Date

Wednesday 26 Aug 2026

Time

11:00 - 11:30

Location

BW019

Supervisor

Sylvestre Bonnet

2nd reviewer

Hermen Overkleeft

Jury

Dennis Hetterscheid

Proteasome inhibitors (ProIs), such as carfilzomib (CFZ), represent the therapeutic base for the treatment of multiple myeloma (MM). However, treatment with CFZ has been associated with cardiovascular side effects thereby hampering its use in treating solid tumors. A potential way to effectuate the applicability of CFZ for the treatment of solid tumors is by incorporating it into photoactivated chemotherapy (PACT) pro-drug complexes. PACT allows site-restricted activation of ProIs upon light irradiation, thereby enabling a possible increase in administered dose and consequently intratumoral drug concentration without the induction of intolerable side effects. Derivatives of CFZ containing either 4-pyridyl-L-alanine, replacing the phenylalanine (1), the leucine furthest away from the epoxyketone warhead (2) and the homo-phenylalanine (3), or containing a 4-pyridylacetyl cap which replaces the morpholino N-cap (4), were conjugated to [Ru(bisaminopyridylterpyridine (baptpy)]2+, thereby generating the far-red light activatable PACT complexes [5](PF6)2 - [8](PF6)2 respectively. Within this thesis project, the biological activities of the free inhibitors 1 – 4 were evaluated in human melanoma A375 cells, specifically their proteasome subunit inhibitory potency using Activity-Based Protein Profiling (ABPP) in A375 cell lysate and effects on cell viability upon treatment of A375 cells using dimethylimidazole (MTT) assays. Findings reveal that 1 – 4 retain strong proteasome inhibitory potency and strongly reduce A375 cell viability. Additionally, intracellular proteasome subunit binding of [6](PF6)2 without and upon light irradiation and of its corresponding ProI (2) were assessed through a newly developed protocol for live-cell ABPP analysis. Results demonstrated a dose dependent decrease in residual proteasome activity proving that [6](PF6)2 and 2 are capable of efficiently crossing the plasma membrane of A375 cells after which both compounds successfully reach intracellular proteasomes and engage with the active sites of the proteasome subunits. Additionally, 655 nm light irradiation effectuates intracellular photosubstitution of [6](PF6)2, thereby liberating and activating its inhibitory molecule 2 inside the cell to generate an additional level of inhibition of proteasome subunit β5 which could not be achieved in the dark.