Tamoxifen aziridine binding to cytosolic proteins from human breast specimens is negatively associated with estrogen receptors, progesterone receptors, pS2, and cathepsin-D |
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Authors: | Domingo Navarro Hilario Doreste Juan J Cabrera Manuel Morales Juan C Díaz-Chico Bonifacio N Díaz-Chico |
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Institution: | (1) Laboratorio de Fisiología, Dept. Endocrinología Celular y Molecular, Centro de Ciencias de la Salud, Universidad de Las Palmas de Gran Canaria, Spain;(2) Servicio de Anatomía Patológica, Hospital Insular de Gran Canaria, Spain;(3) Servicio de Oncología Médica, Hospital de La Candelaria, Santa Cruz de Tenerife, Islas Canarias, Spain |
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Abstract: | 3H]Tamoxifen Aziridine (3H]TAZ) is a derivative of the antiestrogen tamoxifen that covalently labels the Estrogen Receptor (ER), and perhaps other uncharacterized proteins. In a previous article we described that 3H]TAZ binds to a cytosolic protein from human uterine tissues that shares some, but not all, the ER properties. Here we have extended these studies to 3H]TAZ binding to cytosol proteins from human breast cancer specimens, and studied its quantitative association with other molecular markers and clinico-pathological variables. Cytosols were obtained in hypotonic buffer containing 20 mM molybdate and protease inhibitors, incubated with 3H]TAZ, and subjected to Sucrose Gradient Analysis (SGA). A 3H]TAZ labeled peak that consistently migrated with the 4S fractions was found in most of the assayed cytosols (range of 0 to 1278 fmol/mg p.). The 4S peak of 3H]TAZ was partially inhibited by both estrogens and antiestrogens. When 3H]E2 was used instead of 3H]TAZ, only an 8S peak was detected. 3H]TAZ was covalently bound to a protein with an apparent MW of 65 kDa, as determined by SDS-PAGE and fluorography. The mean of 3H]TAZ binding was significantly higher in the subgroups of samples classified as ER-, PR-, pS2- or cathepsin D-, than in the respective positive subgroups (P < 0.01 in all the cases). 3H]TAZ binding was not associated with clinico-pathological variables, except that its mean was significantly larger in tumors larger than 5 cm than in smaller tumors. These results, and those previously reported, suggest that: 1) 3H]TAZ labels a cytosolic protein present in human breast cancers and uterine tissues that does not share all the ER properties, and 2) the 3H]TAZ binding by breast cancer cytosols is negatively associated with markers of estrogenic dependency, and its quantification may provide valuable information on antiestrogen responsiveness of a given tumor. |
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Keywords: | breast cancer estrogen receptors progesterone receptor pS2 cathepsin-D native receptor tamoxifen aziridine |
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