Synthesis and in vitro/in vivo anti-cancer evaluation of curcumin-loaded chitosan/poly(butyl cyanoacrylate) nanoparticles

Abstract

We have synthesized novel cationic poly(butyl) cyanoacrylate (PBCA) nanoparticles coated with chitosan, formulation of curcumin nanoparticles. The size and zeta potential of prepared curcumin nanoparticles were about 200 nm and +29.11 mV, respectively with 90.04% encapsulation efficiency. The transmission electron microscopy (TEM) study revealed the spherical nature of the prepared nanoparticles along with confirmation of particle size. Curcumin nanoparticles demonstrate comparable in vitro therapeutic efficacy to free curcumin against a panel of human hepatocellular cancer cell lines, as assessed by cell viability (3-[4,5-dimethylthiazol-2-yl]2,5-diphenyltetrazolium bromide assay [MTT assay]) and proapoptotic effects (annexin V/propidium iodide staining). In vivo, curcumin nanoparticles suppressed hepatocellular carcinoma growth in murine xenograft models and inhibited tumor angiogenesis. The curcumin nanoparticles’ mechanism of action on hepatocellular cancinoma cells is a mirror that of free curcumin.

Introduction

Curcumin, a yellow polyphenol extracted from the rhizome of turmeric (Curcuma longa), is been extensively used in China, India and southeast tropical Asia as a spice and colouring material, and a treatment for a wide variety of ailments including biliary disorders, hepatic disorders, diabetic wounds, anorexia, rheumatism, and sinusitis (Goel et al., 2008, Aggarwal and Harikumar, 2009, Anand et al., 2007). Recent studies have shown that curcumin, either alone or in combination with other anticancer agents, has potent anticancer effects. This was evidenced by its inhibitory effects on the growth of a number of tumor cell lines in vitro and in vivo, including melanoma, mantle cell lymphoma (MCL), hepatic, prostatic, ovarian, and pancreatic carcinomas (Aggarwal et al., 2003, Siwak et al., 2005, Gururaj et al., 2002, Belakavadi and Salimath, 2005, Shishodia et al., 2005, Zheng et al., 2004). Curcumin has been reported to have diverse effects on signaling molecules, such as downregulation of the expression of angiogenesis-associated genes, activation of the apoptotic mechanisms, and induction of the cell cycle arrest (Gururaj et al., 2002, Belakavadi and Salimath, 2005, Shishodia et al., 2005).

Despite great therapeutic potential for curcumin utilization in a variety of diseases, its clinical development has been hindered due to its fast metabolism and poor water solubility (Wang et al., 1997, Tonnesen et al., 2002, Cheng et al., 2001, Shoba et al., 1998). The main shortcomings of curcumin are its lipophilicity hence, low bioavailability, degradation at alkaline pH and photodegradation. Lipophilic nature of curcumin makes it vulnerable to RES uptake and hence it can’t reach the therapeutic target with therapeutic concentration.

To increase its aqueous solubility and bioavailability, attempts have been made through encapsulation in liposome, polymeric nanoparticle, lipid-based nanoparticle, biodegradable microsphere, cyclodextrin and hydrogel (Bisht et al., 2007, Tiyaboonchai et al., 2007, Kunwar et al., 2006, Sou et al., 2008, Salmaso et al., 2007, Vemula et al., 2006). Interest in nanocarriers for cancer chemotherapy is growing. Nanoparticle-based drug delivery approaches have the potential for rendering hydrophobic agents like curcumin dispersible in aqueous media, thus circumventing the pitfalls of poor solubility.

Design and development of biodegradable controlled drug delivery of therapeutic entities with improved bioavailability is the main research aspect on which extensive work is been done in the past few decades. One of the promising and exciting drug delivery system which can meet the above mentioned requirements is polymeric nanoparticles. Among the various promising polymers like PLA, PLGA, polycaprolactone, polymethylidene malonate, poly(butyl) cyanoacrylate (PBCA) nanoparticles meet ideal requirements for controlled drug delivery and passive targeting such as biodegradability, low toxicity, ability to alter the biodistribution of drugs and easy to synthesize and purify (Behan et al., 2001, Juan et al., 2006, Maksimenko et al., 2008, Reddy et al., 2004).

PBCA nanoparticles have been proved to be an effective drug delivery system for the controlled drug delivery of various pharmacologically active moieties, like anticancer agents, analgesics, antibiotics and peptide (Huang et al., 2007, Miyazaki et al., 2003, Page-Clisson et al., 1998, Tasset et al., 1995). The PBCA nanoparticles are generally prepared by emulsion or dispersion polymerization in an acidic aqueous solutions of surfactants forming a porous structure with high specific area on which various quantities of drugs, dissolved in the medium during or after polymerization, can be loaded (Petri et al., 2007). Surface characteristics like zeta potential and the solubility property can be modulated to meet the prerequisites of intravenous administration.

Curcumin has been conjugated with numerous carriers including phospholipids (Maiti et al., 2007), cyclodextrin (Salmaso et al., 2007), phosphatidyl choline (Marczylo et al., 2007), and liposomes (Kunwar et al., 2006), but very little information is available about its biological activities.

In this study, we describe a simple method of synthesis and characterization of a novel cationic poly(butyl) cyanoacrylate nanoparticles coated with chitosan encapsulated formulation of curcumin that can improve the solubility of curcumin and prevents RES uptake. In various kinds of poly(alkyl) cyanoacrylates (PACAs), PBCA was the most used drug carrier because it can interact with different kind of drugs which will aid in increased curcumin entrapment efficiency. The development of a delivery system that can enable parenteral administration of curcumin in an aqueous phase medium will significantly harness the potential of this promising anti-cancer agent in the clinical arena. We investigated the in vitro and in vivo antitumor activity of curcumin-PBCA nanoparticles against human hepatocellular carcinoma cells. In vivo, curcumin nanoparticles inhibited hepatic carcinoma growth and demonstrated antiangiogenic effects.