Marco Giardiello

Antiretroviral solid drug nanoparticles with enhanced oral bioavailability: production, characterization, and in vitro-in vivo correlation.

Nanomedicine strategies have produced many commercial products. However, no orally dosed HIV nanomedicines are available clinically to patients. Although nanosuspensions of drug particles have demonstrated many benefits, experimentally achieving >25 wt% of drug relative to stabilizers is highly challenging. In this study, the emulsion-templated freeze-drying technique for nanoparticles formation is applied for the first time to optimize a nanodispersion of the leading non-nucleoside reverse transcriptase inhibitor efavirenz, using clinically acceptable polymers and surfactants. Dry monoliths containing solid drug nanoparticles with extremely high drug loading (70 wt% relative to polymer and surfactant stabilizers) are stable for several months and reconstitute in aqueous media to provide nanodispersions with z-average diameters of 300 nm. The solid drug nanoparticles exhibit reduced cytoxicity and increased in vitro transport through model gut epithelium. In vivo studies confirm bioavailability benefits with an approximately four-fold higher pharmacokinetic exposure after oral administration to rodents, and predictive modeling suggests dose reduction with the new formulation may be possible.

Accelerated oral nanomedicine discovery from miniaturized screening to clinical production exemplified by paediatric HIV nanotherapies

Considerable scope exists to vary the physical and chemical properties of nanoparticles, with subsequent impact on biological interactions; however, no accelerated process to access large nanoparticle material space is currently available, hampering the development of new nanomedicines. In particular, no clinically available nanotherapies exist for HIV populations and conventional paediatric HIV medicines are poorly available; one current paediatric formulation utilizes high ethanol concentrations to solubilize lopinavir, a poorly soluble antiretroviral. Here we apply accelerated nanomedicine discovery to generate a potential aqueous paediatric HIV nanotherapy, with clinical translation and regulatory approval for human evaluation. Our rapid small-scale screening approach yields large libraries of solid drug nanoparticles (160 individual components) targeting oral dose. Screening uses 1 mg of drug compound per library member and iterative pharmacological and chemical evaluation establishes potential candidates for progression through to clinical manufacture. The wide applicability of our strategy has implications for multiple therapy development programmes.

Mediation of in Vitro Cytochrome P450 Activity by Common Pharmaceutical Excipients

Polymers and surfactants are commonly used as excipients in oral formulations and are generally considered to be inert. However, relatively few studies have assessed their interaction with enzymes involved in the absorption, distribution, metabolism, and elimination of drugs. We have investigated the impact of twenty-three commonly used excipients (ten polymers and thirteen surfactants) on seven cytochrome P450 (CYP450) isoforms using baculosome-derived CYP450 enzymes across a range of concentrations. Time-course fluorescent readings were then taken to generate IC50 (inhibition) or EC50 (activation) values for excipient effects on CYP450 activity. All excipients had an observable effect activity of at least one CYP450 isoform with the majority of excipients altering substrate metabolism of at least 57% of CYP450s studied. In addition, most excipients were capable of inhibiting and increasing activity of several different CYP450 isoforms. Although the majority of these effects required concentrations outside those achievable therapeutically (>100 μM), almost 20% were seen at concentrations below 100 μM, and these results indicate that several excipients have the potential to modify the pharmacokinetics of administered drugs.

Facile synthesis of complex multi-component organic and organic–magnetic inorganic nanocomposite particles

A generic in situ method for producing triple component hydrophobic inorganic–organic nanocomposite particles, using a combination of modified emulsion templating and freeze-drying, is presented. Model nanocomposite particles have been developed bearing up to three hydrophobic ingredients chosen from polystyrene, oil red and 15–20 nm oleic acid-coated super-paramagnetic iron oxide (Fe3O4) nanoparticles. The technique avoids harsh conditions, in situ polymer synthesis and lengthy workup procedures, and results in high incorporation of magnetic particles (approximately 80% of triple-component nanocomposite particles contain magnetite) with retention of super-paramagnetism (>90% preservation). The nanocomposites have been characterised using dynamic light scattering, and studied under static and flow conditions in the presence of magnetic fields. Drug release was demonstrated using model nanocomposite particles bearing ibuprofen with differing hydrophobic polymer; polycaprolactone and poly(n-butyl methacrylate). Drug release varied with temperature, suggesting the synthetic technique could thus be adopted to develop drug carrier particles with tailored drug release properties.

Research Spotlight: Nanomedicines for HIV therapy

Heterogeneity in response to HIV treatments has been attributed to several causes including variability in pharmacokinetic exposure. Nanomedicine applications have a variety of advantages compared with traditional formulations, such as the potential to increase bioavailability and specifically target the site of action. Our group is focusing on the development of nanoformulations using a closed-loop design process in which nanoparticle optimization (disposition, activity and safety) is a continuous process based on experimental pharmacological data from in vitro and in vivo models. Solid drug nanoparticles, polymer-based drug-delivery carriers as well as nanoemulsions are nanomedicine options with potential application to improve antiretroviral deployment.

