AbstractPoly(ADP-ribose) polymerase (PARP) is a superfamily of nuclear enzymes that can modify histones and other nuclear proteins for the survival of injured, proliferating cells by catalysing the transfer of ADP-ribose units. PARP1 is a DNA-dependent PARP known for its ability to facilitate DNA repair in cancer; PARP14 is a macroPARP and is known to promote the Warburg effect in cancer. Both PARP1 and PARP14 are overexpressed in malignancies, and PARP1 is upregulated when exposed to chemotherapeutics and radiation. Current PARP1 inhibitors are non-selective towards PARP1 and PARP2, and there are no clinically approved PARP14 inhibitors. Although current PARP1 inhibitors provide a positive outcome to clinical treatments, some adverse effects are linked to the co-inhibition of both PARP1 and PARP2. The structural similarity probed the need to understand further the structure of all members in the PARP superfamily. Computational modelling was performed to identify key and unique residues of the superfamily, categorised based on its structural similarity and the subclass.
Generation 1 compounds, synthesised as part of previous work, was evaluated in vitro with commercially available chemiluminescence PARP assay kits, revealing 4-(((1-(4-methoxyphenethyl)piperidin-4-yl)amino)methyl)phenol (compound 2) as a PARP1 lead and N-(2-methoxybenzyl)-1-(4-methoxyphenethyl)piperidin-4-amine (compound 7) as a PARP14 lead. These were derived to form a library of compounds with up to 60 potential products. 24 compounds were synthesised as part of generation 2 and were evaluated with PARP assay kits.
In vitro cell cultures was performed as the final part of this project. HeLa cells, representing cervical cancer and cancer with an expression of PARP1 and PARP14, and PC-3 cells, representing prostate cancer and cancer with an expression of PARP1 were used. Dose-response of the novel compounds and cisplatin was performed. In addition, a time-course experiment and combined dose-response experiment was also performed. The result suggests that both compound 2 and 7 significantly increased the cisplatin-induced lethality.
Future work will include western blotting as part of all microplate assays, radiation exposure to quantify radiosensitization potential, and three-dimensional cell culture to study drug penetration. These high-throughput techniques would allow the rapid development of novel PARP inhibitors.
|Date of Award||2022|
|Supervisor||Stephan Levonis (Supervisor), Stephanie Schweiker (Supervisor) & Catherine McDermott (Supervisor)|