Assessment of cytotoxic artemisinin and its derivatives as DNA damaging inducing agents in triple-negative breast cancer cells
- Authors: Mkhwanazi, Ntando
- Date: 2022-10-14
- Subjects: Breast Cancer , Artemisinin , DNA damage , Antineoplastic agents , Breast Cancer Treatment
- Language: English
- Type: Academic theses , Master's theses , text
- Identifier: http://hdl.handle.net/10962/362960 , vital:65378
- Description: In developing countries, including South Africa, breast cancer is the primary cause of cancer-related deaths among women. TNBC (triple-negative breast cancer) is an aggressive breast cancer subtype that is more prevalent in women of African descent. This subtype lacks the key receptors, namely the estrogen receptor (ER-), progesterone receptor (PR-), and human epidermal growth factor receptor 2 (HER2-) that are the basis of successful targeted therapies for other subtypes of the disease. To date, there are no effective, standardized targeted therapies for TNBC. Artemisinin is an anti-malarial drug and numerous derivatives of the compound have been developed to improve the potency and solubility of the parent compound. Artemisinin and its derivatives have gained attention as potential anti-cancer agents; however, such studies have not yet progressed to clinical trials and the precise mechanism of action of these compounds is yet to be fully explained. In this study, artemisinin, and its known derivative artesunate, as well as a novel derivative, WHN11, were investigated as DNA damage-inducing agents in TNBC. WHN11 was found to be the most potent of the three compounds, displaying an IC50 of 3.20 μM against HCC70 cells, artemisinin displayed an IC50 of 214.70 μM and artesunate displayed an IC50 of 25.48 μM. The compounds were less toxic to the MCF12A non-cancerous cells, with IC50 values 298.30, 87.53, and 8.35 μM for artemisinin, artesunate, and WHN11, respectively, and displayed selectivity indices of 1.39, 3.44 and 2.61 μM for artemisinin, artesunate, and WHN11, respectively. In silico and in vitro studies revealed that the artemisinin compounds bind to DNA through the minor groove. While all three compounds were able to bind to DNA, a comet assay revealed that only artemisinin and artesunate, and not WHN11, were able to cause DNA damage compared to the vehicle control, DMSO. Finally, a topoisomerase I (TOPO I) enzyme assay demonstrated that while the compounds appeared to display a degree of inhibition of TOPO I, as evidenced by a downward shift in the plasmid band on the agarose gel, they were not able to fully inhibit the enzyme to return the plasmid to the supercoiled conformation. In addition, combination studies revealed that artemisinin, artesunate, and WHN11 acted synergistically in combination with camptothecin, but displayed either an additive (artemisinin) or antagonistic (artesunate and WHN11) relationship when used in combination with etoposide. In conclusion, artemisinin, its known derivative artesunate, and novel and highly toxic derivative WHN11, all bind to DNA via the minor groove, however only artemisinin and artesunate, and not WHN11, cause DNA damage, indicating a potentially different mechanism of action of the three artemisinins. All three compounds act synergistically with camptothecin, which suggests interference with topoisomerase activity, partially supported by slight inhibition of TOPO I activity, and could indicate either direct inhibition of the enzyme or interference with enzyme function by competitive binding to the DNA. Further studies could help explore alternate DNA damage assays, to validate these findings, and the effect of the compounds on TOPO II activity could also be assessed. , Thesis (MSc) -- Faculty of Science, Biochemistry and Microbiology, 2022
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- Date Issued: 2022-10-14
An evaluation of the cytotoxic activities of novel artemisinin derivatives: towards targeted therapies for triple-negative breast cancers (TNBC)
- Authors: Kajewole, Deborah Ifeoluwa
- Date: 2020
- Subjects: Uncatalogued
- Language: English
- Type: thesis , text , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/163329 , vital:41029 , doi:10.21504/10962/163329
- Description: Thesis (PhD)--Rhodes University, Faculty of Science, Biochemistry and Microbiology, 2020.
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- Date Issued: 2020
Identification of potential novel roles for Hsp70/Hsp90 organising protein (Hop) using proteomic analysis in human cells
- Authors: Wingate, Ianthe
- Date: 2016
- Language: English
- Type: text , Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/64758 , vital:28598
- Description: Expected release date-May 2018
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- Date Issued: 2016
The effect of extracellular Hsp90β and TGF-β1 on colon cancer biology
- Authors: Perks, Tamarin
- Date: 2015
- Language: English
- Type: text , Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/55896 , vital:26753
- Description: The TGF-β signaling pathway is known to be one of the most commonly mutated pathways in human cancers, while Hsp90 is a bone fide drug target that is involved in regulating the conformation and activity of many oncoproteins. The role of intracellular Hsp90 in cancer has thus far been established and there is a growing link between extracellular Hsp90 and cancer metastasis, as well as the role of TGF-β in metastasis. This study aimed to analyse the interaction between Hsp90 (both intracellular and extracellular) and the TGF-β machinery in cancer cells, as well as to determine the effect of these proteins on cellular responses on the biology of cancer cells. This was achieved by studying the expression of Hsp90; TGF-βRII and TGF-β1 in cancer cell lines of various origins using flow cytometry, ELISA, and western blot analysis. The genetically paired SW480 and SW620 colon cancer cell lines, derived from a primary tumour and lymph node metastasis, respectively, were selected for further study due to differences in expression levels and activation of the TGF-β1 pathway. SW480 cells expressed double the level of TGF-βRII compared to SW620 cells, while SW620 expressed two times more extracellular TGF-β1 than SW480 cells. A direct interaction between TGF-β1 and Hsp90β was determined in vitro, and confirmed in vivo in SW620 cells. Growth, adhesion and migration were analysed in SW480 and SW620 cells. SW480 cells adhered significantly faster than SW620 cells, while SW620 cells had a greater rate of migration. Inhibiting the TGF-β pathway, specifically TGF-βRI, using SB 431542, as well as inhibiting Hsp90 with novobiocin, caused an increase in migration in SW480 cells. Only the addition of TGF-β1 in combination with Hsp90 as well as SB 431542 caused an increase in migration in SW620 cells. The canonical TGF-β1/TGF-βRI/TGF-βRII pathway may be constitutively active in SW620 cells and the inhibition of TGF-βRI may suggest an alternate pathway or receptor in both SW480 and SW620 cells.
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- Date Issued: 2015