ABSTRACT
Purpose: The estrogen receptor (ER) is expressed in over 80% of breast tumors and has been shown to be a significant driver of breast cancer (BC) pathogenesis and therefore a target of effective first-line therapies. While both ionizing radiation (RT) and endocrine therapies (ET) are used for the treatment of ER+ BC, the effect of ET on tumor radiosensitization remains unclear, with concerns it may be radioprotective based on G1 cell arrest with ET treatment. Here we assessed the efficacy and mechanism of ER-mediated radiosensitization using various pharmacologic approaches in ER+ BC. Methods: Radiosensitization with ER inhibitors (tamoxifen [TAM], fulvestrant [FULV], AZD9496) was assessed using clonogenic survival assays. DNA damage was assessed by the neutral comet assay. Efficiency of homologous recombination (HR) or non-homologous end joining (NHEJ) as well as changes in cell cycle, apoptosis, and senescence were assessed. The efficacy of TAM with RT in vivo was assessed with an MCF-7 xenograft model. Results: The selective estrogen receptor modulator TAM radiosensitized ER+ MCF-7 (enhancement ratio [enhR]: 1.14-1.50) and T47D (enhR: 1.33-1.60) cells but not ER-negative SUM-159 cells (enhR: 0.99-1.02). The selective estrogen receptor degrader (SERD) FULV had similar radiosensitization effects in MCF-7 (enhR: 1.33-1.76) and T47D cells (enhR: 0.97-2.81) with no radiosensitization observed in SUM-159 cells (enhR: 1.01-1.03). The novel oral SERD AZD9496 radiosensitized MCF-7 cells (enhR: 1.36-1.56). MCF-7 cells treated with TAM and RT had an increase in dsDNA breaks compared to RT alone as measured by the comet assay (p<0.05) and a decrease in NHEJ-mediated repair with TAM (p<0.05). No changes were observed in HR-mediated repair by Rad51 foci or a reporter (p=NS). RT alone and in combination with TAM or FULV induced similar levels of cell cycle arrest, suggesting that radiosensitization with the combination therapy is cell-cycle independent. There were no significant changes in apoptosis with TAM, FULV, RT, or the combination (p=NS). Although TAM or FULV did induce senescence, ET with RT increased senescence induction (p<0.05). In vivo, combination RT and TAM led to a significant delay in days to tumor doubling (control: 17, TAM: 40, RT: 32, TAM+RT: undefined;p<0.0001), and a significant difference in tumor growth between mice treated with TAM or RT alone compared combination treatment, with no increased toxicities or skin lesions from the combination treatment. Conclusion: Our data suggest that TAM, FULV, or AZD9496 can radiosensitize ER+ breast tumors, and these agents with RT may be more effective for radiosensitization. This work also supports further clinical investigation of the timing of RT for patients receiving ET, including using ET during RT, especially as initiating ET prior to RT has been increasingly utilized as a bridging therapy followed by concurrent ET+RT during the COVID-19 pandemic.