Date of Award

2018

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Marine Biomedical and Environmental Sciences

College

College of Graduate Studies

First Advisor

Demetri Spyropoulos

Second Advisor

John Baatz

Third Advisor

Sherine Chan

Fourth Advisor

Erik Svendsen

Fifth Advisor

Gary Hardiman

Abstract

Individuals living in industrialized regions of Westernized societies are exposed to environmental contaminants by many routes, including plastics, personal care items and other consumer products. Endocrine-active chemicals (EACs), many of which are estrogenic, have been detected in human biofluids and breast tissue, warranting investigation of roles in mammary tumorigenesis. This dissertation explores the molecular changes that occur in breast cancer cells upon treatment with relevant human exposure levels of bisphenol-A (BPA), methylparaben (MP), propylparaben (PP), and decabromodiphenyl ether (DBDE), including mixtures thereof with or without 17β-estradiol (E2). Nanomolar (nM) concentrations of these EACs induced viability increases in MCF-7 (ERα+) similar to picomolar (pM) E2 concentrations, but EAC mixtures did not produce additive effects. MDA-MB-231 viability was unaltered by EACs in the absence of E2 but was significantly increased with exposure to five EACs combined with 50 pM E2. To examine ERα gene regulation, transactivation assays in ERα-transfected HepG2 cells confirmed that nanomolar BPA could induce ER-driven transcription, and suggested an additive effect of EAC mixtures. Non-genomic ERα functions were also investigated in breast cancer cells via high-throughput microscopy and quantitative immunofluorescence with the Hermes/WiScan System. MCF-7 cells revealed a 50% increase in phosphorylated (P)-ERα Serine 167 expression after 30-minute exposure to 100 nM PP, followed by alterations in nuclear/cytosolic localization after 24 hours. Signaling/epigenetic pathways were assessed with fifteen additional protein markers. At 100 nM, EACs induced expression of P-ERK, β-catenin, and epigenetic marks at H3K4me2, H3K9acetyl, and H3K27me2, while decreasing levels of AR and LSD1. Similarities and differences in proteomic expression patterns were observed between E2 and EACs. These pathways were further found to influence breast cancer progression, drug response and resistance; EACs were shown here to increase Tamoxifen IC50 in MCF-7 cells, but further sensitized them to Doxorubicin toxicity. A proteomic marker shown to undergo nuclear localization changes during observed EAC-altered Tamoxifen responses was P-ERα-S167. After extended low-level EAC exposures (60 days), a four-day EAC-withdrawal was sufficient to reset MCF-7 drug responses to control levels. These results reveal that EACs have diverse cellular mechanisms involving genomic and proteomic ERα effects and signaling/epigenetic alterations, highlighting these EACs as important environmental considerations for breast cancer research, clinical care and prevention.

Rights

All rights reserved. Copyright is held by the author.

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