Sclareol-loaded hyaluronan-coated PLGA nanoparticles: Physico-chemical properties and in vitro anticancer features

Highlights

• Sclareol (SCL) has been encapsulated within PLGA nanoparticles

• The nanoencapsulation of SCL favors its administration in physiological media

• The coating of SCL-nanoparticles with hyaluronan increases their anticancer activity on HA+ cells

Abstract

Sclareol (labd‑14‑ene‑8,13‑diol), a phytochemical compound belonging to labdane-type diterpenes, has recently attracted noteworthy attention because of its peculiar pharmacological properties. The foremost obstacle to an efficacious application and use of this molecule is its unfavorable bioavailability due to its poor aqueous solubility. In this investigation sclareol was encapsulated within PLGA nanoparticles with the aim of favoring its administration in physiological media, increasing its anticancer activity and obtaining a stable colloidal formulation. These nanoparticles containing sclareol were characterized by a mean diameter of 100–150 nm, a narrow size distribution and a negative surface charge. The active compound was efficiently retained by the polymeric structure and did not induce any physical destabilization. The coating of the PLGA nanoparticles with hyaluronic acid (1.5 MDa) increased the antitumor efficacy of the encapsulated drug against human breast cancer cells expressing the hyaluronan receptors (MCF-7 and MDA-MB468) while a similar pharmacological effect was obtained on human colon carcinoma cells (CaCo-2). CLSM analysis demonstrated the intracellular localization of fluorescent nanosystems after 3 h incubation.

Introduction

The concept of nanoencapsulation has often been used in recent decades in order to modulate the biopharmaceutical properties of various active compounds [[1], [2], [3]]. Polymeric nanosystems, for example, were found to favor the administration of lipophilic and water-soluble compounds in physiological environments, acting as suitable pharmacokinetic biomodulators [4,5]. The encapsulation of lipophilic drugs within biocompatible materials, in fact, achieves greater therapeutic efficacy besides bringing about a decrease in the most common side effects that were due to the use of organic solvents for their dissolution; this was true, for instance, in the case of taxanes before the development of nab-technology [6,7]. Sclareol (labd‑14‑ene‑8,13‑diol), another natural lipophilic compound belonging to the labdane-type diterpenes, has exhibited significant cytostatic and cytotoxic effects on various human cancer cell lines [8]. Its anticancer features are based on the induction of cell apoptosis promoted by p53-independent mechanisms; in particular, cell death occurs upon the activation of caspases-8 and -9, followed by the activation of caspase-3, and the subsequent degradation of PARP [8,9]. Due to its physico-chemical properties and structural analogy to cholesterol, the drug was encapsulated within liposomes in order to favor its administration in aqueous media. The vesicular formulation evidenced a variable cytotoxicity as a function of its cell line characteristics, besides assuming a different localization in the subcellular compartments with respect to the free form [10,11]. In particular, SCL was totally localized in the cytosol while the liposomal formulation was distributed in the cell membranes, influencing its pharmacological activity as a consequence of its slow release profile from the various bilayers [10]. The aim of this experimental work was to investigate the influence of SCL on the physico-chemical properties of biocompatible polymeric nanoparticles. The polymer used to develop the system able to retain the drug was polyester poly‑lactic‑co‑glycolic acid (PLGA), due to its biodegradability and because it has the approval of the FDA and EMA for pharmaceutical application and human administration [12]. Moreover, the possibility of increasing the cell targeting of the nanosystems by using suitable agents may represent a useful strategy able to maximize the anticancer efficacy of SCL [13,14]; this is why the PLGA nanoparticles were coated with hyaluronan (HA) in order to obtain specific targeting against the cells expressing HA-receptors [15,16]. HA is a glycosaminoglycan made up of repeated units of d‑glucuronic acid and N‑acetyl‑d‑glucosamine linked by alternating β-1,3 and β-1,4 glycosidic bonds; it can be used as a targeting agent against various receptors such as the cluster determinant 44 (CD44), the receptor for hyaluronate-mediated motility (RHAMM), the HA receptor for endocytosis (HARE) and the lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1) [[17], [18], [19]]. It has been shown that high-molecular-weight HA is involved in maintaining cell integrity and water content in the extracellular matrix, while low-molecular-weight HA modulates receptor-mediated intracellular signaling besides being responsible for angiogenesis and inflammatory phenomena [20,21]. The PLGA nanosystems were coated with high-molecular-weight HA (1500 kDa) in order to increase the delivery of the SCL against the HA+ cells as well as prolong the circulation of the particles after systemic administration, avoiding the use of PEG-derivatives [19,20]; the amount of HA adsorbed on the particle surface was investigated by analytical testing and the biological features of the resulting nanocarrier were evaluated.