The design and synthesis of probe molecules to validate the inhibition of epigenetic mechanisms for phenotypic responses

Previously held under moratorium in Chemistry department (GSK) from 9/10/14 until 24 August 2021.The confidentiality statement on each page of this thesis DOES NOT applyThe work described in this thesis concerns the design and synthesis of probe molecules for epigenetic proteins. In many cases, it is unclear what the effect of selective inhibition of epigenetic proteins would achieve in biological systems. This lack of understanding is in part due to the novelty of the area and suitable tool molecules would significantly enable further investigation. Probe molecules have been designed for two epigenetic targets using fragment based drug discovery techniques to grow the templates using structure based design. For the Jumonji D2 family of enzymes, a pyridopyrimidinone core with poor cellular penetration was grown at two positions to identify hydrogen bonding interactions between the ligand and the protein to increase the potency of the series. This resulted in selective compounds with approximately micromolar cellular activity. Two chemically distinct templates were investigated to discover probes for the PCAF bromodomain. Phthalizinone compounds were found to be selective for PCAF, although they had poor aqueous solubility. Improving the solubility of the molecules either abolished potency at PCAF or brought in unwanted off-target activity and work on this series was halted. Using a pyridazinone core, probe compounds for the PCAF bromodomain were found starting from an unselective molecule. Through identification of a putative salt bridge interaction, highly selective and potent compounds were accessed and shown to be capable of displacing PCAF from chromatin. The compounds identified were found to engage their respective targets in cellular systems and are suitable for phenotypic investigation of the inhibition of these epigenetic mechanisms.The work described in this thesis concerns the design and synthesis of probe molecules for epigenetic proteins. In many cases, it is unclear what the effect of selective inhibition of epigenetic proteins would achieve in biological systems. This lack of understanding is in part due to the novelty of the area and suitable tool molecules would significantly enable further investigation. Probe molecules have been designed for two epigenetic targets using fragment based drug discovery techniques to grow the templates using structure based design. For the Jumonji D2 family of enzymes, a pyridopyrimidinone core with poor cellular penetration was grown at two positions to identify hydrogen bonding interactions between the ligand and the protein to increase the potency of the series. This resulted in selective compounds with approximately micromolar cellular activity. Two chemically distinct templates were investigated to discover probes for the PCAF bromodomain. Phthalizinone compounds were found to be selective for PCAF, although they had poor aqueous solubility. Improving the solubility of the molecules either abolished potency at PCAF or brought in unwanted off-target activity and work on this series was halted. Using a pyridazinone core, probe compounds for the PCAF bromodomain were found starting from an unselective molecule. Through identification of a putative salt bridge interaction, highly selective and potent compounds were accessed and shown to be capable of displacing PCAF from chromatin. The compounds identified were found to engage their respective targets in cellular systems and are suitable for phenotypic investigation of the inhibition of these epigenetic mechanisms