Retrospective analysis of phosphorylation status of the estrogen receptor in patients with early stage disease

Abstract

21034 Background: The AKT kinase, when phosphorylated to its activated form (pAKT), mediates many proliferative and survival effects of growth factor receptors such as members of the epidermal growth factor receptor (EGFR) family. Overexpression of AKT in breast neoplasms suggests that AKT activation is involved in breast cancer development and/or progression, and contributes to poor outcome. pAKT is frequently upregulated in Her-2/neu positive breast cancers. Activated AKT is able to phosphorylate in vitro serine 167 (pS167) of estrogen receptor alpha (ERa), resulting in ER-mediated transcriptional activity in the absence of estrogen and possibly contributing to tamoxifen resistance in ERa positive breast cancers. In addition, ERa is phosphorylated at S118 (pS118) by MAPK. Thus, it is possible that pAKT- mediated ERa phosphorylation is implicated in tumorigenesis and resistance to treatment with hormonal therapy in patients with breast cancer. Methods: Utilizing commercially available antibodies, IHC analysis of 125 paraffin-embedded, ERa positive, early breast cancer cases, stratified for stage, were tested for pAKT, PTEN, pS167, and pS118. Staining intensity and number of positive cells were assessed by 4 independent graders. Fisher's exact test was used to determine the significance of associations with a two-sided type I error of 5%. Results: Phosphorylation of AKT occurred in approximately one-half of cases. pS167 was phosphorylated in 66% of pAKT positive cases compared to 35% of pAKT negative cases (p<0.0001). When we analyzed nuclear pAKT staining, this association was more pronounced: 76% of pAKT(+) vs. 37% of pAKT(-) (p<0.0001). 69% of pS118(+) cases were pS167(+) compared to only 15% pS167(+) when pS118 was negative (p<0.0001). In approx. 32% of cases pAKT was present in both the cytoplasm and nucleus. 20% of samples had only cytoplasmic pAKT, while in 4% of cases pAKT was nuclear only, supporting the hypothesis that cytoplasmic phosphorylation of AKT may precede nuclear accumulation. Conclusions: Phosphorylation of pS167ERa is significantly associated with AKT activation (especially in the nucleus) in early breast cancer, supporting the hypothesis that these two events may be causally linked and contribute to breast cancer pathogenesis. No significant financial relationships to disclose.