Hampel, Sonja Margarethe; (2011) Design and Synthesis of G-Quadruplex Binding Small Molecules and their Evaluation as Anti-Cancer Agents. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

G-quadruplexes are specific nucleotide structures of stacked planes of guanines, and can form in G-rich telomeres and promoter regions of oncogenes. Their stabilisation by small molecules has a profound anti-cancer effect, as it leads to telomerase inhibition and telomere uncapping followed by DNA damage response and senescence, as well as chromosomal instability, and down-regulation of oncogene expression. This thesis reports on the design and synthesis of small molecule G-quadruplex ligands and their biophysical and biological evaluation. A series of highly water soluble naphthalene diimides (NDs) with N-methylpiperazine end groups was synthesised, which have high affinity and selectivity for human telomeric G-quadruplexes over duplex DNA, as shown in FRET melting temperature assays. The compounds were conceived by molecular modelling as derivatives of previously described potent NDs, indicating enhanced selectivity through groove interactions between protonated N-methylpiperazine groups and phosphates. Circular dichroism studies indicate that the ligands induce a parallel G-quadruplex topology in solution. In 96 h SRB assays, the compounds show high toxicity to a panel of cancer cell lines including pancreatic cancer cells, with IC50 values in the order of 0.1 μM and up to 100 fold selectivity over a normal fibroblast cell line. Treatment with the compounds at sub-cytotoxic concentrations over several weeks decreases growth of pancreatic cancer cells, resulting in senescence. The fluorescent lead compound was visualised inside the nuclei and cytoplasm of cancer cells by confocal microscopy. It causes DNA damage in cancer cells, which was indicated by detection of the phosphorylated histone H2AX. Cell cycle analysis was undertaken with propidium iodide in relation to further cellular responses to the compounds. The thesis examines the likely mode of action of these compounds in the light of this and further data, and suggests that they are promising candidates for the treatment of pancreatic cancers. The thesis also reports on the design and synthesis of other novel G-quadruplex ligands.

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