School of Chemistry - Theses
Permanent URI for this collection
Search Results
1 - 1 of 1
-
ItemSynthesis and biological evaluation of episilvestrol analogues( 2014)This thesis details the study of the mode-of-action of episilvestrol (2) and related compounds. Improvements were made to the total synthesis of episilvestrol (2) that allowed for sufficient production of this low-abundance natural product for additional manipulation and testing. Various analogues of episilvestrol (2) were synthesized via total synthesis, building on established methods. Biological testing was performed to identify the protein target(s) of silvestrol (1) and episilvestrol (2) for translation inhibition and elucidate the mode-of-action of these anticancer metabolites. Additionally, the total synthesis of the recently isolated natural product 2’’’,5’’’- diepisilvestrol (263) was achieved. Attempts to elaborate the C4’ position of episilvestrol (2) were not successful. Modification of the C2 methyl ester was pursued after it was found that silvestric acid 238 and episilvestric acid 241 retained comparable in vitro translation inhibition activity to the parent natural products. Biotinylated episilvestrol 250 was synthesized from a Huisgen 1,3-dipolar cycloaddition between propargyl amide 244 and biotin azide 249 and found to have satisfactory activity for further testing. Two fluorescent episilvestrol derivatives, 251 and 252, were made in a similar manner and demonstrated analogous activity to 250. Further analogues of episilvestrol (2) were synthesized to study the structure-activity relationship of this compound. It was found that, while 1,2,3,3a,8b-epi episilvestrol 254 had reduced activity as compared to episilvestrol (2), 1’’’, 5’’’-diepisilvestrol 258, 2’’’,5’’’-diepisilvestrol (263), and 1’’’,2’’’,5’’’- triepisilvestrol (268) were all essentially inactive. This demonstrated that the C1’’’ and C2’’’ stereochemistry has a greater effect on translation inhibition. C2 analogues of methyl rocaglate (4) were achieved using the same methodology as the C2 episilvestrol analogues. Propargyl rocaglamide 269 was only ~4 times less active than episilvestrol (2) while biotinylated rocaglamide 270 was almost as active to the parent compound. Simplified analogues of episilvestrol (2) ̶ glycoside 274 and 275, dioxane 281, and the flavone derivative 285 ̶ were found to have far inferior activity. The biotinylated analogues 250, 261, and 270 were used in streptavidin pulldown assays to determine the protein target(s) of these compounds. A direct and exclusive interaction of silvestrol (1) and biotinylated episilvestrol 250 with eIF4AI/II, was identified by Western blot, silver stain, and GC-MS analysis. Crystallography studies to find the binding site of 1 and 2 with eIF4AI were attempted. Investigations into how silvestrol (1) and episilvestrol (2) cause cell death have shown that reduction in the level of anti-apoptosis protein Mc1-1 was not the primary cause of death in a variety of cells. Cell death was also not always controlled by the Bax/Bak pathway. Furthermore, neither proliferation nor differentiation of cells correlated with silvestrol (1) induced cell death. These results suggest that while silvestrol (1) and episilvestrol (2) appear to have only one cellular protein target, eIF4AI/II, they cause cell death via multiple pathways.