Preparation, characterization and pharmacokinetic studies of tacrolimus-dimethyl-β-cyclodextrin inclusion complex-loaded albumin nanoparticles

Abstract

The purpose of the study is to develop a new formulation for clinically used anti-cancer agent tacrolimus (FK506) to minimize the severe side effects. Toward this end, a new formulation method has been developed by complexation of FK506 with an hydrophilic cyclodextrin derivative, heptakis (2,6-di-O-methyl)-β-cyclodextrin (DM-β-CD) using ultrasonic means. The resulting complex displays dramatically enhanced solubility of FK506. Then bovine serum albumin (BSA) nanoparticles were prepared directly from the preformed FK506/DM-β-CD inclusion complex by the desolvation-chemical crosslinking method, with the size of 148.4–262.9 nm. Stable colloidal dispersions of the nanoparticles were formed with zeta potentials of the range of −24.9 to −38.4 mV. The entrapment efficiency of FK506 was increased as high as 1.57-fold. Moreover, notably FK506 was released from the nanoparticles in a sustained manner. As demonstrated, pharmacokinetic studies reveal that, as compared with FK506-loaded BSA nanoparticles, the FK506/DM-β-CD inclusion complex-loaded BSA nanoparticles have significant increase at T_max, t_1/2, MRT and decrease at C_max. In summary, these results suggest that the drug/DM-β-CD inclusion complex-loaded BSA nanoparticles display significantly improved delivery efficiency for poorly soluble FK506 or its derivatives.

Introduction

Tacrolimus (FK506, molecular weight of 822.05, water solubility of 1.3 μg/ml), a hydrophobic macrolide lactones natural product isolated from by Streptomyces tsukubaensis, exerts potent immunosuppressive effects and has been in clinical use as prophylaxis against organ rejection after liver and renal transplantation (Hidetoshi et al., 2001). Recently, it has been reported that FK506 can be widely distributed in the body with a high degree binding of red blood cells and plasma proteins. However, the distribution is significantly affected by individual differences, and the administration routes. For example, the gastrointestinal tract has a narrow therapeutic window due to low bioavailability (Taher et al., 2009), and side effects. In addition, FK506 is known to exhibit low oral bioavailability and a wide range of variability in absorption, ranging from 4 to 89% in kidney and liver transplant recipients (Venkataramanan et al., 1995). For the intravenous administration, because of its low solubility, some solubilizer or injection oil was used, thus inducing greater toxicity.

β-Cyclodextrin (β-CD) and its derivatives, as distinct solubilizers, have received considerable attention, giving prominence to their low biotoxicity and high biocompatibility. As attractive materials for drug inclusion, β-CD can be further chemically modified to improve its physicochemical properties (Hassan and Asghar, 2009, Song et al., 2009). A number of studies have shown that adding β-CD can improve the loading efficiency of nanoparticles and slow down the release of drugs (Alexander and Maria, 2007, Boudad et al., 2001, Maestrelli et al., 2006). Ferreira and collaborators prepared inclusion complexes with hydroxypropyl-β-cyclodextrin and the aqueous solubility of the drug increased linearly with the concentration of the cyclodextrins (Denise et al., 2004). The cavity depth and surface activity of 2,6-di-O-methyl-β-cyclodextrin (DM-β-CD), a derivative of β-CD, improved significantly, as well as the solubility increased as much as by 25 times (Gamal et al., 1986). Various β-CD derivatives have been evaluated to probe their enhancing effect on solubility and stability of FK506 in rats. It was found that DM-β-CD had a dramatic improvement on solubilization and stabilization of FK506 (Hidetoshi et al., 2001).

Modern nanotechnology is considered as an emerging and converging technology (Roco, 2008) and that is said to be one of the key technologies of the 21st century. Nanotechnology is widely seen as having huge potential to bring benefits to many areas of research and application. Studies have shown that albumin nanoparticle has a higher capacity of loading hydrophilic drugs, better performance of controlled release and more stability of storage (MacAdam et al., 1997, Orapin et al., 1993). Furthermore, albumin offers several notable characteristics, including safety, non-toxic, non-immunogenicity, biodegradability and good biocompatibility. Therefore it serves an attractive drug delivery system. After release of drugs, albumin nanoparticles can be absorbed by body through the metabolic decomposition without producing harmful residual substances (Langer et al., 2003, Yun et al., 2007). In addition, the albumin molecule has many functional groups, thus allowing for convenient functionalizations for various purposes. General methods for preparation of albumin nanoparticles include ultrasonic emulsification, desolvation method, pH coagulation, salting, etc. (Langer et al., 2003, Muller et al., 1996, Merodio et al., 2001, Weber et al., 2000). For the desolvation method, several factors can affect albumin nanoparticles such as drugs, the dosage of albumin and the preparation of albumin nanoparticles including dehydrolyzing agent, cross linking agent, pH, cross linking time and stirring speed (Guilin et al., 2008, Hyuncheol et al., 2009, Vogel et al., 2002). With the carrier of albumin, albumin nanoparticles are solid sphere obtained by curing and separation, which can encapsulate and adsorb different kinds of drugs such as polypeptides, vaccines and gene to be used in varied disease. Controlled the nanosize of albumin nanoparticles can not only reduce toxicity but achieve certain sustained effect when used in intravenous. Currently, the research of FK506-loaded nanoparticles has mainly focused on the use of polylactic acid and acrylic acid as the carrier material (Nakaoka et al., 1995, Nakase et al., 2000). The study on the FK506/DM-β-CD inclusion complex-loaded albumin nanoparticles has not been reported.

The purpose of the present study is to develop a novel nano-drug delivery system (NDDS), drug/DM-β-CD inclusion complex-loaded BSA nanoparticles. Firstly, FK506 was complexed to a hydrophilic cyclodextrin derivative via the formation of inclusion complex of the drug with DM-β-CD by an ultrasonic method. Then, FK506/DM-β-CD inclusion complex-loaded bovine serum albumin (BSA) nanoparticles were prepared by desolvation-chemical crosslinking method. The physicochemical characteristics were determined (i.e. entrapment efficiency, loading efficiency, in vitro release, size distribution of the developed nanoparticles). In addition, pharmacokinetic parameters of these nanoparticles were investigated in rats.