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Subcellular and extracellular localization of specific binding sites for triphenylethylene antiestrogens in human
Abstract:
MCF-7 human breast cancer cell homogenates and subcellular organelles were submitted to isopycnic centrifugation on Percoll gradients to investigate the subcellular localization of triphenylethylene antiestrogen specific binding sites (AEBS). Electron microscopy revealed that gradient fractions coincident with the migration of [3H]tamoxifen-AEBS complexes were homogeneously represented by rough and smooth endoplasmic reticulum. Eighty percent of AEBS were localized in the endoplasmic reticulum [45,000 +/- 4,000 sites/cell, mean +/- S.D.), while 20% of these sites were also found in the nuclear fraction (12,000 +/- 1,000 sites/cell, mean +/- S.D.). A similar subcellular distribution of AEBS was observed in human breast cancer bioptic specimens. No differences in [3H]tamoxifen binding affinity between microsomal and nuclear AEBS were observed in MCF-7 and bioptic breast cancer. No major differences in microsomal AEBS levels were observed in the limited number of estrogen receptor-positive or -negative breast cancer specimens we have studied, whereas estrogen receptor-negative samples had higher levels of nuclear AEBS with respect to estrogen receptor-positive tumors. The presence of AEBS was also detected in the human serum of healthy and tumor-bearing subjects. The affinity and the binding specificity of serum AEBS were similar to those of intracellular AEBS. No differences in the levels of serum AEBS were observed between healthy and tumor-bearing subjects [19 +/- 4 and 22 +/- 4 pmoles/ml (mean +/- S.D.) respectively. Human serum AEBS did not appear to be associated to lipoproteins, whereas it migrated as a 5.5 S sedimenting molecule.
Insights
Triphenylethylene antiestrogen binding sites (AEBS) are primarily located in the endoplasmic reticulum and nucleus of breast cancer cells. Serum AEBS show similar binding properties to intracellular AEBS, with no significant differences between healthy and tumor-bearing subjects.
Area of Science:
- Cell Biology
- Endocrinology
- Oncology
Background:
- Antiestrogens are crucial in breast cancer therapy.
- Understanding the cellular localization of antiestrogen binding sites (AEBS) is key to their mechanism of action.
Purpose of the Study:
- To investigate the subcellular localization of triphenylethylene antiestrogen specific binding sites (AEBS) in human breast cancer.
- To compare AEBS distribution and binding characteristics in cell lines, tumor specimens, and serum.
Main Methods:
- Isopycnic centrifugation on Percoll gradients for subcellular fractionation.
- Isotopic labeling with [3H]tamoxifen to detect AEBS.
- Electron microscopy for fraction identification.
- Analysis of AEBS in MCF-7 cells, breast cancer biopsies, and human serum.
Main Results:
- AEBS were predominantly found in the endoplasmic reticulum (80%) and nucleus (20%) of MCF-7 cells and breast cancer biopsies.
- No significant differences in [3H]tamoxifen binding affinity were observed between microsomal and nuclear AEBS.
- Estrogen receptor-negative tumors showed higher levels of nuclear AEBS compared to estrogen receptor-positive tumors.
- AEBS were detected in human serum with similar affinity and specificity to intracellular AEBS, with no significant difference in levels between healthy and tumor-bearing subjects.
Conclusions:
- The endoplasmic reticulum and nucleus are the primary intracellular locations for AEBS in breast cancer.
- Serum AEBS exhibit characteristics similar to intracellular AEBS, suggesting a potential role or marker.
- Nuclear AEBS levels may differ based on estrogen receptor status in breast tumors.