C. H. M. van Deurzen

Substantial breast cancer risk reduction and potential survival benefit after bilateral mastectomy when compared with surveillance in healthy BRCA1 and BRCA2 mutation carriers: a prospective analysis

BACKGROUND To prospectively assess the efficacy of bilateral risk-reducing mastectomy (BRRM) when compared with surveillance on breast cancer (BC) risk and mortality in healthy BRCA1 and BRCA2 mutation carriers. PATIENTS AND METHODS Five hundred and seventy healthy female mutation carriers (405 BRCA1, 165 BRCA2) were selected from the institutional Family Cancer Clinic database. Eventually, 156 BRCA1 and 56 BRCA2 mutation carriers underwent BRRM. The effect of BRRM versus surveillance was estimated using Cox models. RESULTS During 2037 person-years of observation (PYO), 57 BC cases occurred in the surveillance group versus zero cases during 1379 PYO in the BRRM group (incidence rates, 28 and 0 per 1000 PYO, respectively). In the surveillance group, four women died of BC, while one woman in the BRRM group presented with metastatic BC 3.5 years after BRRM (no primary BC), and died afterward, yielding a HR of 0.29 (95% CI 0.02-2.61) for BC-specific mortality. CONCLUSIONS In healthy BRCA1/2 mutation carriers, BRRM when compared with surveillance reduces BC risk substantially, while longer follow-up is warranted to confirm survival benefits.

High protein expression of EZH2 is related to unfavorable outcome to tamoxifen in metastatic breast cancer.

BACKGROUND Metastatic breast cancer (MBC) is a highly heterogeneous disease with great differences in outcome to both chemo- and endocrine therapy. Better insight into the mechanisms underlying resistance is essential to better predict outcome to therapy and to obtain a more tailored treatment approach. We have previously described that increased mRNA expression levels of Enhancer of Zeste homolog (EZH2) are associated with worse outcome to tamoxifen therapy in MBC. Here, we explored whether this is also the case for EZH2 protein expression. PATIENTS AND METHODS A tissue microarray (TMA) was created using formalin-fixed, paraffin-embedded estrogen receptor (ER)-positive primary breast tumor tissues of 250 MBC patients treated with first-line tamoxifen. Quantity and intensity of EZH2 expression were determined by immunohistochemistry (IHC) and both were used to generate and group scores according to a previously described method for scoring EZH2. RESULTS In total, 116 tumors (46%) were considered to be EZH2 positive. The presence of EZH2 protein expression was significantly associated with progression-free survival (PFS) in both univariate [hazard ratio (HR) 1.51, 95% confidence interval (CI) 1.17-1.97, P = 0.002] and multivariate analysis including traditional factors associated with tamoxifen outcome (HR 1.41, 95% CI 1.06-1.88, P = 0.017). Considering quantity irrespective of intensity, tumors with >50% EZH2-positive cells had the worst PFS (HR 2.15, 95% CI 1.42-3.27, P < 0.001), whereas intensity alone did not show a significant association with PFS. Application of other methods of scoring EZH2 positivity resulted in a similar significant association between the amount of EZH2 positive cells and PFS. CONCLUSION In addition to EZH2 mRNA levels, these results suggest that protein expression of EZH2 can be used as a marker to predict outcome to tamoxifen therapy. This provides new rationale to explore EZH2 inhibition in the clinical setting and increases the possibilities for a more personalized treatment approach in MBC patients.

Characterization of male breast cancer: results of the EORTC 10085/TBCRC/BIG/NABCG International Male Breast Cancer Program

