AbstractThe hallmarks of cancer have paved the way for the development of target approached agents which are more selective and active towards cancer cells. Irrespective of this
paradigm shift, there are still many hindrances to effective drug treatment which cause drug resistance such as hypoxia and mild acidity which are found in the tumour microenvironment (TME). One way to circumvent this issue is by the use of hypoxia activated prodrugs (HAPs), but with over 40 years of research there is still no HAP that is approved for use in humans. A new strategy to overcome this barrier is to apply a phenotypic screening approach to test compounds under those conditions which are favourable to the TME such as the combination of hypoxia and mild acidity.
The aim of this study is to evaluate a series of compounds using a phenotypic evaluation strategy based upon testing compounds under (i) normoxia / pHe 7.4, (ii) normoxia / pHe 6.5, (iii) hypoxia / pHe 7.4 and (iv) hypoxia / pHe 6.5 and (v) compare activity in cancer cells to non-cancer cells. The objective of this study is to identify compounds that preferentially and selectively kill cancer cells that reside within the TME that would not have been identified using standard cell culture conditions.
In this study, a series of novel compounds (i) ruthenium, iridium and rhodium based napthyridines, thiourea derivatives, benzylthiourea ligands and aroylthiourea compounds, (ii) Bromo-hexitol prodrugs, (iii) NHC-Ag based complexes, (iv) Minor groove binders and (v) Tyrosine kinase inhibitor (dasatinib) with the attachment of 2-nitroimidazole have been provided by collaborators which will be screened under TME conditions to determine the potency (IC50), under hypoxic conditions (0.1% O2) and in combination of mild acidity (pHe 6.5). As well as potency the selectivity index (SI) which is defined as the ratio of IC50 values in non-cancer cells divided by the IC50 of cancer cells will also be determined. Those compounds that pass these tests are then to be subjected for further mechanistic studies.
The initial studies focused on screening three classic chemotherapeutic drugs, cisplatin, 5- FU, etoposide and the classic HAP tirapazamine under the TME conditions. All three classic chemotherapeutic drugs displayed high activity under normoxic conditions but showed resistance under TME conditions (i.e. hypoxia, mild acidity and the combination of both) with moderate to poor selectivity towards cancer cells. As expected, tirapazamine displayed HAP like abilities. These results were set as the ‘gold standard’ throughout this PhD study and were used to compare with the new novel compounds that were screened. A total of 72 compounds were tested using this screen where 40 (55%) compounds were selective towards the cancer cell lines, 8 (11%) compounds were further activated under normoxic extracellular pH (pHe) 6.5 conditions, 24 (33%) compounds exhibited HAP like properties and finally 3 (4.1%) compounds displayed enhanced toxicity with the combination of hypoxia and pHe 6.5 conditions. The compounds screened in some instances did display superior selectivity and preferential activity towards cells under TME conditions in comparison to the classic chemotherapeutic drugs which is promising.
In conclusion, the phenotypic based screen employed throughout this study demonstrates potential in identifying compounds that show preferential activity and resistance to cells under TME conditions. Both results are valuable in terms of the decision making process for the selection of lead compounds and further studies are required to develop this concept further.
|Date of Award||2023|
|Supervisor||Roger Phillips (Co-Supervisor) & Simon Allison (Co-Supervisor)|