Potential Antiestrogen-Based Cytotoxic Agents and an Affinity Label for the Estrogen Receptor
Wei, Lisa Lining
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Permalink
https://hdl.handle.net/2142/71436
Description
Title
Potential Antiestrogen-Based Cytotoxic Agents and an Affinity Label for the Estrogen Receptor
Author(s)
Wei, Lisa Lining
Issue Date
1984
Department of Study
Physiology and Biophysics
Discipline
Physiology
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Biology, Animal Physiology
Abstract
My thesis research involved studying the role of the estrogen receptor (ER) mediating the cytotoxicity of various tamoxifen derivatives in human breast cancer cells and examining estrogen structure and function using an affinity labeling compound, tamoxifen aziridine (TA).
Using cell proliferation and colony formation assays, derivatives of tamoxifen were evaluated for their ability to act as cytotoxic agents following binding to ER. Tamoxifen-nitrosocarbamate was an effective cytotoxic agent but its effects were not mediated by the ER.
Tamoxifen-nitrosourea in part by binding to ER inhibited cell growth and colony formation of MCF-7L cells. However, tamoxifen-nitrosourea also inhibited the growth and colony formation ability of the ER negative (MDA-MB-231) cells. Further studies indicated that a control compound which contains the nitrosourea moiety but does not bind to ER was harmful to the MDA-MB-231 cells and not to the MCF-7L cells. Thus, it is clear that the two cell lines differ in their sensitivity to tamoxifen-nitrosourea. In MCF-7L cells, tamoxifen-nitrosourea mediates its effects in part through ER and in MDA-MB-231 cells, tamoxifen-nitrosourea acts through an ER independent mechanism.
Tamoxifen aziridine was found to bind irreversibly to ER in cytosol preparations from rat uteri and MCF-7M cells. Binding of {('3)H}TA to ER was highly specific and covalent attachment to the receptor was very efficient. Eighty percent of the receptor in a uterine preparation was irreversibly labeled with {('3)H}TA within 1 hr at 25(DEGREES)C. TA represents the first highly efficient and selective affinity label for the ER. Using SDS gel electrophoresis, the {('3)H}TA labeled cytosolic and nuclear receptor had an M(,r) of approximately 60,000. The molecular weights of cytosolic ER from pituitary and dimethylbenzanthracene-induced mammary tumors were similar to the rat uterine receptor. The pI's of the cytosolic receptor from these three tissues were similar, ca. 6.0.
The biological activity of TA was also examined. TA suppressed uterine weight stimulated by estradiol, inhibted dimethylbenzathracene-induced mammary tumor growth, inhibited cell proliferation of MCF-7L cells, and antagonized estradiol stimulation of plasminogen activator activity. By these criteria, TA behaves as an antiestrogen.
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