Nitric oxide is cytoprotective to breast cancer spheroids vulnerable to estrogen-induced apoptosis

// Yana Shafran 1, * , Naomi Zurgil 1, * , Orit Ravid-Hermesh 1 , Maria Sobolev 1 , Elena Afrimzon 1 , Yaron Hakuk 1 , Asher Shainberg 2 and Mordechai Deutsch 1 1 The Biophysical Interdisciplinary Jerome Schottenstein Center for the Research and Technology of the Cellome, Physics Department, Bar Ilan University, Ramat Gan 52900, Israel 2 The Mina and Everard Goodman Faculty of Life Sciences, Bar Ilan University, Ramat Gan 52900, Israel * These authors have contributed equally to this work Correspondence to: Mordechai Deutsch, email: motti.jsc@gmail.com Keywords: nitric oxide; breast cancer spheroids; estrogen-induced apoptosis; estrogen receptor-positive; live-cell imaging Received: February 15, 2017 Accepted: August 17, 2017 Published: October 07, 2017 ABSTRACT Estrogen-induced apoptosis has become a successful treatment for postmenopausal metastatic, estrogen receptor-positive breast cancer. Nitric oxide involvement in the response to this endocrine treatment and its influence upon estrogen receptor-positive breast cancer progression is still unclear. Nitric oxide impact on the MCF7 breast cancer line, before and after estrogen-induced apoptosis, was investigated in 3D culture systems using unique live-cell imaging methodologies. Spheroids were established from MCF7 cells vulnerable to estrogen-induced apoptosis, before and after exposure to estrogen. Spheroids derived from estrogen-treated cells exhibited extensive apoptosis levels with downregulation of estrogen receptor expression, low proliferation rate and reduced metabolic activity, unlike spheroids derived from non-treated cells. In addition to basic phenotypic differences, these two cell cluster types are diverse in their reactions to exogenous nitric oxide. A dual effect of nitric oxide was observed in the breast cancer phenotype sensitive to estrogen-induced apoptosis. Nitric oxide, at the nanomolar level, induced cell proliferation, high metabolic activity, downregulation of estrogen receptor and enhanced collective invasion, contributing to a more aggressive phenotype. Following hormone supplementation, breast cancer 3D clusters were rescued from estrogen-induced apoptosis by these low nitric oxide-donor concentrations, since nitric oxide attenuates cell death levels, upregulates survivin expression and increases metabolic activity. Higher nitric oxide concentrations (100nM) inhibited cell growth, metabolism and promoted apoptosis. These results suggest that nitric oxide, in nanomolar concentrations, may inhibit estrogen-induced apoptosis, playing a major role in hormonal therapy. Inhibiting nitric oxide activity may benefit breast cancer patients and ultimately reduce tumor recurrence.