Purpose To develop a guideline to improve the accuracy of immunohistochemical (IHC) estrogen receptor (ER) and progesterone receptor (PgR) testing in breast cancer and the utility of these receptors as predictive markers. Methods The American Society of Clinical Oncology and the College of American Pathologists convened an international Expert Panel that conducted a systematic review and evaluation of the literature in partnership with Cancer Care Ontario and developed recommendations for optimal IHC ER/PgR testing performance. Results Up to 20% of current IHC determinations of ER and PgR testing worldwide may be inaccurate (false negative or false positive). Most of the issues with testing have occurred because of variation in preanalytic variables, thresholds for positivity, and interpretation criteria. Recommendations The Panel recommends that ER and PgR status be determined on all invasive breast cancers and breast cancer recurrences. A testing algorithm that relies on accurate, reproducible assay performance is proposed. Elements to reliably reduce assay variation are specified. It is recommended that ER and PgR assays be considered positive if there are at least 1% positive tumor nuclei in the sample on testing in the presence of expected reactivity of internal (normal epithelial elements) and external controls. The absence of benefit from endocrine therapy for women with ER-negative invasive breast cancers has been confirmed in large overviews of randomized clinical trials.

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American Society of Clinical Oncology/College of American Pathologists Guideline Recommendations for Immunohistochemical Testing of Estrogen and Progesterone Receptors in Breast Cancer

Although it is widely accepted that specific intracellular receptor proteins are involved in the oestrogenic regulation of gene expression and growth in reproductive tissues, the precise nature of the regulation is poorly understood. Among the unresolved issues are the distribution and dynamics of the oestrogen receptor protein (oestrophilin) in target tissues in the presence and absence of oestrogens and antioestrogens. The use of radio-labelled and unlabelled receptor ligands to detect and measure oestrogen receptors in tissues has been complicated by the presence of other intracellular steroid-binding proteins1 and by the low concentration of receptors in responsive tissues. We report here the development of an immunocytochemical procedure that is suitable for localizing oestrophilin directly in frozen tissue sections or cells from human and several non-human sources. When monoclonal antibodies to oestrophilin were used to detect receptor in various oestrogen-sensitive tissues, specific staining was confined to the nucleus of all stained cells, suggesting that both cytosol and nuclear forms of the receptor protein may reside in the nuclear compartment.

Monoclonal antibodies localize oestrogen receptor in the nuclei of target cells.

According to the current model of steroid hormone action oestrogen is thought to bind to its receptor in the cytoplasm of target cells1,2 and the oestrogen–receptor complex is then translocated into the nucleus2–4. This model is based on evidence obtained in homogenized cell preparations in which free receptor is associated with the cytosol, whereas steroid-bound receptor is associated with the nuclear fraction. Some data suggest, however, that the unfilled receptor may reside in the nucleus, and that cytosolic localization represents an extraction artefact. We have now reinvestigated the subcellular distribution of unfilled oestrogen receptor using cytochalasin B-induced enucleation to obtain cytoplast and nucleoplast fractions from receptor-containing GH3 cells derived from rat pituitary tumours. We found that cytoplasts prepared from GH3 cells contain little oestrogen-binding activity and that most of the unfilled oestrogen receptors are associated with the nuclear fraction. We therefore suggest that the standard model is in error and that the unoccupied receptor is nuclear in the intact cell.

Nuclear localization of unoccupied oestrogen receptors.

