COMPARING THE EFFECTS OF PROTON AND PHOTON THERAPY ON TUMOR AGGRESSIVENESS IN ADVANCED CANCERS

Radiotherapy, a cornerstone of cancer treatment, can paradoxically induce increased tumor aggressiveness in some cases, complicating patient outcomes. This study compares the effects of proton and photon therapies on the aggressiveness of cancer cells that survive radiation treatments. We examined two types of advanced cancer known for poor prognosis and high recurrence rates: Glioblastoma Multiforme (GBM) and High-Grade Serous Ovarian Cancer (HGSOC). Given the aggressive nature of these cancers and the limited success of available treatments, there is a critical need to develop new strategies that utilize radiation therapy but minimize the adverse effects. Photon radiation, the conventional treatment, utilizes high-energy photons to ionize biomolecules and disrupt cellular functions, leading to cell death. Although photon radiation is effective in killing cancer cells, it is also linked to the development of secondary cancers and more aggressive cancer phenotypes that become resistant to treatment. On the other hand, proton radiation, a more targeted therapy, minimizes collateral damage to surrounding healthy tissues by using hydrogen ions to deliver precise radiation doses to tumors. To compare radiation-induced tumor aggressiveness between proton and photon radiation, we examined pluripotency, epithelial-mesenchymal transition (EMT), and tumor immune evasion traits in irradiated HGSOC and GBM cells. We integrated a SORE6-GFP reporter to identify cancer stem cell (CSC) populations expressing SOX2/OCT4 and a 3’ UTR-ZEB1-GFP reporter to detect mesenchymal cell populations. Transduced cells were treated with 0, 1, 2, 4, and 8 Gy of 250 MeV proton and 6 MeV photon beams, and GFP levels were measured via flow cytometry 72 hours post-radiation. By calculating the ratio of live GFP reporter-expressing cells to the total number of live cells, we observed an increase in the ratio of CSCs and mesenchymal cells following both proton and photon treatments. RT-qPCR analysis assessed gene expression changes in irradiated cells, examining pluripotency genes POU5F1, SOX2, LIN28A, and EMT transcription factors ZEB1, ZEB2, SNAI1, and SLUG. To assess the potential for immune evasion in cells following radiation, we conducted RT-qPCR analysis to examine changes in the expression levels of immune checkpoint genes PD-L1 and HLA-G, as well as MHC class I genes HLA-A, HLA-B, and HLA-C. To determine which type of radiation induced a more immune-favorable response, we developed an Immune Response Score to measure the ratio of immune checkpoint gene expression to MHC class I gene expression. Overall, proton radiation resulted in a more immune-favorable response in cancer cells, with the exception of LN18 cells. This project further examined the responses of a patient-derived HGSOC cell line and its cisplatin-resistant subline to proton and photon radiation treatments. We incorporated the SORE6-GFP reporter, the 3’ UTR-ZEB1-GFP reporter, and an apoptosis assay. Our data indicate that the cisplatin-resistant subline showed greater resistance to both types of radiation compared to the cisplatin-sensitive subline; however, proton radiation induced a higher rate of cell death. When calculating the ratio of live GFP reporter-expressing cells to the total number of experimental cells, we observed that photon radiation therapy increased CSC and mesenchymal cell populations, while proton radiation therapy did not.