A dissertation submitted to the Faculty of Health Sciences, University of
the Witwatersrand, Johannesburg, in fulfillment of the requirements for
the degree of Masters of Science in Medicine (Pharmacology)
Johannesburg,2015 / Breast cancer, is the most frequently diagnosed cancer in women and is associated with
high mortality rates in South Africa. There is a high prevalence of metastatic breast
cancer and triple negative tumours, which are associated with poor prognosis. In this
study, the response of two breast cancer cell lines, MCF-7 and MDA-MB-231, were
evaluated when treated with novel imidazo[1 ,2-a]pyridine amines.
The compounds were synthesized by the School of Chemistry of the University of the
Witwatersrand using the Groebke-Blackburn-Bienayme multicomponent reaction and
tested for purity by elemental analysis. The MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-
diphenyltetrazolium bromide) assay was used to determine the cytotoxic effects of test
compounds on breast cancer cells and the toxic effect of compounds on non-tumorigenic
unstimulated peripheral leukocytes. IC50 values of test compounds were calculated from
sigmoidal dose response curves. The morphology of cells exposed to test compounds
was assessed by fluorescent microscopy using Hoechst 33342, acridine orange and
ethidium bromide. The ability of test compounds to induce apoptosis was measured by a
colorimetric caspase-3 assay and a fluorometric Annexin-V-FITC assay.
Monodansylcadaverine was used to determine if autophagic vacuoles were formed after
exposure to test compounds.
Three imidazo[1,2-a]pyridine amines, JD88, JD253 and JD256, were more cytotoxic to
MCF-7 than to MDA-MB-231 cells. MCF-7 cells showed morphological features
associated with apoptosis, and proteolysis by caspase-3/7 was observed after MCF-7
cells were exposed to JD88 for two hours. Vacuole formation induced by these
compounds was not autophagic in since they did not co-localize with MDC florescent
clusters. This together with the exposure of phosphatidylserine to the outer surface of
MCF-7 cells suggests that apoptosis is induced in these cells. There was no evidence of
cytochrome c translocation to the cytoplasm, which indicates that the intrinsic pathway of
apoptosis is not activated. MDA-MB-231 cells treated with JD88, JD253 and JD256 were
large with multiple nuclei and decondensed chromatin, morphological features
associated with mitotic catastrophe. The cells also showed morphological features
associated with necrosis and apoptosis, which include loss of cell membrane integrity
and cell membrane blebbing respectively. MDA-MB-231 cells exposed to JD88 showed
marked exposure of phosphatidylserine and this was observed to a minor extent in cells
exposed to JD253 and JD256. Proteolysis by caspase-3/7 was activated in MDA-MB-231
cells exposed to JD88 as early as 2 hours after exposure.
In conclusion three compounds; JD88, JD253 and JD256 were able to induce apoptosis
in MCF-7 cells. These compounds were selectively toxic against MCF-7 cells compared
to MDA-MB-231 cells and JD256 in particular was less toxic to leukocytes, which may
translate to fewer serious adverse effects. Addition of a copper dioxygen complex to
these compounds increases activity against both breast cancer cells. JD88 in particular
has shown effective induction of apoptosis and this merits further investigation into its
potential as a lead compound in breast cancer therapy. / AC2016
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:wits/oai:wiredspace.wits.ac.za:10539/21597 |
| Date | January 2015 |
| Creators | Kurebwa, Taurai, Flloyd. |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | application/pdf |
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