In vitro and in vivo anticancer activity of Gomphocarpus Fruticosus extracts and development of LIPID-based nanoparticles
Loading...
Date
2019
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
University of Namibia
Abstract
Cancer is on the rise globally, with incidences and mortalities increasing in developing countries including in Namibia. The ongoing search for efficacious and safe medicines to treat cancer is often cumbered by poor solubility and toxicity of potential treatments. Gomphocarpus fruticosus is a medicinal plant used to treat tumors in amibia through topical application and it has been shown to have anticancer action against several cell lines. I lowcver, due to its cardiotoxic properties, it has not been investigated further. This study was conducted to develop a lipid-based drug delivery system based on Gomphocarpusfruticosus extracts, to alleviate the plants toxicity profile while harnessing its anticancer properties. The phytochemistry of G. fruticosus harvested in Namibia was
investigated using thin layer chromatography and Gas Chromatography-Mass
Spectroscopy. The anticancer activity was evaluated against three cells lines (PC3, I leLa and HcpG2) and cytotoxicity against primary cell lines (Hck293 and KMST6). Apoptosis induction was investigated using the APOPercentageru assay, while general oxidative stress and caspase induction was investigated using the Reactive oxygen species (ROS) assay in HcLa and Hek293 cel ls. The toxicity of the G. fruticosus extracts was also assessed using in vivo toxicity assays in Balb/C mice. Finally, to improve solubility of phytochemicals and reduce non-specific toxicity, a lipid based delivery system was fonnulated through thin-layer hydration of phosphatidylcholine, cholesterol and plant extract (250: J: I 0) in phosphate buffered sal inc The nanoparticlcs were characterized for physically for zeta potential and polydispcrsity using Dynamic Light Scattering and Scanning Electron Microscopy. The encapsulation efficiency of the drug delivery system and its stabi lity in media was also investigated before its biological activity was evaluated
against the lleLa cervical cancer cell line. Phytochemical analysis showed G. fruticosus had alkaloids, coumarins, steroids, anthraquinones, Oavonoids and triterpenoids. Furthermore, anti-inflammatory, anticancer and antioxidant compounds such as betasitosterol, alpha-amyrin, lupeol and their esters and acetates were detected by GCMS. The plant extract showed selective antiproliferativc activity against HeLa cel ls (ICso 8.35"7 ± 0.13 g/L) although the mechanism of cell dcalh was most likely necrosis not apoptosis as expected. This conclusion was supported by detected ROS levels and lack of caspase
cleavage. In vivo toxicity in Balb C mice was observed (LDso 1.968"1 ± 59.16 glkg) and classified as category 3 under GHS. Lipid based nanoparticles showed high encapsulation efficiency of 96.5 I % with a particle size range between 1 30-150 nm and narrow particle size spread. The zeta potential was low at -1.6 ± 0.6 mV, contributing to agglomeration and low stabi lity in biological media. Fluorescent microscopy and flow cytometry analysis showed uptake of G. fruticosus phytosomes based on the rhodamine fluorescence, treatment of HeLa cells with phytosomes (3.976"4 g/L), increased apoptosis. Cells also had reduced viability in a short time space after exposure, alluding to accelerated cell death in comparison to free extract. This study raises the possibility that the toxicity of G. fruticosus can be managed using a lipid based delivery system especially for cervical cancer following optimization in in vivo cancer disease models. It validates the ethnomedicinal use of G. fruticosus and adds value to a plant whose development was previously abandonded due to toxicity. Furthermore, it elucidates the mechanism of action
of G. fruticosus extracts which was not previously known, while describing the
phytochemistry of Namibian G. fruticosus. This study can lead to the development of commercially viable, safe and locally sourced treatment alternatives, especially for cervical cancer.
Description
A dissertation submitted in fulfillment of the requirements for the Degree of Doctor of Philosophy in Science (Biological Sciences)
Keywords
Anticancer, Gomphocarpus Fruticosus, LIPID-based nano-particles