Browsing by Author "Hendricks, Denver"
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- ItemOpen AccessAltered protein expression patterns in oesophageal cancer(2009) Zemanay, Widaad; Hendricks, DenverOesophageal squamous cell carcinoma presents a significant health burden in South Africa. It is one of the most common causes of cancer-related mortality of South African black males, as a result of its asymptomatic progression leading to late diagnosis and poor prognosis. The aim of this study was to identify membrane or membrane-associated proteins that are expressed at different levels in oesophageal tumour tissue when compared to normal tissue. The identification of such proteins would be an important step towards the development of better diagnostic and therapeutic strategies for this disease. Two proteomic approaches, were employed to identify differentially expressed proteins.
- ItemOpen AccessAnalysis of driver gene mutations in oesophageal squamous cell carcinoma(2025) Shipanga, Hendrina Nelao Mwiiwete; Parker, Mohamed; Hendricks, DenverOesophageal cancer (OC) is the eleventh most diagnosed cancer and the seventh most common cause of cancer-related deaths worldwide. The two main subtypes are oesophageal adenocarcinoma (OAC) and oesophageal squamous cell carcinoma (OSCC). OAC is more common in North America and Europe, while OSCC predominantly occurs in Eastern Asia, Sub-Saharan Africa and Latin America. Over 80% of the OSCC cases and deaths worldwide occur in less developed regions, including Sub-Saharan Africa. The asymptomatic development of OSCC, results in late diagnosis of the disease with a poor prognosis, typically ranging from 5-10% at 5-year post-diagnosis in Africa. This study investigated the genomic landscape of OSCC in the South African population by whole-genome sequencing (WGS) and whole-exome sequencing (WES). Normal and tumour DNA and RNA was isolated from OSCC patient biopsies prior to the commencement of any form of chemotherapy or radiotherapy. WGS was performed on 31 samples, while WES was conducted on 67 samples. The mRNA levels of selected genes in OSCC were quantitated by RT-qPCR. KYSE30 cells were used for siRNA-mediated knockdown experiments targeting p14ARF and p16INK4a in OSCC. In silico structural analysis of missense mutations in p14ARF and p16INK4a was conducted using UCSF Chimera tool. WGS analysis identified 35 frequently mutated genes in OSCC, among these, TP53, CDKN2A.p16INK4a, CDKN2A.p14ARF, and KMT2D were identified as OSCC driver genes. Based on the mutation spectrum analysis, samples clustered into two distinct groups, cluster 1 and cluster 2b, characterized by TP53 alterations and mutation rates per megabase (Mb). WES expanded findings across 67 samples, identifying TP53, NFE2L2, CDKN2A.p16INK4a, ZNF750, and NOTCH1 as OSCC driver genes. Samples clustered into three groups: cluster 1, cluster 2a, and cluster 2b, expanding upon the two clusters identified in our WGS analysis. In both WGS and WES analyses, cluster 1 exhibited TP53 mutations and relatively high somatic mutation rates per Mb, while cluster 2 lacked TP53 mutations. Cluster 2 is further subdivided into clusters 2a and 2b in WES. Cluster 2a samples display a high mutation rate per Mb, while cluster 2b samples display fewer genomic alterations. By quantifying the contribution of the mutational signatures to the mutation spectrum, we found a relatively high contribution of mutation signature SBS1, SBS2, and SBS13, implicating aging and AID/APOBEC (activation induced cytidine deaminase/apolipoprotein B mRNA editing enzyme catalytic subunit) activation in OSCC tumourigenesis. WGS analysis revealed three novel mutational signatures that had not been previously identified. Interestingly, these signatures were not observed in the samples analysed by WES, even though the WES cohort included a larger sample size. The significance of these novel mutational signatures remains unclear. Evaluation of selected differentially expressed genes in OSCC involved in cell cycle control, the KEAP1-NFE2L2 (NRF2) pathway, and DNA damage response pathways showed variable expression of these genes in OSCC suggests potential dysregulation of the genes in OSCC. Furthermore, p16INK4a and p14ARF mRNA levels were significantly lower in 61% and 48% of OSCC tumour samples, respectively, while elevated levels were observed in 16% and 25% of tumours, respectively. Knockdown of p14ARF and p16INK4a in KYSE30 cells resulted in dysregulation of key regulators involved in multiple cancer signalling pathways, including cell cycle, apoptosis, and KEAP1-NFE2L2 pathways. This dysregulation could promote cell survival, growth of apoptosis-resistant cells, and resistance to stress, which are critical events in tumorigenesis. In-silico mutation analysis revealed damaging mutations in p16INK4a, such as p.A68V, p.D84N, p.D108H, p.D108N, p.D108Y, and p.L130P. These mutations cause significant structural alterations that disrupt interactions crucial for p16INK4a stability and function, possibly affecting cell cycle regulation and potentially promoting tumorigenesis in OSCC. Our findings highlights novel molecular features of OSCC and provides comprehensive insights into the genomic and molecular mechanisms driving OSCC within the South African population
- ItemOpen AccessAnti-oesophageal cancer activity in extracts of deep-water Marion Island sponges(2005) Davies-Coleman, Michael T; Froneman, William; Keyzers, Robert; Whibley, Catherine; Hendricks, Denver; Samaai, Toufiek; McQuaid, ChristopherOESOPHAGEAL CANCER IS ONE OF THE most common causes of cancer-related deaths in South African black males. The limited efficacy of chemotherapeutic agents to treat this disease has prompted a search for potential new chemical entities with anticancer properties. We report here on the evidence for anti-oesophageal cancer activity in the methanolic extracts of five species of sponges dredged from a depth of approximately 100 m in the vicinity of Marion Island in the Southern Ocean during the autumn of 2004.
