to measure the permeation kinetics of nanoparticles across glove membranes. The technology allows for the measurement of nanoparticle concentration in real-time in the two compartments (DC and AC) and at a detection sensitivity several orders of magnitude higher compared with conventional spectroscopies, thus allowing a much more precise extraction of kinetic parameters. Ultra-small (<10 nm) gold nanoparticles were used as a model nanoparticle contaminant. They were radiolabeled, and their diffusion kinetics was measured in continuous through latex and nitrile polymer membranes. Permeation profiles were recorded at sub-nanomolar sensitivity and in real-time, thus allowing the high precision extraction of kinetic permeation parameters. The technology, methodology, and data extraction process developed in this work could be applied to measure in real-time the kinetics of diffusion of a whole range of potentially toxic molecules and nanoparticles across polymer membranes, including glove membranes.

A diffusion cell adapted to nuclear imaging instruments for the measurement of molecular release and pharmacokinetics across membranes.

Diffusion cells are routinely used in pharmacology to measure the permeation of pharmaceutical compounds and contaminants across membranes (biological or synthetic). They can also be used to study drug release from excipients. The device is made of a donor (DC) and an acceptor (AC) compartment, separated by a membrane. Usually, permeation of molecules across membranes is measured by sampling from the AC at different time points. However, this process disturbs the equilibrium of the cell. Furthermore, analytical techniques used in association with diffusion cells sometimes lack either accuracy, sensitivity, or both. This work reports on the development of nuclear imaging - compatible diffusion cells. The cell is made of a polymer transparent to high-energy photons typically detected in positron emission tomography (PET). It was tested in a finite-dose set-up experiment with a pre-clinical PET system. Porous cellulose membranes (3.5, 25 and 300 kDa), a common excipient in pharmacology, as well as for dialysis membranes, were used as test membranes. The radioisotope 89Zr chelated with deferoxamine B (DFO; 0.65 kDa), was used as an imaging probe (7-10 MBq; 0.2-0.3 nMol 89Zr-DFO). In medicine, DFO is also commonly used for iron removal treatments and pharmacological formulations often require the association of this molecule with cellulose. Permeation profiles were obtained by measuring the radioactivity in the DC and AC for up to 2 weeks. The kinetic profiles were used to extract lag time, influx, and diffusion coefficients of DFO across porous cellulose membranes. A sensitivity threshold of 0.005 MBq, or 3.4 fmol of 89Zr-DFO, was revealed. The lag time to permeation (τ) measured in the AC compartment, was found to be 1.33, 0.5, and 0.19 h with 3.5, 25, and 300 kDa membranes, respectively. Diffusion coefficients of 3.65 × 10-6, 8.33 × 10-6, and 4.74 × 10-5 cm2 h-1 where revealed, with maximal pseudo steady-state influx values (Jpss) of 6.55 × 10-6, 1.76 × 10-5, and 1.29 × 10-5 nmol cm-2 h-1. This study confirms the potential of the technology for monitoring molecular diffusion and release processes at low concentrations, high sensitivities, in real time and in a visual manner.

High-Sensitivity Permeation Analysis of Ultrasmall Nanoparticles Across the Skin by Positron Emission Tomography.

Ultrasmall nanoparticles (US-NPs; <20 nm in hydrodynamic size) are now included in a variety of pharmacological and cosmetic products, and new technologies are needed to detect at high sensitivity the passage of small doses of these products across biological barriers such as the skin. In this work, a diffusion cell adapted to positron emission tomography (PET), a highly sensitive imaging technology, was developed to measure the passage of gold NPs (AuNPs) in skin samples in continuous mode. US-AuNPs (3.2 nm diam.; TEM) were functionalized with deferoxamine (DFO) and radiolabeled with 89Zr(IV) (half-life: 3.3 days, matching the timeline of diffusion tests). The physicochemical properties of the functionalized US-AuNPs (US-AuNPs-PEG-DFO) were characterized by FTIR (DFO grafting; hydroxamate peaks: 1629.0 cm-1, 1569.0 cm-1), XPS (presence of the O═C-N C 1s peak of DFO at 287.49 eV), and TGA (organic mass fraction). The passage of US-AuNPs-PEG-DFO-89Zr(IV) in skin samples was measured by PET, and the diffusion parameters were extracted thereby. The signals of radioactive US-AuNPs-PEG-DFO-89Zr(IV) leaving the donor compartment, passing through the skin, and entering the acceptor compartment were detected in continuous at concentrations as low as 2.2 nM of Au. The high-sensitivity acquisitions performed in continuous allowed for the first time to extract the lag time to the start of permeation, the lag time to start of the steady state, the diffusion coefficients, and the influx data for AuNPs permeating into the skin. PET could represent a highly valuable tool for the development of nanoparticle-containing topical formulations of drugs and cosmetics.

