Abstract P5-03-02: Expression of RANK and RANK ligand (RANKL) in breast carcinoma and distinct breast epithelial cells from BRCA1 mutation carriers

Abstract

Abstract Background: Breast tumors in BRCA1 mutation carriers likely arise from luminal progenitor (LP) cells, previously shown to exhibit aberrant growth properties. Oophorectomy, and possibly tamoxifen, reduce breast cancer risk in BRCA1 carriers, potentially via inhibition of paracrine mediated signaling to stem/progenitor cells. RANKL is a major paracrine effector of progesterone's mitogenic action in mammary epithelium via its receptor RANK, and has a role in ovarian hormone-dependent activation of stem cells. Here we assessed RANK and RANKL expression in breast tumors and normal breast epithelial subtypes from women with mutations of BRCA1 (mBRCA1) or BRCA2 (mBRCA2). Methods: RANK and RANKL expression in breast cancer or normal breast tissue samples with mBRCA1, mBRCA2 or wildtype (WT) BRCA1/2 were analyzed in formalin fixed paraffin embedded (FFPE) sections by IHC. kConFab and The Royal Melbourne Hospital Tissue Bank provided the samples used in this analysis; these samples were obtained with relevant IRB approval. RANK expression on normal breast epithelial cells was measured by flow cytometry. Antibodies against human RANK (N-1H8, N-2B10; Amgen) and RANKL (M366; Amgen) were used in both assays. Incidence of IHC staining was scored as a positive IHC signal of any intensity. The overall expression was generated using the H scoring method which is calculated as the staining intensity of the tumor (0-3) multiplied by the percentage of positively staining cells. Results: Breast tumors from women with mBRCA1 had a higher incidence of RANK expression (68/162; 42%) compared with mBRCA2 (17/113; 15%) or WT (34/314; 11%) and higher overall H score (21.3) compared with mBRCA2 (8.0) or WT (3.4); RANKL expression did not vary greatly between groups: mBRCA1 (13/135; 10%), mBRCA2 (5/114; 4%), WT (23/212; 11%). In normal breast tissue, LP (Lin−EpCAM+CD49f+) and basal/stem cells (Lin−EpCAM−CD49fhi) expressed RANK on their surface. Similar expression patterns were seen in these epithelial subtypes from each BRCA1/2 genotype. Stromal cells (Lin−EpCAM−CD49f−) had minimal RANK expression. Conclusions: RANK expression intensity and incidence scores are both enriched approximately 4-fold in breast tumors from BRCA1 carriers compared with other genotypes. Also, RANK is normally expressed in breast LP cells as well as the basal/stem cell containing population. Ongoing studies will assess functional regulation of LP or mammary stem cell activity by RANKL and determine if the RANKL/RANK signaling pathway affects the aberrant growth characteristics of these cells from BRCA1 mutation carriers. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P5-03-02.