Chu, Jessica Hoi-Yan
(2014)
Investigating in vitro anticancer properties of Malaysian rainforest plants: Acalypha wilkesiana Müll, Arg. Archidendron ellipticum (Blume) Hassk. Duabanga grandiflora Walp. Pseuduvaria macrophylla (Oliv.) Merr.
PhD thesis, University of Nottingham.
Abstract
Malaysia is ranked the 12th richest country for its biological diversity of plant species by the Convention on Biological Diversity and this project aims to contribute to existing knowledge of Malaysian rainforest plants, Acalypha wilkesiana, Duabanga grandiflora, Archidendron ellipticum and Pseuduvaria macrophylla, for the treatment of cancer.
A. wilkesiana (Euphorbiaceae) whole plant EtOH and EtOAc extracts inhibited growth of breast cancer MDA-MB-468 cells (GI50: 22.7 and 15.9 μg/ml) and revealed preference over non-transformed MRC5 fibroblasts (GI50: 46.6 and 53.3 μg/ml, respectively). EtOH and HEX extracts were able to impair cell survival and colony-forming abilities in MDA-MB-468 cells after 24 h. Detection of increased MDA-MB-468 sub-G1 cell populations after 48 h treatment to EtOH and HEX extracts, suggest that cells may be undergoing apoptosis.
A. ellipticum (Leguminosae) crude polar bark and leaf extracts inhibited MDA-MB-468 cell growth (GI50 of bark EtOH, EtOAc extracts: 1.7 and 40.4 μg/ml, respectively and leaf EtOH and EtOAc extracts: 9.3 and 9.3 μg/ml, respectively). However, MDA-MB-468 cell growth was unaffected by HEX extracts (> 200 μg/ml). Separation of crude extracts revealed sub-fractions of greater activity, in particular a 50-fold enhanced potency in sub-fractions of HEX extract thus overcoming masking or antagonistic activity in the crude mixture. Following 24 h, bark and leaf extracts impaired MDA-MB-468 cells’ proliferative and colony-forming abilities at 1X and 2X GI50 values suggesting significant damage was induced leading to observed cellular senescence and inhibition of cell proliferation. After 48 h exposure to EtOH and HEX extracts, MDA-MB-468 cells accumulated cellular damage, possibly affecting microtubule functions resulted in activation of apoptosis as shown by increased of sub-G1 and G2/M MDA-MB-468 cell populations and presence of phosphatidyl serine on the outer membrane of cells. Modest levels of flavonoid and phenolic compounds were found in bark and leaf extracts, which correlated to moderate level of free radical scavenging activity observed.
D. grandiflora (Lythraceae) bark and leaf extracts revealed growth inhibitory effects against colorectal cancer HCT116 cells (GI50 of bark Water, EtOH, EtOAc and HEX extracts: 42.4, 37.5, 21.69 and 28.9 μg/ml, respectively; leaf Water, EtOH, EtOAc and HEX extracts GI50: 38.0, 40.9, 24.7 and > 200 μg/ml, respectively). Separation of crude bark extracts resulted in fractions of greater activity, whereas separation of leaf extracts revealed reduced activity suggesting synergistic activity in the mixture. Following 24 h, bark and leaf extracts impaired HCT116 cells’ proliferative and colony-forming abilities at 1X and 2X GI50 values indicating significant damage incurred leading to observed cellular senescence and cell proliferation inhibition. After 48 h exposure to D. grandiflora extracts, increased sub-G1 HCT116 cell population and a G1/0 cell cycle block accompanied by decreased S and G2/M cell populations were measured. Detection of phosphatidyl serine on cells’ outer membrane and activated apoptotic caspase 3 protein confirmed D. grandiflora extracts induced apoptosis. Highest levels of flavonoid and phenolic compounds were found in polar bark and leaf D. grandiflora extracts, which may correlate to the highest free radical scavenging activity measured.
P. macrophylla (Annonaceae) extracts collectively displayed greatest HCT116 cell growth inhibition (EtOH, EtOAc and HEX extracts GI50: 5.2, 1.6 and 5.4 μg/ml) and separation of EtOH and EtOAc extracts revealed even greater activity in sub-fractions. After 24 h, polar extracts at 2X GI50 completely impaired HCT116 cells’ proliferative and colony-forming abilities suggesting extract-induced significant damage led to inability of cells to proliferate. Analysis of cell cycle distribution revealed increased sub-G1 HCT116 cell population and high levels of early apoptotic HCT116 cells following 48 h exposure to P. macrophylla extracts coupled with detection of caspase 3 activation confirming execution of apoptosis. Modest levels of flavonoid and phenolic compounds were detected in EtOH, EtOAc and HEX extracts, which correlated to modest free radical scavenging activity.
The findings in this project should justify further separation and in vitro investigations.
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