Purpose To develop a guideline to improve the accuracy of immunohistochemical (IHC) estrogen receptor (ER) and progesterone receptor (PgR) testing in breast cancer and the utility of these receptors as predictive markers. Methods The American Society of Clinical Oncology and the College of American Pathologists convened an international Expert Panel that conducted a systematic review and evaluation of the literature in partnership with Cancer Care Ontario and developed recommendations for optimal IHC ER/PgR testing performance. Results Up to 20% of current IHC determinations of ER and PgR testing worldwide may be inaccurate (false negative or false positive). Most of the issues with testing have occurred because of variation in pre-analytic variables, thresholds for positivity, and interpretation criteria. Recommendations The Panel recommends that ER and PgR status be determined on all invasive breast cancers and breast cancer recurrences. A testing algorithm that relies on accurate, reproducible assay performance is proposed. Elements to reliably reduce assay variation are specified. It is recommended that ER and PgR assays be considered positive if there are at least 1% positive tumor nuclei in the sample on testing in the presence of expected reactivity of internal (normal epithelial elements) and external controls. The absence of benefit from endocrine therapy for women with ER-negative invasive breast cancers has been confirmed in large overviews of randomized clinical trials.

Linkage analysis of ten Utah kindreds and one Texas kindred with multiple cases of cutaneous malignant melanoma (CMM) provided evidence that a locus for familial melanoma susceptibility is in the chromosomal region 9p13-p22. The genetic markers analyzed reside in a candidate region on chromosome 9p21, previously implicated by the presence of homozygous deletions in melanoma tumors and by the presence of a germline deletion in an individual with eight independent melanomas. Multipoint linkage analysis was performed between the familial melanoma susceptibility locus (MLM) and two short tandem repeat markers, D9S126 and the interferon-alpha (IFNA) gene, which reside in the region of somatic loss in melanoma tumors. An analysis incorporating a partially penetrant dominant melanoma susceptibility locus places MLM near IFNA and D9S126 with a maximum location score of 12.71. Therefore, the region frequently deleted in melanoma tumors on 9p21 presumably contains a locus that plays a critical role in predisposition to familial melanoma.

https://science.sciencemag.org/content/sci/258/5085/1148.full.pdf

Assignment of a locus for familial melanoma, MLM, to chromosome 9p13-p22.

KLE: a cell line with defective estrogen receptor derived from undifferentiated endometrial cancer.

Immunohistochemical localization of estrogen receptors in human mammary carcinoma using antibodies to the receptor protein

Estrogen receptor assays were performed on tumor specimens from 6 male patients with documented breast cancer. Five out of six (83%) males had positive ER assays with values ranging from 23-384 femtomoles per mg cytosol protein. A review of the literature showed that 32 out of 38 patients (84%) reported were positive for estrogen receptor. Four out of five ER positive males responded to orchiectomy. Estrogen receptors are present in a high percentage of males with breast cancer and may predict response to treatment, similar to female breast cancer.

https://acsjournals.onlinelibrary.wiley.com/doi/pdf/10.1002/1097-0142%2819801015%2946%3A8%3C1781%3A%3AAID-CNCR2820460814%3E3.0.CO%3B2-F

Estrogen receptors in male breast cancer

Estrogen receptor assay: Interlaboratory and intralaboratory variations in the measurement of receptors using dextran-coated charcoal technique: A study sponsored by E.C.O.G.

Neither estrogen dependence nor clinical response to hormone therapy of the tumor is guaranteed by the mere presence of receptors for estrogens (ERs) or progesterones (PRs). Immunohistochemical staining of ER with polyclonal anti-ER antibodies after in vitro transformation has enabled the identification of two types of defective ER among human breast cancers: those that are unable to bind to the nucleus in a hormone-filled state and those that bind to the nucleus as naked ER. Preliminary clinical correlation studies demonstrate that a subclassification of ER(+) tumors based on functional abnormalities of ER may predict refractoriness to hormone therapy.

Shanthi Raam

Papers

KLE: a cell line with defective estrogen receptor derived from undifferentiated endometrial cancer.

Immunohistochemical localization of estrogen receptors in human mammary carcinoma using antibodies to the receptor protein

Estrogen receptors in male breast cancer

Estrogen receptor assay: Interlaboratory and intralaboratory variations in the measurement of receptors using dextran-coated charcoal technique: A study sponsored by E.C.O.G.

Defective estrogen receptors in human mammary cancers: their significance in defining hormone dependence.