Background Male breast cancer (BC) is rare, managed by extrapolation from female BC. The International Male BC Program aims to better characterize and manage this disease. We report the results of part I, a retrospective joint analysis of cases diagnosed during a 20-year period. Methods Patients with follow-up and tumor samples, treated between 1990 and 2010, in 93 centers/9 countries. Samples were centrally analyzed in three laboratories (the United Kingdom, the Netherlands and the United States). Results Of 1822 patients enrolled, 1483 were analyzed; 63.5% were diagnosed between 2001 and 2010, 57 (5.1%) had metastatic disease (M1). Median age at diagnosis: 68.4 years. Of 1054 M0 cases, 56.2% were node-negative (N0) and 48.5% had T1 tumors; 4% had breast conserving surgery (BCS), 18% sentinel lymph-node biopsy; half received adjuvant radiotherapy; 29.8% (neo)adjuvant chemotherapy and 76.8% adjuvant endocrine therapy (ET), mostly tamoxifen (88.4%). Per central pathology, for M0 tumors: 84.8% ductal invasive carcinomas, 51.5% grade 2; 99.3% estrogen receptor (ER)-positive; 81.9% progesterone receptor (PR)-positive; 96.9% androgen receptor (AR)-positive [ER, PR or AR Allred score ≥3]; 61.1% Ki67 expression low (<14% positive cells); using immunohistochemistry (IHC) surrogates, 41.9% were Luminal-A-like, 48.6% Luminal-B-like/HER-2-negative, 8.7% HER-2-positive, 0.3% triple negative. Median follow-up: 8.2 years (0.0-23.8) for all, 7.2 years (0.0-23.2), for M0, 2.6 years (0.0-12.7) for M1 patients. A significant improvement over time was observed in age-corrected BC mortality. BC-specific-mortality was higher for men younger than 50 years. Better overall (OS) and recurrence-free survival (RFS) were observed for highly ER+ (P = 0.001), highly PR+ (P = 0.002), highly AR+ disease (P = 0.019). There was no association between OS/RFS and HER-2 status, Ki67, IHC subtypes nor grade. Conclusions Male BC is usually ER, PR and AR-positive, Luminal B-like/HER2-negative. Of note, 56% patients had T1 tumors but only 4% had BCS. ER was highly positive in >90% of cases but only 77% received adjuvant ET. ER, PR and AR were associated with OS and RFS, whereas grade, Ki67 and IHC surrogates were not. Significant improvement in survival over time was observed.

Ductal carcinoma in situ diagnosed by breast needle biopsy: Predictors of invasion in the excision specimen

BACKGROUND A substantial proportion of women with a pre-operative diagnosis of pure ductal carcinoma in situ (DCIS) has a final diagnosis of invasive breast cancer (IBC) after surgical excision and, consequently, a potential indication for lymph node staging. The aim of our study was to identify novel predictors of invasion in patients with a needle-biopsy diagnosis of DCIS that would help us to select patients that may benefit from a sentinel node biopsy (SNB). PATIENTS AND METHODS We included 153 patients with a needle-biopsy diagnosis of DCIS between 2000 and 2014, which was followed by surgical excision. Several pre-operative clinical, radiological and pathological features were assessed and correlated with the presence of invasion in the excision specimen. Features that were significantly associated with upstaging in the univariable analysis were combined to calculate upstaging risks. RESULTS Overall, 22% (34/155) of the patients were upstaged to IBC. The following risk factors were significantly associated with upstaging: palpability, age ≤40 years, mammographic mass lesion, moderate to severe periductal inflammation and periductal loss of decorin expression. The upstaging-risk correlated with the number of risk factors present: e.g. 9% for patients without risk factors, 29% for patients with 1 risk factor, 37% for patients with 2 risk factors and 54% for patients with ≥3 risk factors. CONCLUSION The identified risk factors may be helpful to predict the upstaging-risk for patients with a needle-biopsy diagnosis of pure DCIS, which facilitates the performance of a selective SNB for high-risk patients and avoid this procedure in low-risk patients.

Micrometastases and isolated tumor cells: relevant and robust or rubbish? (MIRROR): preliminary results of the MIRROR study from the Dutch breast cancer trialists' group (BOOG).

CTRC-AACR San Antonio Breast Cancer Symposium: 2008 Abstracts Abstract #23 Background The sentinel lymph node procedure (SNP) has largely replaced axillary lymph node dissection (ALND) in patients with early breast cancer (BC). Intensified examination of the sentinel node (SN) results in the detection of isolated tumor cells (ITCs) and micrometastases, whereas its relevance is still debated. This retrospective cohort study is the first study on this topic which includes only patients who underwent a SNP, with central review of all SN slides, and with separate analyses for the impact of administration of adjuvant systemic therapy. Methods Patients operated for BC in all Dutch hospitals in the years 1998-2005, having favourable tumor characteristics (> 35 year; tumor size 1-3 cm and differentiation grade I-II OR tumor size ≤ 1 cm irrespective of grade), having undergone a SNP with pN0(i-), pN0(i+), or pN1mi as final N-stage were selected. Patients were classified in cohort I in case of pN0(i-) and no adjuvant systemic therapy (AST), in cohort II in case of pN0(i+)/pN1mi and no AST, and in cohort III in case of pN0(i+)/pN1mi with AST. SNs were centrally reviewed and restaged according to 6th TNM classification. The 5-year disease-free survival (DFS) was analysed for the 3 different cohorts. Results So far, data are available for 1,744 of the 3,240 selected patients (cohort I n=935; cohort II n=340, cohort III n=469) with a median follow-up of 5.5 years. At diagnosis median age was 58, 56 and 56 years (p < 0.01) and median tumor size was 1.2, 1.4 and 1.5 cm (p < 0.0001). Differentiation grade and hormone receptor status were equally distributed among the three cohorts. Adjuvant systemic therapy in cohort III consisted of chemotherapy (10%), hormonal therapy (63%), or both (27%). The 5-yr DFS is 84% in cohort I, 73% in cohort II, and 86% in cohort III (p = 0.003 cohort I vs II; p < 0.0001 cohort II vs III). Correction for baseline characteristics did not lead to a relevant change in the results. Conclusion BC patients with ITCs or micrometastases as final N-stage after SNP have a significantly worse 5-yr DFS as compared to patients having no ITCs or micrometastases, independent of primary tumor characteristics. Adjuvant systemic therapy significantly improves the 5-yr DFS in the group of patients with ITCs or micrometastases in the lymph node(s). At the meeting, updated analyses will be available for the entire MIRROR study population. Support: The Netherlands organization for health research and development (ZonMw) and all Dutch CCCs. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 23.

