Oesophageal squamous cell carcinoma ranks amongst the ten most frequently occurring cancers in the world with some of the highest incidences being reported in the Eastern Cape region of South Africa (Transkei). The etiology of this disease remains obscure. 2-Methoxyestradiol (2 ME), an endogenous estrogen metabolite, may be a defense mechanism against tumors. The aim of this study was to investigate whether 2 ME affects proliferation and/or induces apoptosis in oesophageal cancer cell lines and if so, by what mechanism. 2 ME decreased cell numbers in two oesophageal cell lines investigated. Cells treated with 2 ME showed morphological hallmarks of apoptosis, G2/M arrest and spindle disruption. Increased expression of death receptor 5 protein suggested that the extrinsic pathway was activated to induce apoptosis in oesophageal cancer cells. 2 ME has antitumor effects on oesophageal cancer cells by inducing apoptosis. It can be suggested that 2 ME can be considered to be a potential chemotherapeutic for the treatment of oesophageal cancer. / Dissertation (MSc (Physiology))--University of Pretoria, 2006. / Physiology / unrestricted

This thesis mainly deals with some biochemical aspects regarding the efficacy of novel platinum anticancer compounds alone and in combination with mangiferin, as part of a broader study in which both chemistry and biochemistry are involved. Various novel diamine and N-S donor chelate compounds of platinum II and IV have been developed in which factors such as stereochemistry, ligand exchange rate and biocompatibility were considered as additional parameters. In the first order testing, each of these compounds was tested with reference to their “killing” potential by comparing their rate of killing, over a period of 48 hours with those of cisplatin and oxaliplatin. Numerous novel compounds were tested in this way, using the MTT cell viability assay and the three cancer cell lines MCF7, HT29 and HeLa. Although only a few could be regarded as equal to or even better than cisplatin, CPA7 and oxaliplatin, the testing of these compounds on cancer cells provided useful knowledge for the further development of novel compounds. Three of the better compounds, namely Yol 25, Yol 29.1 and Mar 4.1.4 were selected for further studies, together with oxaliplatin and CPA7 as positive controls, to obtain more detailed knowledge of their anticancer action, both alone and when applied in combination with mangiferin. In addition to the above, resistant cells were produced for each of the three different cell lines tested and all the selected compounds, both in the presence and absence of mangiferin. The effects of these treatments on the activation of NFĸB when applied to normal and resistant cell lines were also investigated. All the compounds induced apoptosis in the cell lines tested as well as alter the DNA cycle at one or more phase. Additionally, combination of these compounds with mangiferin enhanced the above-mentioned effects. Mangiferin decreases the IC50 values of the platinum drugs by up to 3.4 times and, although mangiferin alone did not induce cell cycle arrest, the presence of mangiferin in combination with oxaliplatin and Yol 25 shows an earlier and greatly enhanced delay in the S-phase, while cells treated with CPA7, Yol 29.1 and Mar 4.1.4 in combination with mangiferin showed a later, but greatly enhanced delay in the S-phase. It was also found that mangiferin acts as an NFĸB inhibitor when applied in combination with these drugs, which, in turn, reduces the occurrence of resistance in the cell lines. Resistance to oxaliplatin was counteracted by the combination with mangiferin in HeLa and HT29, but not in MCF7 cells, while resistance to CPA7 was only counteracted in the MCF7 cell line. Yol 25 and Mar 4.1.4 did not seem to induce resistance in HeLa and MCF7 cells, but did in HT29 cells, whereas Yol 29.1 caused resistance in HeLa and HT29 cells, but not in MCF7 cells. Finally, an effort was made to evaluate the different compounds by comparing them with respect to their properties relating to anticancer action with and without the addition of mangiferin.

This dissertation mainly deals with some biochemical aspects regarding the efficacy of novel platinum anticancer compounds, as part of a broader study in which both chemistry and biochemistry are involved. Various novel diamine and N-S donor chelate compounds of platinum II and IV have been developed in which factors such as stereochemistry, ligand exchange rate and biocompatibility were considered as additional parameters. In the first order testing, each of these compounds was tested with reference to their “killing” potential by comparing their rate of killing, over a period of 48 hours with those of cisplatin and oxaliplatin. Some 80 compounds were tested in this way. Although only a few could be regarded as equal to or even better than cisplatin and oxaliplatin, the testing of these compounds on cancer cells provided useful knowledge for the further development of novel compounds. Four of the better compounds, namely Y9, Y14, Y16 and Lt16.2 were selected for further studies to obtain more detailed knowledge of their anticancer action, including some flow cytometric studies. In addition to the above, cisplatin resistant cells were produced for each of the three different cell lines tested, namely, HeLa, HT29 and MCF7 cancer cell lines, by intermittent and incremental exposure to cisplatin (all the cell lines tested became resistant to cisplatin). Each of the selected compounds were exposed to the cells in the same manner, in order to attempt the induction of resistance against these compounds in the three cell lines tested (i.e. whether these cells will become resistant to the various compounds). Each of these selected platinum containing compounds were subsequently tested against the “cisplatin resistant” cell lines in order to determine their efficacy against such cells. One such compound could be singled out, since cervical cancer cells (HeLa cells) do not become resistant to it. This behaviour is similar to that of oxaliplatin against cervical cancer and colon cancer (HT29) cells (oxaliplatin is the number one treatment for colon cancer at present). This compound also proved to be more active against cisplatin resistant cell lines. It was found that all the compounds induced apoptosis in the cell lines tested as well as inhibit the DNA cycle at one or more phase. Finally, an effort was made to evaluate the different compounds by comparing them with respect to their properties relating to anticancer action.

