Biological Activity of Steroid Analogues:Synthesis and Receptor/Enzyme Interactions
Degree GrantorUniversity of Canterbury
Degree NameDoctor of Philosophy
This thesis investigates the biological activity of selected non-steroidal analogues of sex steroid hormones by examining two different effects of analogues on endogenous sex hormone activity. Non-steroidal analogues of sex hormones were synthesised to study their biological interactions with a sex steroid receptor and a sex steroid metabolising enzyme. Chapter One introduces the steroid hormones and their physiology, which leads to a review of the mechanisms by which steroids exert their effects. Their implication in disease is discussed, with particular emphasis on the sex steroids. As the biological activity of steroids is related to their chemical structure, the important features of steroid structure are identified, including the cyclopentanoperhydrophenanthrene nucleus, arrangement of ring substituents and ring junction conformation. The concept of non-steroidal analogues of steroids is introduced, and the harmful or beneficial effects analogues have on endogenous steroid activity are considered. Alteration of steroid activity and its consequences are focussed on two main areas; the potential adverse effects of environmental chemicals which mimic sex steroid activity, and the use of non-steroidal analogues in medicinal chemistry for treating sex steroid related disease. Chapter Two describes an investigation into the 17β-estradiol mimicking activity of non-steroidal analogues. Exogenous chemicals that mimic estradiol are of concern as they may alter endogenous estradiol activity and disrupt endocrine systems. Firstly, an introduction to the field of research concerned with environmental chemicals that mimic steroid hormones is given. The interaction of xenoestrogens with the estrogen receptor is described, as are the methods available for assessing the estrogen mimicking activity of xenoestrogens. The concern for insecticides mimicking estrogen activity is described by reviewing reported activities of insecticides, which leads into a discussion of work carried out as part of this thesis. Metabolites of the pyrethroid insecticides permethrin and cypermethrin, 2.14, 2.15, and 2.16 were synthesised while others were commercially obtained. The interaction of pyrethroid insecticide metabolites with the human estrogen receptor expressed in recombinant yeast (Saccharomyces cerevisiae) was studied, following the establishment and validation of the assay. Metabolites 2.11, 2.12, and 2.14 were found to weakly stimulate estrogen receptor-mediated estradiol responsive gene expression in the yeast assay (105 less active than 17β-estradiol). Since the activity of the metabolites using the yeast assay was greater than for the parent compounds, metabolic pathways need to be considered when assessing the impact of exposure to environmental estrogens. The low estrogenic activity suggests these compounds are not individually contributing significantly to the xenoestrogenic impact on humans, but will add to total xenoestrogen exposure. Chapter Three describes the inhibition of a sex steroid metabolising enzyme, steroid 5a-reductase, by novel non-steroidal compounds. Inhibitors of this enzyme are potentially useful therapeutic agents for regulating the activity of an androgen in prostate disorders. A review of the literature on non-steroidal inhibition of 5a-reductase identified three key structural features known to enhance inhibitor potency; ring substitution, position and nature of ring unsaturation and angular methyl group presence. These features were taken into account in the design of inhibitors synthesised in this thesis (3.55-3.57, 3.59, 3.61, 3.62, 3.110 and 3.111). Inhibitors consisting of non-steroidal 5- or 1-aryl pyridone scaffolds were synthesised to investigate SAR for 4'-substituents. The 5-aryl 1-methyl-2-pyridone/piperidone scaffold of compounds 3.55-3.57 and 3.59 was constructed by Suzuki cross coupling methodology, while the 1-aryl 2-methyl 2,3-dihydro-4-pyridone scaffold of 3.61 and 3.62 was constructed by aza Diels-Alder methodology. Long carbon chain olefin containing tethers 3.107 and 3.108 were synthesised for conjugation to inhibitor 3.57 by cross metathesis to give conjugates 3.110 and 3.111. Compounds 3.55-3.57, 3.59, 3.61, 3.62, 3.110 and 3.111 inhibited the type 1 5a-reductase isozyme expressed by HEK-I cells, with activities comparable to those of related literature compounds. The 1-aryl 2,3-dihydro-4-pyridone 3.62 inhibited both the type 1 and 2 isozymes (expressed by HEK-II cells) of 5a-reductase. The presence of bulky hydrophobic groups (benzoyl, long chain tethers) at the 4' position enhanced the potency of type 1 inhibition by 5-aryl pyridone type compounds in comparison to N,N-diisopropyl- and N-allylacetamide groups. This information provides further understanding of SAR within and across different classes of non-steroidal inhibitors of steroid 5a-reductase towards improved drug design.