PLGA–lecithin–PEG core–shell nanoparticles for controlled drug delivery

Abstract

Current approaches to encapsulate and deliver therapeutic compounds have focused on developing liposomal and biodegradable polymeric nanoparticles (NPs), resulting in clinically approved therapeutics such as Doxil/Caelyx and Genexol-PM, respectively. Our group recently reported the development of biodegradable core–shell NP systems that combined the beneficial properties of liposomal and polymeric NPs for controlled drug delivery. Herein we report the parameters that alter the biological and physicochemical characteristics, stability, drug release properties and cytotoxicity of these core–shell NPs. We further define scalable processes for the formulation of these NPs in a reproducible manner. These core–shell NPs consist of (i) a poly(d,l-lactide-co-glycolide) hydrophobic core, (ii) a soybean lecithin monolayer, and (iii) a poly(ethylene glycol) shell, and were synthesized by a modified nanoprecipitation method combined with self-assembly. Preparation of the NPs showed that various formulation parameters such as the lipid/polymer mass ratio and lipid/lipid–PEG molar ratio controlled NP physical stability and size. We encapsulated a model chemotherapy drug, docetaxel, in the NPs and showed that the amount of lipid coverage affected its drug release kinetics. Next, we demonstrated a potentially scalable process for the formulation, purification, and storage of NPs. Finally, we tested the cytotoxicity using MTT assays on two model human cell lines, HeLa and HepG2, and demonstrated the biocompatibility of these particles in vitro. Our data suggest that the PLGA–lecithin–PEG core–shell NPs may be a useful new controlled release drug delivery system.

Introduction

Over the past two decades, there has been a steady rise in the number of commercially available nanoparticle (NP) therapeutics [1], [2], [3]. Among these products, liposomal drugs [4] and polymer–drug conjugates [5] are two dominant classes, accounting for the majority of clinically approved products. Concurrently, the development of biodegradable polymeric NPs in the ∼100 nm range has become an increasingly exciting field in academic and industrial research [2], [3].

Doxil (liposomal doxorubicin) was the first Food and Drug Administration (FDA)-approved liposomal drug formulation for the treatment of AIDS associated with Kaposi's sarcoma in 1995 [6], [7], [8]. The benefits of liposomal formulations include their ability to encapsulate hydrophilic therapeutic agents at high loading efficiency, shield encapsulated drugs from external conditions, and also be coated with inert and biocompatible polymers such as polyethylene glycol (PEG) for reduced systemic clearance rates and prolonged circulation half-life in vivo [9]. These PEG-end groups may also be functionalized with specific ligands for targeting to specific sites of the cells, tissues and organs of interest [10]. However, liposomes may face some obstacles from their low ability to encapsulate very hydrophobic drugs, burst release of drugs, and having multiple manufacturing steps associated with liposome preparation and purification.

The use of biodegradable polymeric NPs for drug delivery has been gaining momentum and shown significant therapeutic potential [11], [12], [13]. Biodegradable polymers such as poly(d,l-lactic acid), poly(d,l-lactic-co-glycolic acid) and poly(ɛ-caprolactone) and their co-polymers diblocked or multiblocked with PEG have been commonly used to form core–shell structured NPs to encapsulate a variety of therapeutic compounds [14], [15], [16], [17]. These NPs have a number of appealing features: their hydrophobic core is capable of carrying highly insoluble drugs with high loading capacity, while their hydrophilic shell provides steric protection and functional groups for surface modification. Drug release can be manipulated by choosing biodegradable polymers with different surface or bulk erosion rates, and external conditions such as pH and temperature changes may function as a switch to trigger drug release [18]. Worth noting is their ease of manufacturing: amphiphilic co-polymers spontaneously assemble into core–shell structures in aqueous environments [19]. However, to date, polymeric NPs have shown moderate circulation half-lives compared to their liposomal counterparts, despite also being coated with inert and biocompatible polymers such as polyethylene glycol (PEG) [20].

Recently, we developed in our laboratory a protocol for the self-assembly of NPs that combine the properties of liposomes and polymeric NPs [21]. Existing strategies to make such fusion particles (lipopolyplexes) involved multi-step synthesis methods, resulting in inherently inefficient systems which are not easily scalable and show batch-to-batch variation [22], [23]. We engineered a simple, scalable, efficient and more controllable system using a well-defined and predictable formulation strategy. The NPs are formed from three biomaterials: (i) poly(d,l-lactide-co-glycolide) (PLGA) was selected for the hydrophobic core due to its biodegradable nature and ability to encapsulate high amounts of hydrophobic drugs; (ii) soybean phosphatidylcholine (or lecithin) was chosen for a monolayer around the hydrophobic core; and (iii) 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-carboxy(polyethylene glycol)2000 (DSPE–PEG–COOH) intersperses in the lecithin monolayer to form a PEG shell which provides electrostatic and steric stabilizations, a longer circulation half-life in vivo as well as functional-end groups for the attachment of targeting ligands such as antibodies, peptides and aptamers.

Herein we report our studies on the systematic preparation and characterization of these PLGA–lecithin–PEG core–shell NPs. We evaluate parameters that affect the core–shell nanostructure to find an optimal formulation, and subsequently characterize the NPs for physical stability, controlled drug release kinetics, post-formulation purification, storage methods and finally material cytotoxicity. We establish the function of the lipid monolayer ring between the PLGA core and the PEG shell in controlled drug release kinetics. This novel NP system may represent a new way of combining existing lipid and polymer classes of materials for controlled drug delivery applications.