- ItemOpen AccessCharacterization of ADAM10 in cervical and oesophageal cancers(2013) Williams, Cleo Julie-Jean; Leaner, Virna; Hendricks, DenverA Disintegrin And Metalloproteinase 10 (ADAM10) is a cell surface molecule that activates proteins such as: adhesion molecules, cytokines, and growth factors. ADAM10 has been associated with the proliferation and metastasis of cancers including, breast, lung, and colon. While ADAM10 has been investigated in these cancers, very little is known of its role in cervical and oesophageal cancer. The aim of this study was to investigate the endogenous expression of ADAM10 in cervical and oesophageal patient material and representative cell lines.
- ItemOpen AccessDifferentially expressed genes in oesophageal cancer : Retinoic acid receptor-ß2, Trio and Abl-related gene(2003) Hadley, Katie Emma; Hendricks, DenverBibliography: leaves 95-104.
- ItemOpen AccessExpression and regulation of N-Myc Downstream- Regulated gene 1 in squamous cell carcinoma of the oesphagus(2009) Bracher, Jacqueline Claire; Hendricks, DenverSquamous cell carcinoma of the oesophagus is a formidable disease which poses a significant health risk in developing countries where the incidence is frequently high and access to health care facilities is often limited. The identification of genes involved in oesophageal tumourigenesis may provide new targets for therapy and improved diagnostics techniques, thereby improving the prognosis of this pernicious disease. In this study, real-time RT-PCR and immunohistochemistry described the overexpression of N-Myc Downstream-Regulated Gene 1 (NDRG1) in oesophageal squamous cell carcinoma (OSCC) tissue compared to normal tissue in a cohort of South African cancer patients. Despite more than ten years of research into the role of NDRG1 in cancer, the precise function of this protein remains enigmatic. Reports have been contentious, suggesting both tumour suppressor and tumour promoter functions for NDRG1, implicating it in tumourigenic processes such as metastasis and angiogenesis. Our immunohistochemical analysis if NDRG1 expression in OSCC tissue and matched normal epithelium (n=83) showed that NDRG1 expression is elevated by 2.6-fold in cancer tissue compared to normal tissue. Moreover, the expression and localisation of NDRG1 appeared to track with epithelial cell maturation where basal cells of normal oesophageal epithelium displayed plasma membrane-associated NDRG1 while maturing cells were mostly positive for NDRG1 in the cytoplasm and nucleus. Likewise, NDRG1 displayed interesting patterns of localisation in tumour tissue of the xiii oesophagus. Dysplastic tissue and poorly differentiated tumour tissue stained positively for NDRG1 in the plasma membrane, while moderately and well differentiated tumours displayed mixed staining for NDRG1 in the plasma membrane, cytoplasm and nucleus. Analysis of NDRG1 expression in cell lines cultured under anchorage-independent conditions revealed that NDRG1 expression is strongly induced when cells are prevented from adhering to the surface of culture dishes. Induced NDRG1 expression correlated inversely with mRNA expression of invasion genes, MMP-2 and MMP-9, as well as the mRNA expression of angiogenic factors Ang- 1, PDGF-B and VEGF-C but, in contrast, showed positive correlation with the angiogenesis cytokine, VEGF-A. Knock-down of NDRG1 expression with siRNA had no effect on anchorage-independent cell proliferation or apoptosis but did inhibit VEGF-A expression. Moreover, VEGF-A promoter activity, induced by culturing cells under anchorage-independent conditions was shown to be NDRG1-dependent. In order to identify factors that may drive NDRG1 transcription in cultured OSCC cell lines we cloned and partly characterised the NDRG1 promoter. Through the generation of promoter deletion constructs, site-directed mutagenesis and Chromatin Immunoprecipitation (ChIP) assays, we showed that both EGR-1 and cJun/AP-1 are capable of driving transcription of NDRG1 in response to 12-o-tetradecanoylphorbol- 13-acetate (TPA) through activation of PKC/MEK/ERK1/2 and JNK MAPK pathways. Taken together, we describe the regulation of NDRG1 expression by EGR-1 and AP-1 and we show that NDRG1 is overexpressed in squamous cell carcinoma of the oesophagus compared to normal oesophageal tissue. We associate NDRG1 with an xiv oncogenic function in OSCC through its potential role in angiogenesis via modulation of VEGF-A expression.
