EACR25-2345

Targeting endometrial cancer stem cells with an ALDH-mediated photodynamic therapy

B. Serambeque1, B. Costa2, S. Pinto Lopes3, R. Alves4, R. Teixo1, M. Pineiro5, M. Botelho6, T. MVD Pinho e Melo5, M. Carvalho7, M. Laranjo6
1Univ Coimbra, Center for Innovative Biomedicine and Biotechnology (CIBB); Univ Coimbra, Coimbra Institute for Clinical and Biomedical Research (iCBR) Area of Environment Genetics and Oncobiology (CIMAGO), Institute of Biophysics, Faculty of Medicine, Coimbra, Portugal
2Univ Coimbra, Center for Innovative Biomedicine and Biotechnology (CIBB); Univ Coimbra, Coimbra Institute for Clinical and Biomedical Research (iCBR) Area of Environment Genetics and Oncobiology (CIMAGO), Institute of Biophysics, Faculty of Medicine; Univ Coimbra, Coimbra Chemistry Centre - Institute of Molecular Sciences and Department of Chemistry, Coimbra, Portugal
3Univ Coimbra, Coimbra Institute for Clinical and Biomedical Research (iCBR) Area of Environment Genetics and Oncobiology (CIMAGO), Institute of Biophysics, Faculty of Medicine, Coimbra, Portugal
4Univ Coimbra, Center for Innovative Biomedicine and Biotechnology (CIBB); Univ Coimbra, Coimbra Institute for Clinical and Biomedical Research (iCBR) Area of Environment, Genetics and Oncobiology (CIMAGO), Laboratory of Oncobiology and Hematology (LOH) and University Clinics of Hematology and Oncology, Faculty of Medicine; Clinical Academic Centre of Coimbra (CACC), Coimbra, Portugal
5Univ Coimbra, Coimbra Chemistry Centre - Institute of Molecular Sciences and Department of Chemistry, Coimbra, Portugal
6Univ Coimbra, Center for Innovative Biomedicine and Biotechnology (CIBB), Coimbra Institute for Clinical and Biomedical Research (iCBR) Area of Environment Genetics and Oncobiology (CIMAGO), Institute of Biophysics, Faculty of Medicine; Clinical Academic Centre of Coimbra (CACC), Coimbra, Portugal
7Univ Coimbra, Coimbra Institute for Clinical and Biomedical Research (iCBR) area of Environment Genetics and Oncobiology (CIMAGO), Institute of Biophysics and Universitary Clinic of Gynecology, Faculty of Medicine; Gynecology Service, Department of Gynecology, Obstetrics, Reproduction and Neonatology, Unidade Local de Saúde de Coimbra, Coimbra, Portugal
Introduction:

Endometrial cancer (EC) remains a clinical challenge, where cancer stem cells (CSC) have been associated with chemoresistance and recurrence. In particular, a relevant expression of aldehyde dehydrogenase (ALDH) on EC has been involved in disease progression and is associated with stemness pathways, proposing its double potential as a biomarker and a molecular target. To investigate more effective and minimally invasive treatments for EC, our research focuses on a novel approach using CSC-targeted photodynamic therapy (PDT) based on modified 4,5,6,7-tetrahydropyrazolo[1,5-a]pyridine-fused chlorins (PX).

Material and method:

A refined sphere-forming protocol was performed to obtain endometrial CSC from ECC-1, RL95-2 and HEC-1-A cell lines. To evaluate the effectiveness of a CSC-targeted PDT strategy in EC, the colocalisation of a novel class of tetrahydropyrazolo[1,5-a]pyridine-fused chlorins functionalised with aldehyde moieties (A-PX) with ALDH was verified using confocal microscopy. Then, endometrial CSC were incubated with a range of A-PX concentrations (2.5 – 10 µM) for 2 hours, followed by a light irradiation (7.5mW/cm2, 10J). To assess the expression of CSC markers, the effect of CSC-targeted PDT on CD44 and CD24 was examined through flow cytometry. Viability and cell death were determined through luminescence and confocal microscopy.

Result and discussion:

After evidence that ALDH interacts with A-PX in vitro and converts aldehydes into carboxylic acids and A-PX can internalise and accumulate in endometrial CSC, preliminary confocal studies pointed out that A-PX localise in CSC ALDH-enriched zones, facilitating the expected action mediated by ALDH. Moreover, initial findings revealed a high percentage of CD44+/CD24- cells, confirming CSC-like characteristics. CSC-targeted PDT seems not to modulate the stemness features of the ECC-1 CSC-enriched populations. Nevertheless, A-PX-based PDT notably contributed to eliminating endometrial CSC, resulting in a significant reduction in viability and the induction of cell death.

Conclusion:

In conclusion, these results emphasise the potential of the A-PX-mediated strategy to target ALDH in vitro and within CSC, as well as the effectiveness of A-PX-based PDT for eradicating EC cells. This research represents the first steps towards developing a conservative and minimally invasive therapeutic option for challenging cases of EC. FCT supports CIBB (10.54499/UIDB/04539/2020; 10.54499/UIDP/04539/2020; 10.54499/LA/P/0058/2020); CQC (10.54499/UIDB/00313/2020; 10.54499/UIDP/00313/2020; TEMA (10.54499/UIDB/00481/2020), UIDP/00481/2020 (10.54499/UIDP/00481/2020); Projects CarboNCT (10.54499/2022.03596.PTDC) and Chem4LungCare (10.54499/PTDC/QUI-QOR/0103/2021); PhD Scholarship from FCT and European Social Fund to BS (10.54499/2020.07672.BD), BC (10.54499/2022.12013.BD) and RT (SFRH/BD/116794/2016).