In vitro effects of an in silico-modelled 17β-estradiol derivative in combination with dichloroacetic acid on MCF-7 and MCF-12A

OBJECTIVES : To investigate anti-proliferative properties of a novel in silico-modelled 17β-oestradiol derivative (C9), in combination with dichloroacetic acid (DCA), on MCF-7 and MCF-12A cells. MATERIALS AND METHODS : xCELLigence system was employed to determine optimal seeding number for cells, and crystal violet assay was used to assess cell number and to determine IC50 value (24 h) for combination treatment. Light and fluorescent microscopy techniques were used to morphologically detect types of cell death. Flow cytometry was used to analyse cell cycle and apoptosis. RESULTS : Optimal seeding number for 96-well plates was determined to be 5000–10 000 cells ⁄ well for both MCF-7 and MCF-12A cells. IC50 for MCF-7 cells of the combination treatment after 24 h was 130 nM of C9 in conjunction with 7.5 mM of DCA (P < 0.05). In contrast, the same concentration inhibited cell population growth by only 29.3% for MCF- 12As after 24-h treatment (P < 0.05). Morphological studies revealed lower cell density of both types of combination-treated cells. Flow cytometric analyses demonstrated increase in sub-G1 phase in combination- treated MCF-7 cells. CONCLUSIONS : These results demonstrate that the novel 17β-oestradiol derivative C9, in combination with DCA is a potent anti-proliferation treatment, with properties of selectivity towards tumourigenic cells. Thus, this warrants further studies as a potential combination chemotherapeutic agent for further cancer cell lines.This research was supported by grants from the Medical Research Council of South Africa (AL343, AS536), the Cancer Association of South Africa (AS201), the National Research Foundation (NRF) (AL239), the RESCOM of University of Pretoria and the Struwig-Germeshuysen Cancer Research Trust of South Africa (AN074).http://onlinelibrary.wiley.com/journal/10.1111/(ISSN)1365-2184

In vitro evaluation of a novel antimitotic estradiol analog and dichloroacetic acid on breast adenocarcinoma and breast non-tumorigenic cells

In vitro evaluation of a novel antimitotic estradiol analog and dichloroacetae, which restore mitochondrion function in breast adenocarcinoma and breast non-tumorigenic cells, will contribute to the field of combination therapy in cancer.This paper was initially delivered at the Annual Congress of the Biological Sciences Division of the South African Academy for Science and Art, ARC-Plant Protection Research Institute, Roodeplaat, Pretoria, South Africa on 01 October 2010.am201

Docking, synthesis, and in vitro evaluation of antimitotic estrone analogs

In the present study, Autodock 4.0 was employed to discover potential carbonic anhydrase IX inhibitors that are able to interfere with microtubule dynamics by binding to the Colchicine binding site of tubulin. Modifications at position 2’ of estrone were made to include moieties that are known to improve the antimitotic activity of estradiol analogs. 2-ethyl-3-O-sulphamoyl-estra-1,3,5(10),15-tetraen-3-ol-17-one estronem (C9) and 2-ethyl-3-O-sulphamoyl-estra-1,3,5(10)16-tetraene (C12) were synthesized and tested in vitro. Growth studies were conducted utilizing spectrophotometrical analysis with crystal violet as DNA stain. Compounds C9 and C12 were cytotoxic in MCF-7 and MDA-MB-231 tumorigenic and metastatic breast cancer cells, SNO non-keratinizing squamous epithelium cancer cells and HeLa cells after 48 h exposure. Compounds C9 inhibited cell proliferation to 50% of the vehicle-treated controls from 110-160 nM and C12 at concentrations ranging from 180-220 nM. Confocal microscopy revealed abnormal spindle morphology in mitotic cells. Cell cycle analysis showed an increase in the number of cells in the G2/M fraction after 24 h and an increase in the number of cell in the sub-G1 fraction after 48 h, indicating that the compounds are antimitotic and able to induce apoptosis.Medical Research Council of South Africa (AG374, AK076), the Cancer Association of South Africa (AK246), the Struwig-Germeshuysen Cancer Research Trust of South Africa (AJ038) and RESCOM University of Pretoria (A0R984).http://www3.interscience.wiley.com/journal/120118371/grouphome/home.htm

Cytotoxic activity of pentachlorophenol and its active metabolites in SH-SY5Y neuroblastoma cells

