It has been shown that the risk of breast cancer developing in certain morphologically identifiable benign breast lesions correlates with expression of estrogen receptor alpha (ER-alpha). Although ER-alpha and ER-beta genes share a large degree of homology, it is generally thought that their distribution and functions are substantially different in many tissues. Recent development of reliable antibodies to ER-beta has provided this first opportunity to test the hypothesis that the likelihood of malignant transformation in morphologically benign breast lesions can be accurately defined by the distribution and level of ER-beta expression relative to that of ER-alpha. Using a monoclonal antibody, ER-beta protein expression has been analyzed in 53 normal breasts and compared with a cohort of histologically distinct breast lesions of different prognostic risk (54 hyperplasia of usual type, 35 ductal carcinoma in situ, and 141 invasive cancers). All of these tissues were also assessed for ER-alpha. Expression of ER-beta protein was also analyzed in an additional spectrum of benign breast lesions with low or negligible risk of progression to malignancy. The median proportion of cells expressing ER-beta was highest in normal breast lobules (median 94.33%, interquartile range 78.25-99.00) but declined significantly through usual ductal hyperplasia (median 76.67, interquartile range 49.17-95.00, P = 0.002) and ductal carcinoma in situ (median 70.00, interquartile range 59.00-85.00, P = 0.009) to invasive cancer (median 60.00, interquartile range 50.00-80.00, P < 0.001). An appreciable proportion (33.81%) of ER-alpha-negative invasive cancers expressed ER-beta. A high but variable level of ER-beta expression occurred in the benign lesions. The data from the intact histologic tissues were evaluated with respect to the relative expression of ER-alpha and ER-beta in five mammary cell lines of different behavioral phenotype (MCF7, ZR-75, T47D, MDAMB231, HUMA121). The highly significant differences in expression and distinct tissue distributions of ER-alpha and ER-beta within the histologic lesions of defined risk, together with the data from the cell lines, support the original hypothesis that the tissue concentration, relative occurrence, and/or interaction of these two types of estrogen receptor may play an important role in modulating mammary tumorigenesis.

This study has tested the hypothesis that comparison of protein and mRNA expression for ERa and ERb1 by human breast cancers provides novel information relating to the clinical and pathological characteristics of human breast cancers. Expression of ERa and ERb1 was identified in 167 invasive cancers from postmenopausal women treated only with endocrine therapy. The cohort included 143 cases receiving only adjuvant Tamoxifen following surgery. ERa and ERb1 expression was analysed by immunohistochemistry and reverse transcription RT -PCR and compared with clinical progression of individual cancers. ERa protein was closely associated with the corresponding RNA detected by RT -PCR (Chi-square, Po0.001). In contrast, ERb1 protein and mRNA were inconsistent. Although an association was identified between ERa and ERb mRNAs (Chi-square, Po0.001) and between ERa protein and ERb1 mRNA (Chi-square, Po0.027), no association was identified for the ERa and ERb1 proteins detected by immunohistochemistry. ERb1 was not associated with outcome. However, in the absence of ERa, ERb1 protein expression was associated with elevated cell proliferation. There was a trend for the ERb1 protein-positive cases to have a worse outcome, both within the group as a whole as well as within the ERa-positive Tamoxifen-treated cases. This study has confirmed the hypothesis that expression of ERa is an important determinant of breast cancer progression, and has further demonstrated that ERb1 may play a role in the response of breast cancers to endocrine therapy.

The anterior gradient protein-2 (AGR2) is inducible by oestrogen and itself can induce metastasis in a rat model for breast cancer. Here, a rabbit antibody to recombinant human AGR2 was used to assess its prognostic significance in a retrospective cohort of 351 breast cancer patients treated by adjuvant hormonal therapy. The antibody stains 66% of breast carcinomas to varying degrees. The percentage of positive carcinoma cells in tumours directly correlates with the level of AGR2 mRNA (Spearman's rank correlation, P ¼ 0.0007) and protein (linear regression analysis r 2 ¼ 0.95, P ¼ 0.0002). There is a significant association of staining of carcinomas for AGR2 with oestrogen receptor a (ERa) staining and with low histological grade (both Fisher's Exact test Po0.0001). In the ERapositive cases, but not the ERa-negative cases, when subdivided into the separate staining classes for AGR2, there is a significantly progressive decrease in patient survival with increased staining (log rank test, P ¼ 0.006). The significant association of staining for AGR2 with patient death over a 10-year period (log rank test P ¼ 0.007, hazard ratio ¼ 3) only becomes significant at 6 years of follow-up. This may be due to the cessation of adjuvant hormonal therapy at an earlier time, resulting in adverse re-expression of the metastasis-inducing protein AGR2.