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. 2008 Feb;52(3):355-69.
doi: 10.1111/j.1365-2559.2007.02955.x.

Expression of small breast epithelial mucin (SBEM) protein in tissue microarrays (TMAs) of primary invasive breast cancers

G P Skliris  1 , F Hubé, I Gheorghiu, M M Mutawe, C Penner, P H Watson, L C Murphy, E Leygue, Y Myal
Affiliations

Expression of small breast epithelial mucin (SBEM) protein in tissue microarrays (TMAs) of primary invasive breast cancers

G P Skliris et al. Histopathology. 2008 Feb.

Abstract

Aims: Small breast epithelial mucin (SBEM) is a recently described gene product that shows promise as a new breast biomarker. The aim was to investigate for the first time SBEM protein expression in a large cohort (n = 300) of invasive breast cancers, its relationship to established clinical variables and its association with clinical outcome.

Methods and results: Immunohistochemical analysis was performed on tissue microarrays consisting of 149 oestrogen receptor (ER) alpha- and 151 ERalpha+ breast cancers. Overall, 18% of tumours were SBEM+ (n = 53/300). However, SBEM protein was more frequently observed in ER- (22%) than in ER+ cancers (13%; P = 0.049). A significant association with psoriasin/S100A7 expression (P < or = 0.0001) was observed in the entire cohort. SBEM was also positively associated with HER-2 (P = 0.046) in ER- cancers, and increased levels of SBEM were strongly associated with higher tumour grade (P = 0.0015). Furthermore, SBEM expression showed a trend towards an association with reduced overall survival and relapse-free survival in the ER+ cohort (P = 0.063 and P = 0.072, respectively).

Conclusions: Our results suggest that SBEM may identify a unique subset of breast cancers with poor prognosis and may have future implications for therapeutic management of this disease.

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Figures

Figure 1
Figure 1
SBEM RNA and protein expression in a small panel of 10 human breast cancers. RNA and protein were extracted from breast cancers (n = 10) as described in Materials and Methods. Numbers (1–10) represent individual tumours. A, RT-PCR analysis. ‘N’ is a negative RT-PCR, i.e. reverse transcription performed minus MMLV enzyme, and ‘B’ is a PCR negative control. Lane M, PhiX174 RF DNA/Hae III DNA ladder. B, Northern blot analysis. Breast cancers were analysed as described in Materials and methods. Fifteen micrograms of total RNA from tumours 1–7, three from tumour no. 8, 10 μg from no. 9 and 6 μg from no. 10 were separated on agarose gels and hybridized using SBEM32P-labelled probes. Loading controls are provided with 18s and 28s RNA bands on the bottom panel M, ladder. C, Western blot analysis. Total proteins (25 μg) were analysed by sodium dodecyl sulphate–polyacrylamide gel electrophoresis using the SBEM H39C51 monoclonal antibody. M = ladder.
Figure 3
Figure 3
SBEM expression, determined by immunohistochemistry in ER+ and ER− tissue microarrays. A, ER+ cancer stained with the SBEM monoclonal antibody (H218C31) showing no expression (H-score of 0). B, ER+ tumour showing low expression (H-score of 30). C, ER+ breast cancer showing medium expression (H-score of 60). D, ER+ tumour showing high expression (H-score of 225). E, ER− breast cancer (ER and PR levels of 2.3 and 8.9 fmol/mg protein, respectively) immunoreactive for the H218C31 SBEM-specific antibody showing high expression. F, Serial section of the ER− breast cancer, where the primary antibody was incubated with ×20-fold excess SBEM peptide and abolished the specific stain.
Figure 2
Figure 2
Immunohistochemical analysis of SBEM expression in the small panel of 10 human breast cancers. Immunohistochemistry was performed as described in Materials and Methods. Patient/sample identification is indicated on top left of each picture, and the corresponding H-score for IHC is presented on the bottom right corner. H-scores were derived from semiquantitative assessment of both staining intensity (scale 0–3) and the percentage of positive cells (0–100%) and, when multiplied, generate a score ranging from 0 to 300. Bar = 50 μm.
Figure 4
Figure 4
Kaplan–Meier graphs for overall survival and relapse free survival-time to progression for SBEM expression in the entire breast cancer cohort (A,B), ER α− (C,D) and ERα+ cancers (E,F). Symbols on the graph lines represent censored data, P-values are given for log rank tests. A (top left), n = 300; low SBEM events = 80, high SBEM events = 23; B (top right), n = 300; low SBEM events = 95, high SBEM events = 26; C (left), n = 149; low SBEM events = 57, high SBEM events = 16; D (right), n = 149; low SBEM events = 55, high SBEM events = 17; E (bottom left), n = 151; low SBEM events = 23, high SBEM events = 7; F (bottom right), n = 151; low SBEM events = 40, high SBEM events = 9.
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