- ItemOpen AccessFunctional analysis of N-MYC downstream regulated gene 1 (NDRG1) in Oesophageal squamous cell carcinoma(2009) Wei, Wei; Hendricks, DenverOesophageal squamous cell carcinoma (OSCC) ranks as one of the deadliest tumours with a high incidence in developing countries in the areas of Southern Africa, Middle East and Far East. Moreover, its unfavourable prognosis is further complicated by the lack of knowledge about the molecular biology of this disease. In this thesis, we describe our work analysing the function of N-myc downstream regulated gene 1 (NDRG1, also known as Cap43 or Drg-1) in the neoplastic progression and maintenance of OSCC. Although NDRG1 has previously been implicated in breast, prostate, colon and liver carcinoma, the exact role of NDRG1 in OSCC still remains unclear. According to the immunohistochemical analysis of clinical OSCC tissue samples (n=52), NDRG1 expression was gradually increased in tumour tissue versus normal, indicating the potential involvement of NDRG1 in the neoplastic progression of OSCC. We next performed ectopic NDRG1 gain-of-function and loss-of-function studies using transfectants established from transduced OSCC cell lines (KYSE30 and KYSE150) by lentiviral vector mediated gene delivery. In KYSE30 cells, although no substantial effects on in vitro cell proliferation and differentiation were observed with altered NDRG1 expression, the ectopic overexpression of NDRG1 was found to be positively linked to metastasis, angiogenesis and apoptotic evasion as measured in cell culture. Accordingly, in the nude mouse xenograft model system, NDRG1 overexpression promoted the in vivo growth and metastasis of KYSE30 derived xenografts, which could be attributed to the reduced apoptotic and enhanced angiogenic activities promoted by this gene. Nevertheless, no significant phenotypic changes were observed in response to NDRG1 knock-down, suggesting that this gene was not essential for the neoplastic progression of OSCC. Moreover, null effect of either ectopic NDRG1 overexpression or knock-down were observed in KYSE150 cells, indicating ix that the function of NDRG1 may be largely dependent on the cellular context (Chapter 2). In addition to direct functional assays, evidence from analysing the regulation pattern of NDRG1 in OSCC cells was also presented to provide clues to indirectly predict the function of NDRG1 in OSCC. In Chapter 3, we demonstrated that NDRG1 could be actively regulated by various oncogenic stimuli such as cellular stress (genotoxicity and hypoxia) and mitogenic factors (EGF and IGF). Although these oncogenic regulatory effects on NDRG1 expression in OSCC cells may be dichotomous, the functional significance of NDRG1 upregulation, especially by hypoxia and EGF signalling, is highlighted. In our studies, the regulatory pattern of NDRG1 in OSCC is highly consistent with its oncogenic function revealed in ectopic studies (Chapter 2), further suggesting that phenotypic changes observed in the functional studies may not be artifactual, but may reflect the role of NDRG1 in the neoplastic progression of OSCC in physiological conditions. Taken together, our current data implicate NDRG1 as an effective but non-essential promoter in the neoplastic progression of oesophageal squamous cell carcinoma. Although the mechanism still needed to be further explored, our study suggests important clues regarding these mechanistic roles considering the impact of this gene on apoptosis, metastasis and angiogenesis.
