Male breast cancer remains understudied despite evidence of rising incidence. Using a co-ordinated multi-centre approach, we present the first large scale biomarker study to define and compare hormone receptor profiles and survival between male and female invasive breast cancer. We defined and compared hormone receptor profiles and survival between 251 male and 263 female breast cancers matched for grade, age, and lymph node status. Tissue microarrays were immunostained for ERα, ERβ1, -2, -5, PR, PRA, PRB and AR, augmented by HER2, CK5/6, 14, 18 and 19 to assist typing. Hierarchical clustering determined differential nature of influences between genders. Luminal A was the most common phenotype in both sexes. Luminal B and HER2 were not seen in males. Basal phenotype was infrequent in both. No differences in overall survival at 5 or 10 years were observed between genders. Notably, AR-positive luminal A male breast cancer had improved overall survival over female breast cancer at 5 (P = 0.01, HR = 0.39, 95% CI = 0.26-0.87) but not 10 years (P = 0.29, HR = 0.75, 95% CI = 0.46-1.26) and both 5 (P = 0.04, HR = 0.37, 95% CI = 0.07-0.97) and 10 years (P = 0.04, HR = 0.43, 95% CI = 0.12-0.97) in the unselected group. Hierarchical clustering revealed common clusters between genders including total PR-PRA-PRB and ERβ1/2 clusters. A striking feature was the occurrence of ERα on distinct clusters between genders. In female breast cancer, ERα clustered with PR and its isoforms; in male breast cancer, ERα clustered with ERβ isoforms and AR. Our data supports the hypothesis that breast cancer is biologically different in males and females suggesting implications for clinical management. With the incidence of male breast cancer increasing this provides impetus for further study.

Increased eukaryotic translation initiation factor 4E (eIF4E) expression occurs in many cancers, and makes fundamental contributions to carcinogenesis by stimulating the expression of cancer-related genes at post-transcriptional levels. This key role is highlighted by the facts that eIF4E levels can predict prognosis, and that eIF4E is an established therapeutic target. However, eIF4E activity is a complex function of expression levels and phosphorylation statuses of eIF4E and eIF4E-binding proteins (4E-BPs). Our hypothesis was that the combined analyses of these pathway components would allow insights into eIF4E activity and its influence on cancer. We have determined expression levels of eIF4E, 4E-BP1, 4E-BP2 and phosphorylated 4E-BP1 within 424 breast tumours, and have carried out analyses to combine these and relate the product to patient survival, in order to estimate eIF4E activity. We show that this analysis gives greater prognostic insights than that of eIF4E alone. We show that eIF4E and 4E-BP expression are positively associated, and that 4E-BP2 has a stronger influence on cancer behaviour than 4E-BP1. Finally, we examine eIF4E, estimated eIF4E activity, and phosphorylated 4E-BP1 as potential predictive biomarkers for eIF4E-targeted therapies, and show that each determines selection of different patient groups. We conclude that eIF4E's influence on cancer survival is modulated substantially by 4E-BPs, and that combined pathway analyses can estimate functional eIF4E.

Oestrogen receptors (ERs) are critical regulators of the behaviour of many cancers. Despite this, the roles and regulation of one of the two known ERs – ERβ– are poorly understood. This is partly because analyses have been confused by discrepancies between ERβ expression at mRNA and proteins levels, and because ERβ is expressed as several functionally distinct isoforms. We investigated human ERβ 5′ untranslated regions (UTRs) and their influences on ERβ expression and function. We demonstrate that two alternative ERβ 5′UTRs have potent and differential influences on expression acting at the level of translation. We show that their influences are modulated by cellular context and in carcinogenesis, and demonstrate the contributions of both upstream open reading frames and RNA secondary structure. These regulatory mechanisms offer explanations for the non-concordance of ERβ mRNA and protein. Importantly, we also demonstrate that 5′UTRs allow the first reported mechanisms for differential regulation of the expression of the ERβ isoforms 1, 2 and 5, and thereby have critical influences on ERβ function.

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Incidence rates of male breast cancer (MBC) are rising. MBC etiology is poorly understood with most of our current knowledge regarding its biology, natural history and treatment extrapolated from our knowledge of female breast cancer (FBC). Retrospective studies on MBC have suffered from small numbers of cases available from any one centre thus a significant problem in studying this disease is accruing sufficiently large numbers to allow comparative analysis of possible prognostic markers. Using a co-ordinated multi-centre approach, the aim of this study was to conduct the first large scale study to address the relevance of the expression of recognised biomarkers in FBC in the same disease in males. Five hundred and twenty three cases were obtained retrospectively and assimilated into TMAs, including 260 MBCs and 263 cases of stage-matched FBCs. MBC comprised 21 grade 1, 121 grade 2, 68 grade 3, 50 unknown, mean age 67 (range 39-90) with 167 ductal, 4 lobular, 10 papillary, 10, mucinous, 4 DCIS, 1 mixed and 64 unknown. FBC comprised 29 grade 1, 140 grade 2, 94 grade, mean age 58 (range 27-92) with 220 ductal, 23 lobular, 14 mixed and 6 unknown. Four µm TMA sections were analysed using the following biomarkers: hormone receptors (ERα, ERβ1, ERβ2, ERβ5, total PR, PRA, PRB, AR), apoptosis markers (p53, bcl2), basal (CK5/6, CK14) and luminal epithelial markers (CK18, CK19), E-cadherin and HER2. Biomarkers were scored according to published criteria; for ERβ isoforms both nuclear and cytoplasmic immunoreactivity was determined Statistical analysis was conducted using SPSS. Luminal A (ERα+, and/or PR+, HER2-) was seen in 93% of MBC vs. 84% of FBC, Luminal B (ERα+, and/or PR+, HER2+) or HER2 subgroup (ERα-, PR-, HER2+) was not seen in MBC but found in 6% and 2% of FBC, respectively. Basal-like tumours (ERα-, PR-, HER2-, CK5/6+) were infrequent (MBC 2%, FBC 1%) and in MBC these tumours also expressed ERβ isoforms. No differences were observed in grade, stage or LN status between genders. Univariate analysis showed ERα, ERβ1, ERβ5, PRA, AR, p53 were significantly associated with FBC while cytoplasmic ERβ2, bcl2 and e-cadherin were associated with MBC (all P<0.001). Although membranous HER2 was not seen in MBC, many cases displayed nuclear staining. Biomarker profile with respect to clinical outcome is on-going. This work has shown the luminal A phenotype is common in MBC and that gender-specific biomarkers are expressed. As MBC is becoming more common, this information may be useful in identifying biomarkers which might affect outcome. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 2109.