Fabrication of 6-gingerol, doxorubicin and alginate hydroxyapatite into a bio-compatible formulation: enhanced anti-proliferative effect on breast and liver cancer cells

Manatunga et al. Chemistry Central Journal https://doi.org/10.1186/s13065-018-0482-6 (2018) 12:119 Chemistry Central Journal Open Access RESEARCH ARTICLE Fabrication of 6‑gingerol, doxorubicin and alginate hydroxyapatite into a bio‑compatible formulation: enhanced anti‑proliferative effect on breast and liver cancer cells Danushika C. Manatunga1, Rohini M. de Silva1* , K. M. Nalin de Silva1,2, Dulharie T. Wijeratne3, Gathsaurie Neelika Malavige3 and Gareth Williams4 Abstract Ample attention has been devoted to the construction of anti-cancer drug delivery systems with increased stability, and controlled and targeted delivery, minimizing toxic effects. In this study we have designed a magnetically attractive hydroxyapatite (m-HAP) based alginate polymer bound nanocarrier to perform targeted, controlled and pH sensitive drug release of 6-gingerol, doxorubicin, and their combination, preferably at low pH environments (pH 5.3). They have exhibited higher encapsulation efficiency which is in the range of 97.4–98.9% for both 6-gingerol and doxorubicin molecules whereas the co-loading has accounted for a value of 81.87 ± 0.32%. Cell proliferation assays, fluorescence imaging and flow cytometric analysis, demonstrated the remarkable time and dose responsive anti-proliferative effect of drug loaded nanoparticles on MCF-7 cells and HEpG2 cells compared with their neat counter parts. Also, these systems have exhibited significantly reduced toxic effects on non-targeted, non-cancerous cells in contrast to the excellent ability to selectively kill cancerous cells. This study has suggested that this HAP based system is a versatile carrier capable of loading various drug molecules, ultimately producing a profound anti-proliferative effect. Keywords: Hydroxyapatite, 6-Gingerol, Doxorubicin, MCF-7, HEpG2 Introduction Doxorubicin is an extensively used first line chemotherapeutic [1, 2] with an excellent effectiveness over a range of cancer types including breast cancer and liver cancer [3–7]. It is an anthracycline which exerts its anti-proliferation effect by intercalating with double stranded DNA, which could in turn arrest cell division and expression of vital proteins, and ultimately lead to cell death [4, 5]. However, later on it was observed that this particular drug is heavily associated with cardiotoxicity, neurotoxicity, myelosuppression, non-targeted killing of normal or *Correspondence: 1 Department of Chemistry, University of Colombo, Colombo 00300, Sri Lanka Full list of author information is available at the end of the article healthy cells, and the development of multi drug resistance (MDR), which has restricted its clinical efficacy and given rise to the recurrence of the cancers [8–11]. It has also been observed that the conjugation of doxorubicin with nanoparticulate systems such as superparamagnetic iron oxide nanoparticles would be an ideal approach to minimize the MDR while leading to enhanced cytotoxic effect over the drug resistant cancer cells [12, 13]. In addition, as a replacement approach for doxorubicin, the use of natural products as anti-cancer and cancer preventive agents has gained much attention over the past 30 years [14]. In this context, plant derived phytochemicals are preferred as they are generally less toxic and well tolerated by normal cells. These compounds generally contain a pool of active compounds © The Author(s) 2018. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/ publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Manatunga et al. Chemistry Central Journal (2018) 12:119 such as alkaloids, phenolics, tannins and flavonoids with very high activity, including anti-oxidant, antiinflammatory, anti-angiogenic, anti-microbial, anticancer activity [15]. Curcumin, gingerol, β-carotene, quercetin and linamarine are some of the commonly investigated compounds of plant extracts that are very effective and heavily investigated for the safer development of anti-cancer drugs [16–18]. 6-Gingerol is a polyphenolic active ingredient of the ginger rhizome, Zingiber officinale [19, 20], which is capable of reducing the growth of many cancer types [17, 21, 22]. 6-Gingerol can interfere with number of cell signaling pathways that control the balance between the cell apoptosis and proliferation [23]. These beneficial effects have been mainly assessed for breast cancer and in liver carcinoma [19, 24]. Moreover, it has also shown anti-microbial, anti-viral, cardio-protective, anti-hyperglycemic, anti-lipidemic and immunomodulatory effects [25–27]. Nevertheless, 6-gingerol has various drawbacks such as temperature, pH, and oxygen sensitivity, light instability, and poor aqueous solubility, hindering its potential applicability [28, 29]. Therefore, the development of drug carrier systems for the safer delivery of 6-gingerol in a targeted and controlled manner is highly essential. Therefore, attention has been devoted to the development of a nanoparticle based delivery for these compounds [30]. However, the use of carriers for the delivery of 6-gingerol is limited to a few studies [20, 26, 28, 31]. The co-delivery approach of 6-gingerol with toxic chemotherapeutics such as doxorubicin and cis-platin is another area of 6-gingerol utilization as it could synergistically act along with these drug molecules due its chemo preventive and chemo sensitive properties [20]. 6-Gingerol has been very effective in the elimination of the problem of MDR, seen with many chemotherapeutics [32]. Furthermore, the synergistic effect of 6-gingerol on neuroprotective, hepatoprotective, and anti-emetic properties has been exhibited when coadministering with doxorubicin [25, 32–37]. Nevertheless, it is worth noticing that the use nanoparticle based targeted and controlled drug delivery carriers for the dual loading of doxorubicin and 6-gingerol and enhancing their properties is not reported elsewhere. Therefore, in this study we have attempted to use a novel magnetic hydroxyapatite (m-HAP) nanoparticle system as an effective drug carrier for the controlled and pH sensitive delivery of 6-gingerol, doxorubicin and the dual drugs to inhibit the proliferation of breast and liver carcinoma cells targeting the development of a universal type drug carrier. Page 2 of 13 Materials and methods Materials 6-Gingerol (> 98.0%, HPLC), Doxorubicin hydrochloride (98.0–102.0%, HPLC), calcium nitrate tetrahydrate (Ca(NO3)2·4H2O, 99%, ACS), diammonium hydrogen phosphate ((NH4)2HPO4, > 99.0%), ammonium iron(II) sulfate hexahy (...truncated)