- ItemOpen AccessInvestigating the Role of Cigarette Smoke and Alcohol in Oesophageal Squamous Cell Carcinoma(2022) Gamieldien, Raabie'ah; Hendricks, Denver; Parker, IqbalThe World Health Organisation (WHO) has estimated cancer to be one of the leading causes of death to people under the age of 70. Every year there are about 18.1 million new cancer cases worldwide; oesophageal cancer (OC) ranks seventh out of all incidence cases and sixth in mortality. OC has a poor 5-year overall survival rate of about a 15% (Arnal et al., 2015). This occurs as OC is largely asymptomatic and patients often seek medical assistance at a late stage of their cancer. This late diagnosis and a lack of efficient treatment has rendered OC a serious world health problem. Oesophageal Squamous Cell Carcinoma (OSCC) is the more common subtype of and accounts for about 90% of incident oesophageal cancers every year (Abnet et al., 2018) with the majority of OSCC cases occur in developing countries. The objective of this study was to investigate cigarette smoking and alcohol consumption as they have widely been reported as risk factors for OSCC. The project explored the impact of cigarette smoke condensate (CSC) treatment and EtOH on the expression of genes in cultured oesophageal cancer cells. Prior to treating the cells for expression analysis, cytotoxicity experiments were conducted to determine treatment conditions of CSC and EtOH which were sub-lethal, for the cell types and timeframes investigated. It was found that concentrations of 40 µg/ml CSC and 50 mM EtOH did not cause cell death for the time period of three days. Furthermore, we also showed that cell viability was maintained up to 10 days of treatment. RNA-sequencing then revealed a wide variety of genes that were differentially expressed in the OSCC cells treated with these selected concentrations of CSC and EtOH. One gene found to be differentially expressed in two RNA-Seq analyses was confirmed to be upregulated by RT-qPCR. Seven genes, AHRR, ALDH3A1, CYP1A1, CYP1B1, GSTM3, GSTM4 and UGT1 A6, involved in xenobiotic metabolism and a number of other metabolic pathways were also altered in response to CSC and EtOH treatment. However, these genes' involvement require further confirmation by RT-qPCR. This result of this study confirms that we have designed a reliable experimental system to investigate the role of EtOH and CSC in the development of OSCC. These results gave us a deeper insight into the genes and pathways affected by CSC and EtOH which may contribute to OSCC. Additionally, the cytotoxicity data can be use for future experimental work and the RNA-seq data can be used for further investigation into the development of OSCC.
- ItemOpen AccessInvestigating the role of growth hormone receptor in oesophageal squamous-cell carcinoma.(2013) Krivochiev, Boris; Hendricks, Denver; Hadley, KateSquamous-cell carcinoma of the oesophagus is a formidable disease which poses a significant health risk in developing countries where incidence is high and survival is low. Investigating the poorly understood mechanisms involved in oesophageal tumourigenesis may provide a platform to develop improved diagnostic techniques and therapies. The growth hormone (GH) signalling axis is important for proper cellular and organ system function. The axis has been shown to play a role in a number of cancers.
- ItemOpen AccessAn investigation into the molecular mechanisms induced by derivatives of natural products in oesophageal cancer(2014) Shunmoogam-Gounden, Nelusha; Hendricks, DenverCurrent chemotherapies for oesophageal cancer display poor efficacy and tolerability, highlighting an unmet need for novel chemotherapeutic agents. Artemisinin derivatives, currently used to treat malaria, were recently shown to possess potent anticancer activity. This study investigated the potential of two first generation artemisinin derivatives (artesunate and dihydroartemisinin), together with novel artemisinin hybrid compounds, as cancer chemotherapeutic agents and explored the mechanism of action in oesophageal cancer. Artesunate and dihydroartemisinin including seventeen other artemisinin derivatives were screened against oesophageal cancer cells using the 3 - [4,5-dimethylthiazol-2 -yl]-2,5 - diphenyltetrazolium bromide (MTT) assay and GraphPad Prism Software to calculate IC 50 (50% inhibitory concentration) values. Novel halogenated artemisinin - isatin hybrid compounds displayed the best activity against oesophageal cancer cells, and were more potent than artesunate and dihydroartemisinin in a small panel of oesophageal, breast and cervical cancer cell lines tested. The novel derivatives induced a G0/ G1 cell cycle arrest whilst the parental compounds induced a G2/ M block of the cell cycle, using flow cytometry. This suggested a different mechanism of action for the novel compounds. Dihydroartemisinin and the most active novel hybrid, EXP57EA, were investigated to understand their molecular mechanisms of action in oesophageal cancer.