Gold Nanoparticles in Radiotherapy and Recent Progress in Nanobrachytherapy.

Over the last few decades, gold nanoparticles (GNPs) have emerged as "radiosensitizers" in oncology. Radiosensitizers are additives that can enhance the effects of radiation on biological tissues treated with radiotherapy. The interaction of photons with GNPs leads to the emission of low-energy and short-range secondary electrons, which in turn increase the dose deposited in tissues. In this context, GNPs are the subject of intensive theoretical and experimental studies aiming at optimizing the parameters leading to greater dose enhancement and highest therapeutic effect. This review describes the main mechanisms occurring between photons and GNPs that lead to dose enhancement. The outcome of theoretical simulations of the interactions between GNPs and photons is presented. Finally, the findings of the most recent in vivo studies about interactions between GNPs and photon sources (e.g., external beams, brachytherapy sources, and molecules labeled with radioisotopes) are described. The advantages and challenges inherent to each of these approaches are discussed. Future directions, providing new guidelines for the successful translation of GNPs into clinical applications, are also highlighted.

Intratumoral Injection of Low-Energy Photon-Emitting Gold Nanoparticles: A Microdosimetric Monte Carlo-Based Model.

Gold nanoparticles (Au NPs) distributed in the vicinity of low-dose rate (LDR) brachytherapy seeds could multiply their efficacy thanks to the secondary emissions induced by the photoelectric effect. Injections of radioactive LDR gold nanoparticles (LDR Au NPs), instead of conventional millimeter-size radioactive seeds surrounded by Au NPs, could further enhance the dose by distributing the radioactivity more precisely and homogeneously in tumors. However, the potential of LDR Au NPs as an emerging strategy to treat cancer is strongly dependent on the macroscopic diffusion of the NPs in tumors, as well as on their microscopic internalization within the cells. Understanding the relationship between interstitial and intracellular distribution of NPs, and the outcomes of dose deposition in the cancer tissue is essential for considering future applications of radioactive Au NPs in oncology. Here, LDR Au NPs (103Pd:Pd@Au-PEG NPs) were injected in prostate cancer tumors. The particles were visualized at time-points by computed tomography imaging ( in vivo), transmission electron microscopy ( ex vivo), and optical microscopy ( ex vivo). These data were used in a Monte Carlo-based dosimetric model to reveal the dose deposition produced by LDR Au NPs both at tumoral and cellular scales. 103Pd:Pd@Au-PEG NPs injected in tumors produce a strong dose enhancement at the intracellular level. However, energy deposition is mainly confined around vesicles filled with NPs, and not necessarily close to the nuclei. This suggests that indirect damage caused by the production of reactive oxygen species might be the leading therapeutic mechanism of tumor growth control, over direct damage to the DNA.

Low-Dose Prostate Cancer Brachytherapy with Radioactive Palladium-Gold Nanoparticles.

Prostate cancer (PCa) is one of the leading causes of death among men. Low-dose brachytherapy is an increasingly used treatment for PCa, which requires the implantation of tens of radioactive seeds. This treatment causes discomfort; these implants cannot be removed, and they generate image artifacts. In this study, the authors report on intratumoral injections of radioactive gold nanoparticles (Au NPs) as an alternative to seeds. The particles (103 Pd:Pd@Au-PEG and 103 Pd:Pd@198 Au:Au-PEG; 10-14 nm Pd@Au core, 36-48 nm hydrodynamic diameter) are synthesized by a one-pot process and characterized by electron microscopy. Administrated as low volume (2-4 µL) single doses (1.6-1.7 mCi), the particles are strongly retained in PCa xenograft tumors, impacting on their growth rate. After 4 weeks, a tumor volume inhibition of 56% and of 75%, compared to the controls, is observed for 103 Pd:Pd@Au-PEG NPs and 103 Pd:Pd@198 Au:Au-PEG NPs, respectively. Skin necrosis is observed with 198 Au; therefore, Au NPs labeled with 103 Pd only are a more advisable choice. Overall, this is the first study confirming the impact of 103 Pd@Au NPs on tumor growth. This new brachytherapy procedure could allow tunable doses of radioactivity, administered with smaller needles than with the current technologies, and leading to fewer image artifacts.

Towards a Multifunctional Electrochemical Sensing and Niosome Generation Lab-on-Chip Platform Based on a Plug-and-Play Concept.

