X‐box binding protein 1 (XBP‐1) is stimulated by endoplasmic reticulum stress as part of the unfolded protein response (UPR), which can promote apoptosis or cell survival. Non‐conventional splicing, stimulated during the UPR, converts mRNA for “unspliced” XBP‐1U to “spliced” XBP‐1S mRNA. XBP‐1 mRNA is oestrogen‐responsive, but XBP‐1S confers oestrogen independence and anti‐oestrogen resistance to breast cancer cell lines. We therefore evaluated XBP‐1 mRNA splicing as a factor in response of breast cancer patients to endocrine treatment. XBP‐1 isoforms were measured by quantitative RT‐PCR in 100 primary breast cancer patients treated with adjuvant tamoxifen (including 30 ERα‐negative cases). In ERα‐positive cases, levels of XBP‐1U mRNA correlated with ERα mRNA levels and were lower in grade 3 tumors. Higher levels of XBP‐1U mRNA were significantly associated with breast cancer survival (Log‐rank p = 0.002; Cox hazard ratio (HR) 0.2, p = 0.005), independent of grade, size, nodal status and progesterone receptor status. However, in the full cohort, higher ratios of XBP‐1S/XBP‐1U mRNA (indicating enhanced splicing) were associated with poor survival (Log‐rank p = 0.03; Cox HR 2.3, p = 0.03) and related factors: ERα‐negative status, progesterone receptor negative status, grade 3 tumors and greater proliferation. Significant associations with poor outcome were also seen for XBP‐1 splicing in ERα‐positive cases. Our findings, that XBP‐1 isoforms are differently associated with outcome of endocrine therapy for patients, can be explained by higher levels of dominant‐negative XBP‐1U favouring apoptosis of tumor cells and higher levels of XBP‐1S increasing tumor survival.

S100P, an EF-hand calcium-binding protein, has been reported to be associated with the progression of many types of cancers. Transfection of an expression vector for S100P into a benign, nonmetastatic rat mammary cell line causes a 4- to 6-fold increase in its level in all four transformant cell clones. When the resultant transformant cell lines are introduced in turn into the mammary fat pads of syngeneic Furth-Wistar rats, there is a significant 3-fold increase in local muscle invasion and a significant induction of metastasis in 64% to 75% of tumor-bearing animals. In a group of 303 breast cancer patients followed for up to 20 years, antibodies to S100P immunocytochemically stain 161 primary tumors. Survival of patients with S100P-positive carcinomas is significantly worse by about 7-fold than for those with negatively stained carcinomas. There is also a significant association between the class level of immunocytochemical staining of the carcinoma cells and decreased patient survival. Positive staining for S100P is significantly associated with that for two other metastasis-inducing proteins, S100A4 and osteopontin. Patients with tumors that stained positively for both S100P and S100A4 have a significantly reduced survival of 1.1% over patients with either S100 protein alone. Multivariate regression analysis identifies S100P, S100A4, and osteopontin as the most significant independent indicators of death in this group of patients. These results suggest that stratification of patients into groups according to expression of multiple metastasis-inducing proteins may lead to a more accurate prediction of patient survival. (Cancer Res 2006; 66(2): 1199-207)

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=0.0007) and protein (linear regression analysis r²=0.95, P=0.0002). There is a significant association of staining of carcinomas for AGR2 with oestrogen receptor α (ERα) staining and with low histological grade (both Fisher's Exact test P<0.0001). In the ERα-positive cases, but not the ERα-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=0.006). The significant association of staining for AGR2 with patient death over a 10-year period (log rank test P=0.007, hazard ratio=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.