Abstract 1602: Effects of Letrozole and radiotherapy on cell cycle and death in a breast cancer cell line.
Authors: Durmak Isman Burcu , Gokturk Dilek , Sercan Zeynep , Ates Halil , Karaguler Zafer , Bilkay Gorken Ilknur 
Published: August 2014

Abstract
Preclinical and clinical studies have shown that hormone receptor-positive tumors have molecular, biological and clinical differences. Tamoxifen is the standard endocrine therapy in premenopausal women, whereas in ER (+), postmenopausal patients the optimal adjuvant hormonal therapy is aromatase inhibitors either as initial therapy or after tamoxifen treatment. Studies have shown that concurrent use of tamoxifen sensitizes cells to radiotherapy (RT) and increases RT induced pulmonary fibrosis. We aimed to investigate biological effects of the concurrent use of letrozole with radiotherapy on the breast cancer cells; using cell lines MCF7, which does not express the aromatase enzyme; and MCF7aro which is a aromatase expressing subclone. Cells were exposed to letrozole at varying doses of 100, 500, 1000 nM; for 24, 72 and 144 hours. Irradiations were performed using a cobalt-60 source with doses 2 and 4 Gy. Cell death was evaluated by measuring caspase 3 activation and by cell surface annexin V / PI staining. Beclin expression levels were determined by western blot. Cell cycle analyses were performed by DNA content staining/flow cytometry. All experiments were done in triplicates. Caspase-3 and annexin-V / PI results showed no significant difference for early apoptosis, late apoptosis and necrosis between the letrozole treated and untreated aromatase non-expressing MCF7 cells. In the aromatase expressing MCF7aro cells, we observed that there was a general reduction in cell death in cells treated with letrozole; with a trend towards apoptosis as a cell death modality rather than necrosis. We also observed increased beclin protein levels -which may imply an increase in autophagy- in MCF7 cells which have received 4 Gy irradiation, independent of the presence of letrozole indicating an acute stress response when expose to high doses of radiation. On the other hand beclin levels were increased in all experimental groups of MCF-7aro cells independent of the presence letrozole and/or radiation dose which supports the notion of an autophagic response under stress. The presence of the aromatase gene itself may be a positive factor for cell survival under stress conditions. Cell cycle analyses revealed 5% more cells accumulated at G1 in letrozole treated MCF7aro cells. Our preliminary findings suggest a reduction of cell death in hormone-sensitive, receptor-positive and aromatase enzyme expressing cells, after concurrent use of letrozole and RT; with apoptosis being the primary cell death modality. This observation correlates with our findings that autophagy may have also increased (based on beclin protein levels) in these cells. It is still not clear whether letrozole treated aromatase expressing cells have a survival advantage when treated concurrently with letrozole and RT. More extensive studies are needed to be able to evaluate the effects of the concurrent use of letrozole and radiotherapy on tumor cell death.
Citation Format: Burcu Durmak Isman, Dilek Gokturk, Zeynep Sercan, Halil Ates, Zafer Karaguler, Ilknur Bilkay Gorken. Effects of Letrozole and radiotherapy on cell cycle and death in a breast cancer cell line. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1602. doi:10.1158/1538-7445.AM2013-1602