In this paper, we present a new modular lab on a chip design for multimodal neurotransmitter (NT) sensing and niosome generation based on a plug-and-play concept. This architecture is a first step toward an automated platform for an automated modulation of neurotransmitter concentration to understand and/or treat neurodegenerative diseases. A modular approach has been adopted in order to handle measurement or drug delivery or both measurement and drug delivery simultaneously. The system is composed of three fully independent modules: three-channel peristaltic micropumping system, a three-channel potentiostat and a multi-unit microfluidic system composed of pseudo-Y and cross-shape channels containing a miniature electrode array. The system was wirelessly controlled by a computer interface. The system is compact, with all the microfluidic and sensing components packaged in a 5 cm × 4 cm × 4 cm box. Applied to serotonin, a linear calibration curve down to 0.125 mM, with a limit of detection of 31 µ M was collected at unfunctionalized electrodes. Added sensitivity and selectivity was achieved by incorporating functionalized electrodes for dopamine sensing. Electrode functionalization was achieved with gold nanoparticles and using DNA and o-phenylene diamine polymer. The as-configured platform is demonstrated as a central component toward an "intelligent" drug delivery system based on a feedback loop to monitor drug delivery.

Automated and reconfigurable platform for niosome generation based on a microfluidic architecture.

Drug delivery at the nano-scale is becoming an important topic in nano and regenerative medicine as it can offer a very localized therapy. Therefore, niosomes are one of the most important vehicles to release drug at the nanoscale. In this paper, we present a new automated microsystem for niosome generation on-demand. Used niosome were based on a mixture of cholesterol and dicetyl phosphate with chloroform. Three compact micropumps are connected to a microfluidic substrate in order to generate 100 nm noisome vesicles. Through this paper we also investigated the impact of using 150 µm pseudo-Y and cross shape microchannel on the diameter of vesicles. We have observed reliable results with Y-shaped microchannel, which was able to generate vesicles down to 91 nm. All the system is based on a low-cost fabrication process using dry photo resist.

Rapid Nucleation of Iron Oxide Nanoclusters in Aqueous Solution by Plasma Electrochemistry.

Progresses in cold atmospheric plasma technologies have made possible the synthesis of nanoparticles in aqueous solutions using plasma electrochemistry principles. In this contribution, a reactor based on microhollow cathodes and operating at atmospheric pressure was developed to synthesize iron-based nanoclusters (nanoparticles). Argon plasma discharges are generated at the tip of the microhollow cathodes, which are placed near the surface of an aqueous solution containing iron salts (FeCl2 and FeCl3) and surfactants (biocompatible dextran). Upon reaction at the plasma-liquid interface, reduction processes occur and lead to the nucleation of ultrasmall iron-based nanoclusters (IONCs). The purified IONCs were investigated by XPS and FTIR, which confirmed that the nucleated clusters contain a highly hydrated form of iron oxide, close to the stoichiometric constituents of α-FeOOH (goethite) or Fe5O3(OH)9 (ferrihydrite). Relaxivity values of r1 = 0.40 mM(-1) s(-1) and r2/r1 = 1.35 were measured (at 1.41 T); these are intermediate values between the relaxometric properties of superparamagnetic iron oxide nanoparticles used in medicine (USPIO) and those of ferritin, an endogenous contrast agent. Plasma-synthesized IONCs were injected into the mouse model and provided positive vascular signal enhancement in T1-w. MRI for a period of 10-20 min. Indications of rapid and strong elimination through the urinary and gastrointestinal tracts were also found. This study is the first to report on the development of a compact reactor suitable for the synthesis of MRI iron-based contrast media solutions, on site and upon demand.

Metal chelate grafting at the surface of mesoporous silica nanoparticles (MSNs): physico-chemical and biomedical imaging assessment.

