A winning strategy to improve the anticancer properties of Cisplatin and Quercetin based on the nanoemulsions formulation

Abstract

The aim of the present study was the proposal of a strategy to permit the safe use of the powerful antitumor agent Cisplatin. Considering the possibility of reducing the oxidative stress responsible for the Cisplatin induced nephrotoxicity by using antioxidants, such as flavonoids, we proposed the concomitant therapeutic association of Cisplatin with Quercetin. Considering the poor solubility of Quercetin, nanoemulsions have been proposed to encapsulate and facilitate the use of the drug. Originally, the concomitant encapsulation of Quercetin with Cisplatin has been also assessed. The obtained formulations have been characterized and tested against two human cell models, namely the human breast cancer MDA-MB-231 and the normal HEK-293 renal cells. We demonstrated that the new formulated nanoemulsions have improved the anticancer properties of both the molecules and, most importantly, the synergistic effect of the Cisplatin/Quercetin against the MDA-MB-231. Moreover, the dramatic cytotoxic effects of Cisplatin against the human renal HEK-293 cells have been significantly diminished by the use of nanoemulsions containing both the molecules, whereas the antioxidant properties of encapsulated Quercetin have been improved. Finally, the simple process used to obtain the nanoemulsions and the physico-chemical properties compatible with parenteral administration, the stability and the improved pharmaceutical effects contribute to the high potential of this strategy to be studied in future in vivo studies.

Introduction

Cisplatin is a powerful antitumor agent, widely used in clinical, especially for the treatment of solid tumors (uterine, testicular, bladder, lung, head and neck tumors). It interacts with cellular proteins and lipids, forms DNA adducts that cause cell cycle arrest in the G2/M phase [[1], [2], [3], [4]] and can also interfere with cytoskeleton organization [5]. However, the onset of resistance phenomena and its severe adverse effects, including nephrotoxicity, intestinal toxicity and myelosuppression, are some major limitations for its effectiveness and use in clinical medicine [[6], [7], [8], [9]]. The mechanism underlying the Cisplatin-induced nephrotoxicity is not yet known, but in vitro and in vivo studies indicate that is closely associated with an increase of lipid peroxidation [6,[10], [11], [12]]. Thus, the reactive oxygen species (ROS) seem to play a central role in renal injury induced by such chemotherapy [6,13]. The oxidative stress responsible for the Cisplatin-induced nephrotoxicity can be reduced, for instance, using different antioxidants, such as flavonoids. Among these, Quercetin is one of the most abundant flavonoids in the human diet and it is commonly found in most edible fruits and vegetables [14]. Quercetin is an extraordinary scavenger of reactive oxygen species, a metal chelator and it is capable of inhibiting lipid peroxidation, blocking the enzyme xanthine oxidase as demonstrated in in vitro and in vivo systems. Indeed, literature data showed that Quercetin possesses a high radical scavenging activity, even at low concentrations, and the highest antioxidant potential compared to the standard antioxidants (vitamin C and Trolox) and to other powerful antioxidants (Resveratrol, Ferulic Acid, Gallic Acid, Kampferol, etc.) [15,16].

Moreover, many studies indicated that a pre-treatment with Quercetin had a protective role against the Cisplatin-induced nephrotoxicity [2,6,17].

Together with the powerful antioxidant effect, Quercetin exhibits an excellent anti-proliferative activity, particularly toward the highly aggressive and metastatic breast cancer cells (MDA-MB-231) [18]. Furthermore, the combination of Quercetin and Cisplatin could have a synergistic effect, resulting more effective in the induction of growth suppression and apoptosis, compared to the single treatment [6,9]. However, as for Cisplatin, the Quercetin clinical use is very limited because of its poor solubility in water (0.00215 g/L at 25 °C) [19] and bioavailability [20]. Recent studies have shown that the administration of drugs encapsulated in nanodelivery systems shows a greater efficacy together with the reduction of undesired effects [21,22]. The repurposing of old approved drugs, to treat cancer and overcome pharmacokinetic issues, drug over-dosage and consequent side effects onset, and resistance phenomena, has been strongly supported by nanotechnologies [23]. In this scenario, nanoemulsions evolved as carriers particularly appealing for the hydrophobic drugs delivery. Nanoemulsions (NE) belong to the lipid-based drug delivery systems. They consist of nanoscale oil droplets, are highly biocompatible and have been successfully used to ameliorate the drugs solubilisation, stability and membranes permeation [[24], [25], [26]]. The NE formulation through low energy approaches is a simple and low cost process that produce fine droplets and can be easily implemented [27]. In a previous work of our team, NE have been successfully developed from a very simple spontaneous emulsification process, and they have been shown to be able to encapsulate drugs with limited solubility, with high loading rates [26,28].

For these reasons, we have decided to propose NE to increase the water solubility and the bioavailability of Quercetin, in order to increase its antioxidant and antitumor activities. In this way, we aimed also at improving the therapeutic profile of Cisplatin. These developed formulations have been fully characterized for their physicochemical properties and the antioxidant potential of Quercetin loaded NE has been determined by the means of ORAC assay.

Finally, the improved anticancer properties of NE encapsulating Cisplatin and Quercetin have been evaluated against the triple negative and highly aggressive MDA-MB-231 breast cancer cells and the diminished cytotoxicity of Cisplatin against the Hek-293 human embryonic kidney epithelial cells has been proved.