Targeting endometrial cancer stem cells through aldehyde dehydrogenase-mediated photodynamic therapy based on enhanced ring-fused chlorins

This study presents the development of an aldehyde dehydrogenase (ALDH)-mediated photodynamic therapy (PDT) approach targeting endometrial cancer stem cells (CSC). Novel 4,5,6,7-tetrahydropyrazolo[1,5-a]pyridine-fused chlorins (APX) were synthesised incorporating aldehyde groups, which are enzymatically oxidised by endogenous ALDH, highly expressed in CSC-enriched populations, into carboxylic acids, thereby enhancing intracellular retention and improving PDT efficacy. APX molecules were characterised and evaluated in vitro using endometrial cancer cells and CSC-enriched populations, and in vivo using a xenograft model. Assessments included reactive oxygen species (ROS) generation, cell viability, photoactivation-induced cell death, cell cycle progression, and ALDH expression in endometrial cancer cells. In CSC, the interaction between APX and endogenous ALDH, APX subcellular localisation, and CSC-associated marker expression were evaluated post-treatment. APX3 exhibited high absorbance potential at 652 nm, a moderate fluorescence quantum yield of 0.21, and promising photodynamic activity in endometrial cancer cells. Our findings showed that APX3 produced a robust photodynamic effect, inducing cell death via both necrosis and apoptosis, primarily through Type 1 photodynamic reactions mediated by superoxide anion production, whilst also affecting ALDH expression. In vivo, APX3-PDT effectively suppressed tumour growth, increasing caspase-3 expression and decreasing ALDH1 levels, demonstrating its therapeutic potential. In endometrial CSC, endogenous ALDH interacted with APX in vitro, converting aldehyde moieties into carboxylic acids. APX accumulated preferentially in ALDH-enriched zones of endometrial CSC, decreasing their viability and inducing cell death through necrosis. These findings underscore the potential of targeted PDT using APX ring-fused chlorins as an effective strategy for selectively eliminating ALDH+ endometrial CSC.