Mesoporous silica nanoparticles (MSNs) are being developed as drug delivery vectors. Biomedical imaging (MRI and PET) enables their tracking in vivo, provided their surface is adequately grafted with imaging probes (metal chelates). However, MSNs are characterized by huge specific surfaces, and high-quality metal chelate anchoring procedures must be developed and validated, to demonstrate that their detection in vivo is associated to the presence of nanoparticles and not to detached metal chelates. MCM-48 nanospheres (M48SNs, 150 nm diam., 3-D pore geometry) were synthesized and functionalized with diethylenetriaminepentaacetic acid (DTPA). The strong grafting of DTPA was confirmed by 29Si MAS-NMR, XPS, FTIR and TGA. The particles were labeled with paramagnetic ions Gd3+ (for MRI) as well as radioactive ions 64Cu2+ (for PET; half-life: 12.7 h). Gd3+-DTPA-M48SNs formed a stable colloid in saline media for at least 6 months, without any sign of aggregation. The relaxometric properties were measured at various magnetic fields. The strength of DTPA binding at the surface of MSNs was also assessed in vivo, by injecting mice (i.v.) with Gd3+/64Cu2+-DTPA-M48SNs. Vascular retention and urinary clearance were monitored by MRI, whereas the PET modality provided dynamic and quantitative assessment of biodistribution and blood/organ clearance. No significant 64Cu activity was detectable in the bladder. The study confirmed the very limited detachment of DTPA from M48SNs cores once injected in vivo. The transit of MSNs through the liver and intestinal tract, does not lead to evidence of Gd3+/64Cu2+-DTPA in the urine. This physico-chemical and biodistribution study confirms the quality of DTPA attachment at the surface of the particles, necessary to allow further development of PET/MRI-assisted MSN-vectorized drug delivery procedures.

Multidentate block-copolymer-stabilized ultrasmall superparamagnetic iron oxide nanoparticles with enhanced colloidal stability for magnetic resonance imaging.

Ultrasmall superparamagnetic iron oxide nanoparticles (USPIOs) with diameters <5 nm hold great promise as T1-positive contrast agents for in vivo magnetic resonance imaging. However, control of the surface chemistry of USPIOs to ensure individual colloidal USPIOs with a ligand monolayer and to impart biocompatibility and enhanced colloidal stability is essential for successful clinical applications. Herein, an effective and versatile strategy enabling the development of aqueous colloidal USPIOs stabilized with well-defined multidentate block copolymers (MDBCs) is reported. The multifunctional MDBCs are designed to consist of an anchoring block possessing pendant carboxylates as multidentate anchoring groups strongly bound to USPIO surfaces and a hydrophilic block having pendant hydrophilic oligo(ethylene oxide) chains to confer water dispersibility and biocompatibility. The surface of USPIOs is saturated with multiple anchoring groups of MDBCs, thus exhibiting excellent long-term colloidal stability as well as enhanced colloidal stability at biologically relevant electrolyte, pH, and temperature conditions. Furthermore, relaxometric properties as well as in vitro and in vivo MR imaging results demonstrate that the MDBC-stabilized USPIO colloids hold great potential as an effective T1 contrast agent.

Manganese-impregnated mesoporous silica nanoparticles for signal enhancement in MRI cell labelling studies.

Mesoporous silica nanoparticles (MSNs) are used in drug delivery and cell tracking applications. As Mn(2+) is already implemented as a "positive" cell contrast agent in preclinical imaging procedures (in the form of MnCl2 for neurological studies), the introduction of Mn in the porous network of MSNs would allow labelling cells and tracking them using MRI. These particles are in general internalized in endosomes, an acidic environment with high saline concentration. In addition, the available MSN porosity could also serve as a carrier to deliver medical/therapeutic substances through the labelled cells. In the present study, manganese oxide was introduced in the porous network of MCM-48 silica nanoparticles (Mn-M48SNs). The particles exhibit a narrow size distribution (~140 nm diam.) and high porosity (~60% vol.), which was validated after insertion of Mn. The resulting Mn-M48SNs were characterized by TEM, N2 physisorption, and XRD. Evidence was found with H2-TPR, and XPS characterization, that Mn(II) is the main oxidation state of the paramagnetic species after suspension in water, most probably in the form of Mn-OOH. The colloidal stability as a function of time was confirmed by DLS in water, acetate buffer and cell culture medium. In NMR data, no significant evidence of Mn(2+) leaching was found in Mn-M48SNs in acidic water (pH 6), up to 96 hours after suspension. High longitudinal relaxivity values of r1 = 8.4 mM(-1) s(-1) were measured at 60 MHz and 37 °C, with the lowest relaxometric ratios (r2/r1 = 2) reported to date for a Mn-MSN system. Leukaemia cells (P388) were labelled with Mn-M48SNs and nanoparticle cell internalization was confirmed by TEM. Finally, MRI contrast enhancement provided by cell labelling with escalated incubation concentrations of Mn-M48SNs was quantified at 1 T. This study confirmed the possibility of efficiently confining Mn into M48SNs using incipient wetness, while maintaining an open porosity and relatively high pore volume. Because these Mn-labelled M48SNs express strong "positive" contrast media properties at low concentrations, they are potentially applicable for cell tracking and drug delivery methodologies.