As knowledge regarding mechanisms of pentachlorophenol (PCP) toxicity in neuronal cell lines is limited, the aim of the study was to evaluate the effects of PCP and its active metabolites, tetrachloro-1,4-benzoquinone (TCBQ) and tetrachlorohydroquinone (TCHQ) in human neuroblastoma SH-SY5Y cells. All compounds induced cytotoxic effects in time- and dose-dependent manners, and resulted in differential modes of cell death. Reduced mitochondrial membrane potential (ΔᴪM) and oxidative damage lead to apoptosis and necrosis following TCBQ and PCP exposure, respectively. Time-dependent investigations revealed transient ΔᴪM recovery in TCHQ exposed cells, and redox stress. Sufficient ΔᴪM recovery allowed apoptosis completion in TCHQ exposed cells, whereas overwhelming metabolic and oxidative stress saw a conversion from apoptotic to necrotic-like cell death. The onset of mitochondrial dysfunction preceded that of redox damage for all compounds, indicating that oxidative damage is secondary to ΔᴪM insult. Cytotoxic events were further linked to cell cycle. S phase and G2/M blocks were observed after 12 h exposure to TCBQ and TCHQ, respectively, while a G1 block occurred after 24 h exposure to PCP. This study provides new insight regarding time-dependant toxic effects of PCP and its metabolites in human neuronal cells.The National Research Foundation, South Africa and Research Committee of the School of Medicine, Faculty of Health Sciences, University of Pretoria.http://www.elsevier.com/locate/tiv2020-08-01hj2020PharmacologyPhysiolog

2-Methoxyestradiol-3,17,0,0-bis-sulfamate (2MEBM) induces two types of cell death in a cervical adenocarcinoma cell line

2-MEBM induced apoptosis and autophagy in HeLa cells and also disrupted the cellular microtubule network. This contributes to the understanding of the in vitro mechanism of action of 2MEBM as a potential anticancer drug.This paper was initially delivered at the Annual Congress of the Biological Sciences Division of the South African Academy for Science and Art, ARC-Plant Protection Research Institute, Roodeplaat, Pretoria, South Africa on 01 October 2010.http://www.satnt.ac.zaam2014ay201

Flow cytometric comparison of platelets from a whole blood and finger-prick sample : impact of 24 hours storage

In this study, we investigate the validity and laboratory utility of flow cytometry when analyzing platelet activation by studying CD41, CD42b, CD62P and CD63. We compare flow cytometry results from citrated whole blood and finger-prick samples directly after collection and also after storing both a finger-prick and whole blood sample for 24 hours. Citrated whole blood and finger-prick samples were taken from three healthy individuals on two occasions, and a total of 60 000 cells were analyzed for each of the 4 phycoerythrin-labelled monoclonal antibodies. Half of each sample was analyzed immediately after sampling while the other half was kept in the fridge at 6°C for 24 hours before analysis. No significant difference was found between the sampling methods or the period of time before analysis. Results therefore suggest that an appropriately prepared finger-prick sample can be used for platelet function analysis, and samples can be stored for 24 hours in the fridge at 6°C before analysis.http://www.maneyonline.com/loi/hemhb2016Physiolog

In vitro effects of a new and novel antimitotic compound in human breast adenocarcinoma metastatic epithelial cells

This study contributes to the understanding of molecular mechanisms and cell signaling events associated with in vitro anticancer responses of a new and novel antimitotic compound, 2-etiel-3-sulfamaat- 1,3,5(10)16-tetraeen (C19).This paper was initially delivered at the Annual Congress of the Biological Sciences Division of the South African Academy for Science and Art, ARC-Plant Protection Research Institute, Roodeplaat, Pretoria, South Africa on 01 October 2010.http://www.satnt.ac.zaam201

Differential signaling involved in Sutherlandia frutescens-induced cell death in MCF-7 and MCF-12A cells

Ethnopharmacological relevance: The scientific study of natural products traditionally used in anticancer preparations has yielded several therapeutically relevant compounds. One of these traditional preparations with potentially beneficial properties is aqueous extracts of Sutherlandia frutescens, a shrub indigenous to the Western Cape region of South Africa. Aims of the study: To evaluate in vitro efficacy of these preparations on the MCF-7 breast adenocarcinoma and MCF-12A non-tumorigenic cell lines in terms of cell proliferation, cell morphology and possible induction of cell death. Materials and Methods: Crystal violet staining was used to evaluate cell proliferation, lightand fluorescence microscopy were used to investigate both intracellular and extracellular morphological features of apoptosis and autophagy (e.g. membrane blebbing, condensed chromatin and intracellular lysosomes), while flow cytometry quantified cell cycle changes and induction of apoptosis through analysis of the flip-flop translocation of phosphatidylserine.The Medical Research Council of South Africa (AK076; AL343). The Cancer Association of South Africa (AK246).The National Research Foundation of South Africa (AL239) and the Struwig-Germeshuysen Cancer Research Trust of South Africa (AN074).http://www.elsevier.com/locate/jethpharmhb201

