M. Zhou; Center Hospital Chinese Academy of Medical Sciences, Shenzhen Center, Shenzhen, China
Purpose/Objective(s): Considerable evidence suggests that breast cancer therapeutic resistance and relapse can be driven by polyploid giant cancer cells (PGCCs). PGCCs are involved in tumor repopulation following radiotherapy. Although the majority of PGCCs induced by irradiation underwent cell death, some exhibited proliferative capacity, and the number increases with the stages of disease and therapeutic stress. Given the importance of PGCCs, it remains challenging to dig effective anti-PGCCs therapy. In this study, we propose identifying promising radiotherapy with repurposed old drugs combination therapy to inhibit all cancer cells or only PGCCs. In this manner, we screen a drug library combined with irradiation and identify promising combination therapy that inhibit all cancer cells or only PGCCs (e.g., regulators of HDAC, proteasome, and ferroptosis). Materials/
Methods:
(1)PGCCsInducing: The breast cancer cell line MDA-MB-231, SUM-159, SUM-149 cells were irradiated with various doses X-rays (0Gy, 2Gy, 4Gy, 8Gy,10 Gy) using a linear accelerator, after 5 day culture, we harvest irradiation-induced PGCCs and do drug screening experiments.
(2)Drug library Setting: We make up a library including 4000 compounds (NIH Anti-cancer Compound Library (96-well)-L3000) and small targeting compounds).
(3) Combination therapy screening and PGCCs cell toxicity test: We screen the drug library, the drug combination and irradiation combined with drug library. Then based on the single-cell morphological analysis pipeline for the swift differentiation of compounds targeting non-PGCCs, PGCCs, or both, which was published in the previous study, we identify PGCCs and make PGCCs cell toxicity analysis to find out promising anti-PGCCs treatments.
(4) Statistical analyses were conducted using R (version 4.1), scientific 2-D graphing and statistics software, and a programming environment. Two-tailed Student’s t-test compared two groups, while paired 2-way ANOVA and Fisher’s Least Significant Difference (LSD) test compared multiple groups, significance set at P < 0.05 (*P < 0.05, **P < 0.01, ***P < 0.001).
Results: (1) PGCCs existed after various dose of irradiation(2,4,8,10Gy ), and the percentage of these PGCCs grew rapidly in the following days, reaching peak value around day 5. (2) Irradiation combined with some drugs could kill PGCCs induced by irradiation, such as Bortezomib, MG-132, Carfilzomib, Ixazomib (proteasome inhibitors), Imidazole ketone erastin (IKE), RSL3, FINO2, ML162, ML210 (ferroptosis inducers), Vorinostat(SAHA), Romidepsin, Panobinostat (HADC inhibitors). Conclusion: While the importance of PGCCs in cancer treatment resistance has been widely acknowledged, there is no effective way to eradicate them. This study identified 3 main classes of compounds effectively killed irradiation induced PGCCs combined with irradiation: HDAC inhibitors, proteasome inhibitors, and ferroptosis inducers. Further mechanistic and in vivo investigations will be performed in our future research endeavors inducers.