Using second harmonic generation to predict patient outcome in solid tumors

BackgroundOver-treatment of estrogen receptor positive (ER+), lymph node-negative (LNN) breast cancer patients with chemotherapy is a pressing clinical problem that can be addressed by improving techniques to predict tumor metastatic potential. Here we demonstrate that analysis of second harmonic generation (SHG) emission direction in primary tumor biopsies can provide prognostic information about the metastatic outcome of ER+, LNN breast cancer, as well as stage 1 colorectal adenocarcinoma.MethodsSHG is an optical signal produced by fibrillar collagen. The ratio of the forward-to-backward emitted SHG signals (F/B) is sensitive to changes in structure of individual collagen fibers. F/B from excised primary tumor tissue was measured in a retrospective study of LNN breast cancer patients who had received no adjuvant systemic therapy and related to metastasis-free survival (MFS) and overall survival (OS) rates. In addition, F/B was studied for its association with the length of progression-free survival (PFS) in a subgroup of ER+ patients who received tamoxifen as first-line treatment for recurrent disease, and for its relation with OS in stage I colorectal and stage 1 lung adenocarcinoma patients.ResultsIn 125 ER+, but not in 96 ER-negative (ER-), LNN breast cancer patients an increased F/B was significantly associated with a favorable MFS and OS (log rank trend for MFS: p = 0.004 and for OS: p = 0.03). On the other hand, an increased F/B was associated with shorter PFS in 60 ER+ recurrent breast cancer patients treated with tamoxifen (log rank trend p = 0.02). In stage I colorectal adenocarcinoma, an increased F/B was significantly related to poor OS (log rank trend p = 0.03), however this relationship was not statistically significant in stage I lung adenocarcinoma.ConclusionWithin ER+, LNN breast cancer specimens the F/B can stratify patients based upon their potential for tumor aggressiveness. This offers a “matrix-focused” method to predict metastatic outcome that is complementary to genomic “cell-focused” methods. In combination, this and other methods may contribute to improved metastatic prediction, and hence may help to reduce patient over-treatment.

Mutational signatures impact the breast cancer transcriptome and distinguish mitotic from immune response pathways

Background: Bladder cancer affects >70,000 patients annually in the United States. Despite its high incidence, therapeutic options are limited in early or late stage. We wanted to identify key metabolic pathways that were altered in bladder cancer development and progression. Experimental Design: We performed global metabolomics profiling of benign urothelium, high-grade non-muscle invasive bladder cancer and advanced, muscle-invasive bladder cancer using GC-MS and LC-MS platforms. This analysis was coupled with publicly available data on transcriptomics of key enzymes, to determine pathways that may be suitable for future therapeutics development. Results: Categorical pathways globally dysregulated in cancer relative to benign urothelium included glucose, TCA cycle, lipid, amino acid and nucleotide pathways. Bladder cancers demonstrated Warburg metabolism, with elevated glucose utilization to drive glycolysis and sorbitol pathway intermediates. Elevated late TCA cycle intermediates, coupled with higher levels of amino acids and dipeptides, suggest the possibility of anaplerotic activity in bladder cancer as a mechanism to sustain energy production. Medium and long chain fatty acids were produced at the expense of dicarboxylic fatty acids. Muscle-invasive bladder cancers showed enhanced use of COX and LOX metabolomics pathways and a possible role for inflammation in regulating NAD+ synthesis in muscle-invasive bladder cancer. Transcriptomic profiling validated that the majority of metabolomics pathway alterations corresponded to gene expression changes of enzymes responsible for metabolite production. Conclusions: This study identifies multiple parallel metabolomics changes unique to non-muscle invasive and muscle-invasive bladder cancer that can be used to justify testing novel therapeutics targeting metabolic pathways in bladder cancer. Citation Format: Divya Sahu, Yair Lotan, Bryan Wittmann, Bruce Neri, Donna Hansel. Metabolomics analysis reveals distinct profiles of non-muscle invasive and muscle-invasive bladder cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2.