A combination of an antimitotic and a bromodomain 4 inhibitor synergistically inhibits the metastatic MDA-MB-231 breast cancer cell line

CITATION: Mqoco, T., et al. 2019. A combination of an antimitotic and a bromodomain 4 inhibitor synergistically inhibits the metastatic MDA-MB-231 breast cancer cell line. BioMed Research International, 2019(1850462):1-13. doi:10.1155/2019/1850462The original publication is available at https://www.hindawi.com/journals/bmri/Current chemotherapeutic agents have many side effects and are toxic to normal cells, providing impetus to identify agents that can effectively eliminate tumorigenic cells without damaging healthy cells. The aim of this study was to examine whether combining a novel BRD4 inhibitor, ITH-47, with the antimitotic estradiol analogue, ESE-15-ol, would have a synergistic effect on inhibiting the growth of two different breast cancer cell lines in vitro. Our docking and molecular dynamics studies showed that compared to JQ1, ITH-47 showed a similar binding mode with hydrogen bonds forming between the ligand nitrogens of the pyrazole, ASN99, and water of the BRD4 protein. Data from cell growth studies revealed that the GI50 of ITH-47 and ESE-15-ol after 48 hours of exposure was determined to be 15 μM and 70 nM, respectively, in metastatic MDA-MB-231 breast cancer cells. In tumorigenic MCF-7 breast cancer cells, the GI50 of ITH-47 and ESE-15-ol was 75 μM and 60 nM, respectively, after 48 hours of exposure. Furthermore, the combination of 7.5 μM and 14 nM of ITH-47 and ESE-15-ol, respectively, resulted in 50% growth inhibition of MDA-MB-231 cells resulting in a synergistic combination index (CI) of 0.7. Flow cytometry studies revealed that, compared to the control, combination-treated MDA-MB-231 cells had significantly more cells present in the sub-G1 phase and the combination treatment induced apoptosis in the MDA-MB-231 cells. Compared to vehicle-treated cells, the combination-treated cells showed decreased levels of the BRD4, as well as c-Myc protein after 48 hours of exposure. In combination, the selective BRD4 inhibitor, ITH-47, and ESE-15-ol synergistically inhibited the growth of MDA-MB-231 breast cancer cells, but not of the MCF-7 cell line. This study provides evidence that resistance to BRD4 inhibitors may be overcome by combining inhibitors with other compounds, which may have treatment potential for hormone-independent breast cancers.NRF Thuthuka grantCancer Association of South AfricaMedical Research CouncilSchool of Medicine Research CommitteeStruwig Germeshuysen Trusthttps://www.hindawi.com/journals/bmri/2019/1850462/Publisher’s versio

Short communication : effects of a 17-beta estradiol analogue on gene expression and morphology in a breast epithelial adenocarcinoma cell line : a potential antiproliferative agent

2-Methoxyestradiol is a 17-beta estradiol derivative with antitumor activity. Due to low bioavailability, a new estrogen analogue was in silico-designed. The latter, a sulphamoylated analogue of 2-methoxyestradiol, was evaluated for its potential antiproliferative effect by means of expression microarray analysis and transmission electron microscopy in the highly metastatic breast adenocarcinoma (MDA-MB-231) cell line. Data indicate changes in gene expression pertaining to induction of apoptosis and autophagy as types of cell death, arrest of the cell cycle, and impairment of the cytoskeleton and induction of reactive oxygen species. Transmission electron microscopy confirmed morphological characteristics of apoptosis and autophagy. Altered cell cycle- and cytoskeletal-related gene expression profiles demonstrated that the estradiol analogue acts as an antimitotic agent in this highly metastatic breast cell line. Data also showed that the newly designed estrogen analogue exerts an antiproliferative effect in this cancer cell line culminating in both apoptosis and autophagy as type of cell death paving the way for further investigations into its potential as anticancer agent.Cancer Association of South Africa, the Struwig Germeshuysen Trust, RESCOM (Research Council of the University of Pretoria), National Research Foundation and The Medical Research Council.http://www.biomedres.infohb201