par Vanhalle, Lucas 
Président du jury Debaste, Frédéric
Promoteur Valkenier, Hennie
Publication Non publié, 2025-06-09
Président du jury Debaste, Frédéric
Promoteur Valkenier, Hennie
Publication Non publié, 2025-06-09
Mémoire
| Résumé : | Cancer remains a leading cause of mortality in Western Europe, with approximately 20% of cases proving refractory to conventional therapeutic interventions. Brain, oesophageal and lung cancers represent particularly challenging malignancies requiring novel treatment approaches with fundamentally distinct mechanisms of action. Copper homeostasis dysregulation has emerged as a promising therapeutic strategy, as copper plays essential roles in cancer cell proliferation, metastasis and chemoresistance. Copper dyshomeostasis can notably be induced through metalloallostery and the recently discovered cuproptosis pathway, mediated by ionophores. This master thesis investigates the copper ion transmembrane transport capacities of commercially available copper ionophores alongside Cuphoralix, a novel synthetic copper(I) ionophore developed through collaboration between the EMNS and LCO laboratories. This comparative analysis primarily seeks to gain a better understanding of the activity and selectivity for Cu+ and Cu2+ transport in liposomes by these compounds whilst assessing their differences relative to Cuphoralix. The compounds examined in this study comprises disulfiram, elesclomol, pyrithione, neocuproine, Cu-ATSM and copper ionophores I/IV. Copper ion transmembrane transport was assessed using liposomes containing encapsulated fluorescent probes that bind with transported copper ions. Calcein probe was employed for Cu2+, whilst BCS probe was used for Cu+. The transmembrane transport activity of ionophores was evaluated by monitoring fluorescence quenching upon copper complexation with the fluorescent probes. Rapid fluorescence decrease indicates effective transport across the liposome membrane. Transport studies at 1:1000 ionophore-to-lipids ratios revealed distinct mechanistic profiles. Elesclomol and Cu-ATSM showed no transmembrane transport capacity for either ion due to reduction requirements and unsuitable coordination geometry for Cu+ complexation. Neocuproine exhibited moderate Cu2+ transport but high Cu+ transport efficiency. Pyrithione and clioquinol showed moderate Cu+ transport but efficient Cu2+ transport. Disulfiram demonstrated high transport capacity for both ions, though apparent copper(I) transport likely involves redox reaction rather than direct Cu+ transmembrane transport. Cuphoralix exhibited selective Cu+ transport with no Cu2+ activity. EC50 determinations revealed clioquinol as the most potent Cu2+ transporter, followed by disulfiram and pyrithione. Complementary studies using genetically modified yeast cells lacking endogenous copper transport mechanism were conducted to evaluate copper transport restoration capacity of studied compounds. However, significant contaminations and cytotoxicity of several ionophores limited definitive conclusions regarding their transport efficacy in living systems. This comparative analysis supports Cuphoralix’s unique selectivity for copper(I) ions and its probable exclusive membrane distribution mechanisms, distinguishing it from conventional ionophores that require deprotonation or reduction steps. These findings corroborate Cuphoralix’s potential as a novel therapeutic agent targeting copper homeostasis in refractory cancers, warranting further investigations through cellular cytotoxicity studies and mechanistic characterisation. |