No description available.

No description available.

Amifostine (WR-2721, Ethyol®), a phosphorylated thiol, demonstrates the unique ability to protect normal but not tumour tissue from cytotoxic damage induced by radiation therapy and chemotherapy. This study tested the effect of amifostine's active metabolite, the free thiol, WR-1065, on the cytotoxicity of standard anticancer drugs against human A2780 ovarian and MCF7 breast cancer cell lines in vitro, using the well-characterised sulphorhodamine B assay. 50% inhibitory concentration (IC50) values were determined for each of 16 different anticancer drugs in the presence and absence of the highest nontoxic dose of WR-1065 from concentration-response curves constructed in triplicate and based on 18 replicate cell culture plates for each tested drug concentration. Pretreatment with WR-1065 had no statistically significant effect on the IC50 value of any of the 16 drugs tested against either the A2780 or MCF7 human tumour cells. These data expand upon previous reports showing that amifostine does not protect tumours from the cytotoxic effects of anticancer agents. The ability of amifostine to protect against dose-limiting toxicity to a variety of normal tissues without protection of tumour should enhance the efficacy ratio of a wide range of standard anticancer drugs.

Cytotoxic natural products represent a class of cancer drug candidates that have remained largely untapped as payloads in the antibody-drug conjugate (ADC) therapeutic modality. The epothilones, a class of exquisitely cytotoxic natural products and their synthetic analogs, are a prime example, and our work has focused on the development of epothilones as ADC payloads. Strategies toward this goal have included total synthetic efforts toward four structurally distinct epothilone analogs equipped with linker functionality and structural modifications designed to improve metabolic and chemical stability. In addition, we have pursued the synthesis of octreotide- epothilone conjugates, structures designed to target epothilones into cells that overexpress somatostatin receptors 2 and 5. Biological evaluation via in vitro cellular assays revealed one of our epothilone analogs as a promising epothilone-inspired ADC payload. Synthetic efforts toward these goals will be discussed.

One of the reasons for poor response of tumors is their three dimensional structure (micro-environment), which forms a penetration barrier to anti-tumor agents. A drug has to cross the interstitial space of tumors and many layers of cells to reach the interior of a tumor. The main objective of this study was to develop two new in vitro models to evaluate the in vivo availability of antitumor compounds to solid tumors. Two in vitro models, basement membrane p:1atrix of Engelbreth-Holm Swarm (EHS) mouse sarcoma (Matrigel) and human colorectal adenocarcinoma (Caco-2) multilayers were developed. The permeability characteristics of SR271425 (N-[1-{ [2-(diethylamino) ethyl] amino} -7 -methoxy-9-oxo-9H -thioxanthen-4-yl] methylformamide) a novel antitumor compound and three of its thioxanthone analogs (probable metabolites) were studied to evaluate the in vitro models.

No description available.

Podophyllotoxin, a naturally occurring lignan isolated from several species of Podophyllum, is used as a precursor to the clinical chemotherapeutic agents teniposide and etoposide. The biosynthesis of podophyllotoxin is not fully understood, but its optical activity, like that of most lignans, is suggestive of enzyme-mediated processes. It has been proposed that the formation of podophyllotoxin begins with stereo-controlled coupling of a hydroxy cinnamyl alcohol derivative and a substituted hydroxy Cinnamic acid, although no "coupling" enzyme has been isolated to date. Further biosynthetic modifications of the coupled compound could lead to matairesinol and/or yatein, which have been proposed as potential biological precursors of podophyllotoxin. Although no firm evidence has been obtained to date, conversion of matairesinol to yatein has been postulated. This conversion would, however, involve biosynthetic steps which, though common for hydroxycinnamates, are unprecedented at the dimeric level. Conversion of yatein to podophyllotoxin has been demonstrated, with the conversion involving a stereo-controlled cyclization and subsequent stereospecific hydroxylation. In order to investigate the biosynthesis of podophyllotoxin, leading from the postulated precursors matairesinol and yatein, a series of stereospecific deuterium-labelled matairesinol and yatein derivatives was proposed and the synthetic methodology for each compound developed. The methodology used to obtain deuterium-labelled compounds can be extended to generating tritium-labelled compounds as well. With sufficient quantities of a number of the deuterium-labelled compounds, feeding studies can now be carried out in Podophyllum plants. Isolation and analysis of podophyllotoxin, from plants fed with labelled yatein, will allow determination of the stereochemical nature of yatein cyclization. Isolation and analysis of yatein, from plants fed with labelled matairesinol, will indicate whether matairesinol is indeed a precursor to yatein (and, hence, podophyllotoxin). The information obtained from the synthesis and incorporation of such labelled compounds should then provide a clearer understanding of some interesting but, as yet, unestablished biotransformations. / Master of Science