- ItemOpen AccessNatural product derivative activates autophagy in cancer cells(2016) Andong Koung Edzidzi, Ursula-Claire; Hendricks, DenverArtemisinin, a natural product and its derivatives are potent antimalarial compounds, which have shown anticancer activity. In this study, we further characterized a novel artemisinin derivative namely EXP57EA which was previously designed and synthesized by the Chemistry Department at the University Of Cape Town. We determined the effect of EXP57EA on a panel of cancer cell lines, characterized the mode of cell death and also performed preliminary investigations of the signaling pathways that trigger the mode of cell death. Dihydroartemisinin (DHA), EXP57EA and cisplatin were screened on a selected panel of cancer cell lines: 3 esophageal cancer cell lines WHCO1, WHCO5, KYSE150; one breast cancer cell line MDA-MB-231 and one cervical cancer cell line SiHa. The 3-[4, 5-dimethylthiazol-2-yl]-2, 5- diphenyltetrazolium bromide (MTT) assay, and analysis with GraphPad prism software were used to calculate IC₅₀ values. EXP57EA displayed toxicity in the panel of cancer cell lines studied, and had lower IC₅₀ values (IC₅₀ values were ranging from 15.8 μM to 25.1 μM) than DHA and cisplatin. DHA was only active in two cells lines: WHCO1 (21.3 μM) and WHCO5 (77.3 μM), IC₅₀ values of cisplatin were ranging from 31.2 μM to 108.1 μM. EXP57EA was further investigated to understand the mode of cell death activated in the panel of cancer cell lines. The results showed that EXP57EA did not induce apoptosis in any of the cell lines studied, whereas DHA induced apoptosis, based on the PARP cleavage assay. In contrast, treatment with EXP57EA induced the appearance of vacuoles in treated cells compared to untreated cells, which was suggestive of autophagy. Autophagy was monitored by analyzing the expression level of two autophagy markers, Beclin1 and LC3-II by western blot. It was observed that EXP57EA treatment caused changes in the expression levels of both Beclin1 and LC3-II. We showed that EXP57EA induced elevated levels of autophagy, based on an increase in the flux of autophagy in the treated cells, since the lysosomal inhibitors ammonium chloride (NH₄Cl) and chloroquine substantially blocked LC3-II turnover in WHCO1 (confirmed previous result in our laboratory) and SiHa cancer cell lines. Furthermore, we also showed that treatment with EXP57EA resulted in increased expression of CHOP (by Real-Time PCR), and activated the PERK/eIF2α pathway, since treatment of WHCO1 cells with EXP57EA stimulated phosphorylation of eIF2α, suggesting that ER stress might be involved in mediating EXP57EA-induced cell death. Our results also suggested that EXP57EA activated the JNK pathway since treatment of WHCO1 and WHCO5 cells with EXP57EA stimulated phosphorylation of cjun and resulted in elevated levels of total c-jun. These results suggested the JNK pathway might also be involved in EXP57EA-induced cell death. However, the proposed involvement of the PERK/eIF2α pathway and the JNK pathway in EXP57EA-mediated autophagy is of a preliminary nature, and further work will have to be done to confirm the involvement of these pathways. This study showed that EXP57EA may have potential as an anticancer drug lead.
- ItemOpen AccessNovel chemotherapeutic agents for oesophageal cancer(2012) Khuzwayo, Londiwe Mandisa; Hendricks, DenverChemotherapeutic agents such as cisplatin, doxorubicin and 5-fluorouracil are currently used in the treatment of a variety of cancers, including oesophageal cancer. Although these agents have been part of our therapeutic repertoire for many years, there are several side effects that have been associated with their use in cancer therapy. Thus there is a need to develop novel chemotherapeutic agents with improved activity and less severe side effects. In this project, twelve compounds, consisting of 5 platinum dichloride complexes, four iminophosphine ligands and three gold (I) chloride complexes that were synthesised by a PhD student in our laboratory were tested for anticancer activity and the mode of action of these compounds was also explored.
- ItemOpen AccessProliferative and survival pathways in oesophageal cancer(2011) Esau, Luke Emmanuel; Hendricks, DenverOesophageal squamous cell carcinoma (OSCC) is the 8th most common cancer worldwide with high incidence in areas that include China, Iran and South Africa. The current treatment available for OSCC does not significantly enhance patient survival. A better understanding of proliferative and survival pathways activated in OSCC could allow identification of more specific therapeutic targets, potentially improving management of OSCC. Cell surface receptors are known to play important roles in relaying signals from the extracellular environment...The aim of this study was to determine the role of EGFR, IGF-1R and CXCR2 in proliferation and survival of OSCC cells.