ESTRO 2024 - Abstract Book

S5193

Radiobiology - Microenvironment

ESTRO 2024

179

Digital Poster

Differential lipid peroxidation on tumour vs normal tissues as a key determinant of FLASH effect

Nuria Vilaplana Lopera, Jiyoung Kim, Iain Tullis, Ellie Walker, Ammar Abu-Halawa, Jia-Ling Ruan, Salome Paillas, Kristoffer Petersson, Ejung Moon

University of Oxford, Oncology, Oxford, United Kingdom

Purpose/Objective:

FLASH radiation (RT) uses ultra-high dose rates to enhance the therapeutic window by protecting normal tissue damage without diminishment of tumour cell killing. Compared to conventional RT, the FLASH effect has been attributed in part to the interplay between oxygen depletion and reactive oxygen species (ROS) production. Recently, it was proposed that production of lipid peroxidation may be a key to mediate the FLASH effect. However, detailed biological mechanisms of FLASH-induced lipid peroxidation still need to be investigated. In our study, using both in vitro and in vivo settings, we evaluated lipid peroxidation in normal and tumour cells/tissues and proposed a possible mechanism contributing to the FLASH effect.

Material/Methods:

Radiation (RT) was given at FLASH core established in the Department of Oncology at University of Oxford. FLASH RT was delivered at an average dose rate of 2-3 kGy/s with 3.5 µs electron pulses (a repetition rate of 300 Hz). Conventional RT was treated with 3.5 µs electron pulses (a repetition rate of 25 Hz) at an average dose rate of 0.1 Gy/s. To determine FLASH-induced lipid peroxidation, we treated MDA-MB-231 (a human breast cancer cell line) and A549 (a human lung cancer cell line) with FLASH and conventional radiation. MCF10A (a human normal breast epithelial cell line) was used to determine normal cell responses. C11 BODIPY was used to detect lipid peroxidation in cells after RT. RT-induced lipid peroxidation was further evaluated by staining mouse xenograft tumour and normal intestine tissues with 4-HNE. Finally, prussian blue staining was used to detect iron accumulation on both human tissue microarray and mouse tissues.

Results:

In cancer cells, we demonstrated that lipid peroxidation is induced by both conventional and FLASH RT in a dose dependent manner. Ferroptosis is a novel cell death pathway that depends on iron and lipid peroxidation. Our clonogenic assay with or without ferrostatin-1, a ferroptosis inhibitor, confirmed that both FLASH and conventional RT led to ferroptosis in cancer cells. Xenograft tumour tissue staining using 4-HNE, a marker for lipid peroxidation, further validated similar levels of lipid peroxidation by FLASH and conventional RT. Interestingly, in a normal cell line and healthy mouse intestine tissues, we found that FLASH-RT resulted in less lipid peroxidation when compared to conventional radiation. As an underlying mechanism, we hypothesise that differences in basal iron levels between normal and tumour tissues lead to the differential lipid peroxidation and ferroptosis. Our iron staining on patient tissue microarray confirmed higher levels of iron in tumours than in healthy tissues and mouse