ESTRO 36 Abstract Book

S252 ESTRO 36 2017 _______________________________________________________________________________________________

Purpose or Objective Treatment resistance is an important cause of adverse outcome in breast cancer. Several mechanisms involved in resistance to either radiotherapy or endocrine therapy (tamoxifen) have been elucidated. Since tamoxifen resistant breast cancer cells were found to be less sensitive to irradiation, we aimed to investigate common pathways involved in radiotherapy and tamoxifen resistance. Material and Methods The estrogen receptor positive breast cancer cell line MCF7 was grown to radioresistance by exposing them to multiple fractions of 4 Gy irradiation, adding up to a total dose of at least 50 Gy (MCF7RT). Tamoxifen resistant MCF7 cells were created by culturing with gradually increasing concentrations of 4-hydroxy-tamoxifen up to 10 µM (MCF7TAM). Changes in expression profiles in MCF7RT and MCF7TAM cells compared to parental MCF7 cells were investigated by RNA sequencing, and common pathways identified. QPCR was used to confirm the RNA sequencing data, and to investigate expression of genes of interest after irradiation and tamoxifen treatment. Results Genes involved in interferon signaling were significantly increased in both MCF7RT and MCF7TAM compared to parental MCF7 cells. For further experiments, five interferon stimulated genes (ISGs; representing different parts of the signaling pathway) were evaluated, namely DDX60, STAT1, OAS1, IFI6 and IFI27. Differential expression of these five ISGs in the resistant MCF7 cells, and in treatment resistant T47D cells (also an estrogen receptor positive breast cancer cell line), was confirmed by qPCR. Expression of ISGs was induced by treatment: all five genes were increased 24h and 48h after irradiation with 4 Gy. Tamoxifen treatment (1 or 10 mM for 24h) also led to increased ISG expression. In patients treated with tamoxifen (KMplot, n=849), high expression of ISGs correlated with a worse outcome (figure 1). Additionally, in a cohort of advanced breast cancer patients (n=344), ISGs were co-expressed and correlated with a tumor infiltrating lymphocyte signature, which in turn was associated with a shorter time to progression after tamoxifen treatment in these patients.

Performed scans are summarized in table 1.

In total 22/32 patients (69%) showed hypoxia at baseline in primary tumor and/or lymph nodes. In the 11 hypoxic tumors a significant negative correlation was found between baseline HX4-TBR and tumor blood flow (r=- 0.618, p=0.043) and blood volume (r=-0.736, p=0.010). (Fig 1).

After nitroglycerin administration, 4 of the 15 patients (27%) with a baseline hypoxic tumor and/ or nodes that received a second HX4-scan could not be classified as hypoxic anymore. In the group of tumors with baseline hypoxia we found a significant negative correlation between the change in blood volume and HX4-HF (r=- 0.829, p=0.042) and HX4-TBR (r=-0.829, p=0.042). In non- hypoxic tumors there was no correlation between the HX4- TBR and either of the perfusion parameters. Conclusion In this trial 27 percent of patients with baseline hypoxic tumors and nodes become normoxic after treatment with nitroglycerin. Dynamic contrast enhanced CT-scans demonstrate that this effect on hypoxic tracer uptake is negatively correlated with an effect on tumor perfusion in hypoxic tumors. These results support the hypothesis that hypoxia scans and/or DCE-CT scans could form a tool to select patients for a nitroglycerin patch adjuvant to anti- cancer treatment (radiotherapy, chemotherapy, targeted agents or immunotherapy). An animation summarizing our results is available at https://youtu.be/udJSBYaRv9w. OC-0482 Interferon stimulated genes: a common pathway in tamoxifen- and radioresistance in breast cancer A. Post 1 , M. Smid 2 , A. Nagelkerke 1 , J. Martens 2 , J. Bussink 1 , C. Sweep 3 , P. Span 1 1 Radboud university medical center, Radiation Oncology, Nijmegen, The Netherlands 2 Erasmus MC Cancer Institute, Department of Medical Oncology and Cancer Genomics Netherlands, Rotterdam, The Netherlands 3 Radboud university medical center, Laboratory Medicine, Nijmegen, The Netherlands

Figure 1 Conclusion

We show here that expression of ISGs is increased in MCF7 cells resistant to radiotherapy or tamoxifen, compared to parental MCF7 cells. Since the expression of ISGs already increased after a single dose of irradiation or tamoxifen treatment, we hypothesize that after extended treatment the interferon signaling pathway is upregulated and confers a survival advantage to the cells, ultimately leading to treatment resistance. We are currently investigating the effect of pathway inhibition on the sensitivity of resistant MCF7 cells to irradiation and