Arsenic and 17-b-estradiol bind to each other and neutralize each other’s signaling effects Sukhdeep Kumar a , Tapan K. Mukherjee b , Purnananda Guptasarma a, * a Center for Protein Science, Design and Engineering (CPSDE), Department of Biological Sciences, Indian Institute of Science Education and Research (IISER) Mohali, Knowledge City, Sector-81, SAS Nagar, Punjab 140306, India b Department of Biotechnology, Maharishi Markandeshwar University, Mullana, Ambala, Haryana 133207, India article info Article history: Received 8 June 2016 Accepted 17 June 2016 Available online xxx Keywords: Arsenic Reproduction Endocrinology Estrogen Health Molecular recognition abstract We report that arsenic trioxide (ATO) and 17-beta-estradiol (E2) abolish each other’s independent cell signaling effects in respect of cell survival and proliferation/migration of breast cancer (MCF-7) cells. The possibility that this is due to binding of ATO to E2 was confirmed through difference absorption spec- troscopy, chromatography-coupled voltammometry and 1-D 1 H and 13 C NMR spectroscopy. Binding leads to attenuation of E2’s hydroxyl 1 H peaks at its C17 and C3 carbon positions. The results suggest that ATO and E2 can titrate each other’s levels, potentially explaining why sustained arsenic exposure tends to be associated with delays in age of menarche, advanced age of menopause, poorer sperm quality, higher overall morbidity in men, and lower incidences of breast cancer in women in some arsenic-contaminated areas. © 2016 Elsevier Inc. All rights reserved. 1. Introduction The semi-metal, arsenic, which contaminates water in many parts of the world [1,2] has been called both a poison [3] and a medicine [4] because it has been shown to have causative [5], curative [6], or protective [7] effects, in different types of cancer. Arsenic is also an acknowledged stimulator of reactive oxygen species generation pathways [8] and an acknowledged disruptor of endocrine function and reproductive fitness [9]. In humans and rodents, variously, exposure to arsenic results in delayed menarche [10,11], early menopause [12], poorer fertility and poor gamete quality in men [13,14]. These effects ostensibly owe to arsenic’s suspected dose-dependent ability to cause lowering of plasma levels of the steroid hormone, estrogen [15]. In principle, arsenic could cause lowering of estrogen levels by (a) affecting estrogen synthesis, (b) effecting estrogen degradation, or (c) by somehow neutralizing functional estrogen. Current opinion holds that arsenic acts either as an agonist or competitive antagonist of estrogen, depending on the latter’s endogenous levels [16,17], with the semi-metal interacting with specific cysteine residues located at the estrogen binding site of the estrogen receptor alpha (ER-a) [18]. Some reports propose that arsenic binds to ER-a to inactivate it [19]. Interestingly, other reports propose that arsenic instead binds to ER-a to reactivate it [20]. Arsenic has been called a ‘xenoestrogen’, capable of inducing effects similar to es- trogens and other known metal xenoestrogens, such as cadmium. Competitive inhibition experiments involving arsenic, estrogen and ER-a, have been thought to suggest that arsenic binds to ER-a with aK i of 5 nM [19]. Notably, we searched the literature and did not find any actual physical or structural evidence which demonstrates that arsenic binds to specific cysteine residues on ER-a, as has been postulated [18], or that arsenic-bound ER-a molecules are inactive and estrogen-non-responsive. We hold, therefore, that scope still exists for alternative and/or additional interpretations of the competitive inhibition data that we have referred to above [19]. One such alternative interpretation - which, surprisingly, appears to never have been proposed, or explored, previously - is the pos- sibility that arsenic binds directly to estrogen. Below, we provide both suggestive as well as direct evidence of arsenic’s binding to estrogen, using arsenic trioxide (ATO) and 17-b-estradiol (E2) as model reagents. The findings have important potential implications for human health, which are mentioned in the concluding section. 2. Materials and methods In-vitro culture of MCF-7 breast cancer cells: MCF-7 breast cancer * Corresponding author. E-mail address: guptasarma@iisermohali.ac.in (P. Guptasarma). Contents lists available at ScienceDirect Biochemical and Biophysical Research Communications journal homepage: www.elsevier.com/locate/ybbrc http://dx.doi.org/10.1016/j.bbrc.2016.06.087 0006-291X/© 2016 Elsevier Inc. All rights reserved. Biochemical and Biophysical Research Communications xxx (2016) 1e6 Please cite this article in press as: S. Kumar, et al., Arsenic and 17-b-estradiol bind to each other and neutralize each other’s signaling effects, Biochemical and Biophysical Research Communications (2016), http://dx.doi.org/10.1016/j.bbrc.2016.06.087