Flow cytometric analysis of the physical and protein-binding characteristics of solid drug nanoparticle suspensions

AIM Oral and intramuscular sustained-release antiretroviral solid drug nanoparticles (SDNs) are in development but there is limited understanding of whether nanoparticles or dissolved drug predominate systemically. MATERIALS & METHODS A flow cytometric method was developed to analyze SDNs in biological fluids such as plasma, including the putative formation of a protein corona. RESULTS SDNs were found to be stable in plasma and could be observed using the techniques developed here. In addition, transferrin, fibrinogen and albumin were found to be associated with SDNs upon incubation. CONCLUSION This methodology has enabled us to determine protein interactions of SDNs in solution without the requirement of separation from the matrix. This will enable further studies of their biological fate.

High-throughput nanoprecipitation of the organic antimicrobial triclosan and enhancement of activity against Escherichia coli

Enhancing the activity of existing antimicrobial agents may help to address the emergence of resistant bacteria. Nanoparticles of antimicrobial agents have previously been shown to provide potential activity enhancements and here we report a high-throughput nanoprecipitation approach to identify viable nanosuspensions of the antimicrobial compound triclosan. Through careful choice of the components of the nanoprecipitation, amorphous nanosuspensions were created, freeze-dried and redispersed in water with z-average diameters varying from 170-290 nm. Particle size was shown to be controlled by a series of factors including polymer/surfactant concentration and concentration of triclosan solution prior to nanoprecipitation. A ten-fold decrease (i.e. higher activity) in the Escherichia coli (E. coli) inhibitory concentration (IC50) of triclosan, compared to an aqueous control, was observed for nanoparticles prepared using Pluronic® F68 and the cationic surfactant Hyamine. This overall approach offers a rapid route for identifying viable nanosuspensions and enhancing the properties of commercially available biologically active compounds with poor water-solubility.

The Application of Nanotechnology to Drug Delivery in Medicine

Abstract This chapter examines the application of nanotechnology in medicine, known as nanomedicine, with a focus on the therapeutic application of nanomedicine. The different types of nanomaterials used in nanomedicine will be categorized based on their composition into three categories: solid drug nanoparticles, polymer therapeutics, and nanocarriers. A detailed description of the characteristics, design features, and synthetic approaches for each of these nanomaterial categories and their corresponding subcategories will be provided. The absorption, distribution, metabolism, and elimination processes will be introduced in context to the pharmacological benefits provided by nanomedicines. All the current clinically approved nanomedicines will be discussed from the perspective of the category of nanomaterial and their therapeutic application. The safety aspects of nanomedicines will then be examined with regards to the immunology of nanomaterials. The economic and infrastructural status, future direction and global initiatives present in nanomedicine will be outlined, before in the final section of the chapter, the different nanomedicines currently in clinical trials will be explored, and possible future directions of research will be summarized.

Augmented Inhibition of CYP3A4 in Human Primary Hepatocytes by Ritonavir Solid Drug Nanoparticles

Ritonavir is a protease inhibitor utilized primarily as a pharmaco-enhancer with concomitantly administered antiviral drugs including other protease inhibitors. However, poor tolerance, serious side effects, and toxicities associated with drug-drug interactions are common during exposure to ritonavir. The aim of this work was to investigate the impact of nanoformulation on ritonavir pharmacological properties. Emulsion-templated freeze-drying techniques were used to generate ritonavir (10 wt %) solid drug nanoparticle formulations. A total of 68 ritonavir formulations containing various mixtures of excipients were assessed for inhibition of CYP3A4 in baculosomes and primary human hepatocytes. Accumulation and cytotoxicity were assessed in HepG2 (hepatocytes), Caco-2 (intestinal), THP-1 (monocytes), A-THP-1 (macrophage), and CEM (lymphocytes). Transcellular permeation across Caco-2 cells was also assessed. From 68 solid drug nanoparticle formulations tested, 50 (73.5%) for baculosome and 44 (64.7%) for human primary hepatocytes exhibited enhanced CYP3A4 inhibition relative to an aqueous ritonavir solution. Sixty-one (89.7%) and 49 (72%) solid drug nanoformulations had higher apical to basal permeation across Caco-2 cells than aqueous solution of ritonavir after 60 and 120 min, respectively. No significant difference in cellular accumulation was observed for any solid drug nanoparticle for any cell type compared to aqueous ritonavir. However, incubation with the vast majority of solid drug nanoparticle formulations resulted in lower cytotoxicity of ritonavir than detected with an aqueous solution. These data provide in vitro proof of concept for improved inhibition of CYP3A4 by ritonavir through formation of solid drug nanoparticles. Nanodispersions also showed enhanced permeability across Caco-2 cells lower cytotoxicity across hepatic, intestinal, and immune cell types compared to an aqueous solution of ritonavir.

Intracellular delivery of nano-formulated antituberculosis drugs enhances bactericidal activity: Intracellular delivery of nano-formulated anti-TB drugs

Tuberculosis kills more people worldwide than any other infectious disease. Treatment requires multiple drug therapy administered over long periods (6–24 months). The emergence of multidrug‐resistant strains is a major problem, and with few new drugs in the pipeline, a novel modus operandi is urgently required. Solid drug nanoparticles (SDNs), a new development in nanomedicine, offer a fresh therapeutic approach. Here, we show that SDNs are more effective (50‐fold) at killing pathogenic mycobacteria than aqueous forms of the same drug and can target mycobacteria internalised by macrophages, where bacilli reside. We demonstrate synthesis of dual and triple drug loaded SDNs, facilitating combination tuberculosis therapy. Our results suggest that by employing SDNs of existing antibiotics, it may be possible to improve drug delivery and therefore reduce drug dosage to lessen side effects and fight drug resistance.