The overexpression or amplification of the human epidermal growth factor receptor 2 gene (HER2/neu) is associated with high risk of brain metastasis (BM). The identification of patients at highest immediate risk of BM could optimize screening and facilitate interventional trials. We performed gene expression analysis using complementary deoxyribonucleic acid-mediated annealing, selection, extension and ligation and real-time quantitative reverse transcription PCR (qRT-PCR) in primary tumor samples from two independent cohorts of advanced HER2 positive breast cancer patients. Additionally, we analyzed predictive relevance of clinicopathological factors in this series. Study group included discovery Cohort A (84 patients) and validation Cohort B (75 patients). The only independent variables associated with the development of early BM in both cohorts were the visceral location of first distant relapse [Cohort A: hazard ratio (HR) 7.4, 95 % CI 2.4–22.3; p < 0.001; Cohort B: HR 6.1, 95 % CI 1.5–25.6; p = 0.01] and the lack of trastuzumab administration in the metastatic setting (Cohort A: HR 5.0, 95 % CI 1.4–10.0; p = 0.009; Cohort B: HR 10.0, 95 % CI 2.0–100.0; p = 0.008). A profile including 13 genes was associated with early (≤36 months) symptomatic BM in the discovery cohort. This was refined by qRT-PCR to a 3-gene classifier (RAD51, HDGF, TPR) highly predictive of early BM (HR 5.3, 95 % CI 1.6–16.7; p = 0.005; multivariate analysis). However, predictive value of the classifier was not confirmed in the independent validation Cohort B. The presence of visceral metastases and the lack of trastuzumab administration in the metastatic setting apparently increase the likelihood of early BM in advanced HER2-positive breast cancer.

Background:To investigate whether copy number gain of MET or hepatocyte growth factor (HGF) affect trastuzumab sensitivity in HER2-positive metastatic breast cancer (MBC).Methods:We analysed 130 HER2-positive MBC treated with trastuzumab-based therapy. MET and HGF gene copy numbers (GCN) were assessed by fluorescence in situ hybridisation (FISH) in primary breast cancer samples. Receiver operating characteristic analysis was applied to find the best cutoff point for both MET and HGF GCN.Results:MET FISH-positive cases (N=36, mean ⩾3.72) had a significantly higher trastuzumab failure rate (44.4% vs 16.0% P=0.001) and a significantly shorter time to progression (5.7 vs 9.9 months; HR 1.74; P=0.006) than MET FISH-negative cases (N=94, mean <3.72). Hepatocyte growth factor GCN was evaluated in 84 cases (64.6%). Receiver operating characteristic analysis identified 33 HGF FISH-positive patients (mean HGF GCN ⩾3.01). HGF FISH-positive status was significantly associated with higher risk of failure (30.3% vs 7.8% P=0.007) as compared with HGF FISH-negative cases (N=51, mean <3.01). MET and HGF FISH-positive status was highly correlated (P<0.001) and combination of both biomarkers did not increase predictive value of either considered separately.Conclusion:High GCNs of MET and HGF associate with an increased risk of trastuzumab-based therapy failure in HER2-positive MBC.

IntroductionWe investigated the status of estrogen receptor alpha (ERα), progesterone receptor (PR), and epidermal growth factor receptor 2 (HER2) in primary tumor and in the corresponding brain metastases in a consecutive series of breast cancer patients. Additionally, we studied factors potentially influencing conversion and evaluated its association with survival.MethodsThe study group included 120 breast cancer patients. ERα, PR, and HER2 status in primary tumors and in matched brain metastases was determined centrally by immunohistochemistry and/or fluorescence in situ hybridization.ResultsUsing the Allred score of ≥ 3 as a threshold, conversion of ERα and PR in brain metastases occurred in 29% of cases for both receptors, mostly from positive to negative. Conversion of HER2 occurred in 14% of patients and was more balanced either way. Time to brain relapse and the use of chemotherapy or trastuzumab did not influence conversion, whereas endocrine therapy induced conversion of ERα (P = 0.021) and PR (P = 0.001), mainly towards their loss. Receptor conversion had no significant impact on survival.ConclusionsReceptor conversion, particularly loss of hormone receptors, is a common event in brain metastases from breast cancer, and endocrine therapy may increase its incidence. Receptor conversion does not significantly affect survival.

BackgroundA better understanding of immune response in breast cancer brain metastases (BCBM) may prompt new preventive and therapeutic strategies.MethodsImmunohistochemical expression of stromal tumor-infiltrating lymphocytes (TILs: CD4, CD8, CTLA4), macrophage/microglial cells (CD68), programmed cell death protein 1 receptor (PD-1), programmed cell death protein 1 receptor ligand (PD-L)1, PD-L2 and glial fibrillary acid protein was assessed in 84 BCBM and their microenvironment.ResultsMedian survival after BCBM excision was 18.3 months (range 0–99). Median number of CD4+, CD8+ TILs and CD68+ was 49, 69 and 76 per 1 mm2, respectively. PD-L1 and PD-L2 expression in BCBM was present in 53 % and 36 % of cases, and was not related to BCBM phenotype. PD-1 expression on TILs correlated positively with CD4+ and CD8+ TILs (r = 0.26 and 0.33), and so did CD68+ (r = 0.23 and 0.27, respectively). In the multivariate analysis, survival after BCBM excision positively correlated with PD-1 expression on TILs (hazard ratio (HR) = 0.3, P = 0.003), CD68+ infiltration (HR = 0.2, P < 0.001), brain radiotherapy (HR = 0.1, P < 0.001), endocrine therapy (HR = 0.1, P < 0.001), and negatively with hormone-receptor-negative/human epidermal growth factor receptor 2 (HER2)-positive phenotype of primary tumor (HR = 2.6, P = 0.01), HER2 expression in BCBM (HR = 4.9, P = 0.01).ConclusionsPD-L1 and PD-L2 expression is a common occurrence in BCBM, irrespective of primary tumor and BCBM phenotype. Favorable prognostic impact of PD-1 expression on TILs suggests a beneficial effect of preexisting immunity and implies a potential therapeutic role of immune checkpoint inhibitors in BCBM.

BARD1 and RAD51 are frequently overexpressed in brain metastases from breast cancer and may constitute a mechanism to overcome reactive oxygen species-mediated genotoxic stress in the metastatic brain.