ESTRO 2024 - Abstract Book

S5324

Radiobiology - Tumour biology

ESTRO 2024

296

Proffered Paper

Feasibility of ferroptosis inducers as clinical radiosensitizers.

Lisa Kerkhove 1 , Febe Geirnaert 1 , Hugo Vandenplas 2 , Adrián Gutiérrez, 1 , Thierry Gevaert 1 , Inès Dufait 1 , Mark De Ridder 1

1 Vrije Universiteit Brussel, Radiotherapy, Jette, Belgium. 2 Vrije Universiteit Brussel, Medical Oncology, Jette, Belgium

Purpose/Objective:

Radioresistance, predominantly due to a hypoxic tumor environment, remains a hurdle in the field of radiotherapy (RT). Recently, it has been confirmed that RT triggers ferroptosis, a type of iron-dependent regulated cell death (RCD), alongside the established forms of RCD (apoptosis and necroptosis). However, the exact contribution of ferroptosis within RT-induced RCD remains uncertain. Previous findings demonstrated that sulfasalazine (SSZ) enhanced radiosensitivity in two hypoxic human colorectal cancer (CRC) cell lines, partially by induction of ferroptosis 1 . Consequently, the first aim of this study was to fully elucidate the role of ferroptosis in RT-induced RCD and assess its potential as a clinical target. Despite a decade of research into ferroptosis as an anti-cancer treatment, recent research has highlighted the involvement of ferroptosis in fibrosis development, a process underlying chronic toxicity after RT. Hence, a secondary objective was to investigate the role of ferroptosis in normal tissue toxicity after RT. This research is crucial to enhance the prospects of ferroptosis inducers (FINs) as clinical radiosensitizers.

Material/Methods:

This study included three human cell lines (DLD-1, HCT116 and HT29), one murine CRC cell line (CT26), and a human intestinal epithelial cell line (HIEC6) as normal tissue control. Radiomodulatory effects of SSZ (100 – 500 µM) were explored in vitro in 2D and 3D, via colony formation assays (CFA) and spheroid growth. Additional evidence was gathered by performing in vivo experiments in a DLD-1 xenograft model. The importance of the different types of cell death was evaluated using different cell death inhibitors; 10 µM Z-VAD-FMK (apoptosis), 10 µM necrostatin-1 (necroptosis) and 5 µM Ferrostatin-1 (ferroptosis). Afterwards, cells were irradiated under hypoxic (1x16Gy/ 1x20Gy) or normoxic conditions (1x6Gy/ 4x2Gy) followed by CFA. These results were confirmed in 3D spheroids. Reactive oxygen species (ROS) generation and ferroptosis induction after SSZ treatment were evaluated in HIEC6 cells and compared to CRC cells.

Results:

Prior research elucidated that SSZ enhanced the radiosensitivity of two human CRC cell lines (DLD-1 and HCT116) under hypoxic conditions (enhancement ratio of 1.9 and 1.6) in vitro and in vivo . The radiomodulatory features in DLD 1 can be attributed to triggering ferroptosis, while no ferroptosis induction was present in HCT116 1 .

This study seeks to assess the importance of ferroptosis in RT-induced RCD in DLD-1 and HCT116. Due to the expected variability in RT response, two additional CRC cell lines were included.