Performance of BRCA1/2 mutation prediction models in male breast cancer patients

To establish whether existing mutation prediction models can identify which male breast cancer (MBC) patients should be offered BRCA1 and BRCA2 diagnostic DNA screening, we compared the performance of BOADICEA (Breast and Ovarian Analysis of Disease Incidence and Carrier Estimation Algorithm), BRCAPRO (BRCA probability) and the Myriad prevalence table (“Myriad”). These models were evaluated using the family data of 307 Dutch MBC probands tested for BRCA1/2, 58 (19%) of whom were carriers. We compared the numbers of observed vs predicted carriers and assessed the Area Under the Receiver Operating Characteristic (ROC) Curve (AUC) for each model. BOADICEA predicted the total number of BRCA1/2 mutation carriers quite accurately (observed/predicted ratio: 0.94). When a cut‐off of 10% and 20% prior probability was used, BRCAPRO showed a non‐significant better performance (observed/predicted ratio BOADICEA: 0.81, 95% confidence interval [CI]: [0.60‐1.09] and 0.79, 95% CI: [0.57‐1.09], vs. BRCAPRO: 1.02, 95% CI: [0.75‐1.38] and 0.94, 95% CI: [0.68‐1.31], respectively). Myriad underestimated the number of carriers in up to 69% of the cases. BRCAPRO showed a non‐significant, higher AUC than BOADICEA (0.798 vs 0.776). Myriad showed a significantly lower AUC (0.671). BRCAPRO and BOADICEA can efficiently identify MBC patients as BRCA1/2 mutation carriers. Besides their general applicability, these tools will be of particular value in countries with limited healthcare resources.

Isolated Tumor Cells in Axillary Lymph Nodes of Breast Cancer Patients: Differential Prognostic Impact of Single Tumor Cell(s) Versus Tumor Cell Clusters, and Microanatomic Location. New Results from the Dutch MIRROR Study.

Background The MIRROR study is the largest breast cancer cohort study on pN0(i+) and pN1mi in the sentinel node era, with central pathology review plus restaging according to the 6 th AJCC classification, and separate analyses on the impact of systemic adjuvant therapy. In patients not receiving systemic therapy, pN0(i+) as final N-status was shown to be an equally independent prognosticator for disease-free survival as pN1mi compared with pN0 (SABCS 2008, #23, oral). However, pN0(i+) is a heterogeneous group, consisting of single tumor cell(s) and clusters of cells, and consisting of locations in either sinus or parenchyma. We determined the differential impact of size and microanatomic location of nodal involvement on disease-free survival. Methods Women operated for breast cancer in all Dutch hospitals in the years 1998-2005, with favorable characteristics (tumor size 1-3 cm and differentiation grade I-II OR tumor size Results The 5-year disease-free survival of patients with lymph nodes containing single tumor cell(s) (n=93) was 78.6%, as compared to a 5-year disease-free survival of 77.1% of patients with small clusters of cells (n=404). After adjustment for age, log tumor size, grade, axillary treatment, and hormone receptor status, the adjusted hazard ratio for events was 1.10 (95% CI 0.67-1.78, NS) for the presence of tumor cell clusters as compared to the presence of single tumor cell(s). When including log metastasis size within the pN0(i+) group as a continuous variable in the Cox regression model, increasing metastatic size was borderline significantly associated with reduced 5-years disease-free survival with an adjusted hazard ratio of 1.27 (95% CI 1.00 – 1.61, p=0.05). The adjusted hazard ratio for events was 0.93 (95% CI 0.57-1.51, NS) for the parenchymal (n=112) versus sinusoidal microanatomic location (n=395). Conclusion Single tumor cell(s) bear similar prognostic information as small tumor cell clusters, even though results do suggest that within the pN0(i+) group increasing size of nodal involvement is associated with reduced survival. Microanatomic location does not seem to have prognostic relevance. Support: The Netherlands organization for health research and development (ZonMw) and the Dutch Breast Cancer Trialists9 Group (BOOG) Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 306.

Progression of ductal carcinoma in situ to invasive breast cancer: comparative genomic sequencing

Several models have been described as potential mechanisms for the progression of ductal carcinoma in situ (DCIS) to invasive breast cancer (IBC). The aim of our study was to increase our understanding of DCIS progression by using massive parallel sequencing of synchronous DCIS and IBC. We included patients with synchronous DCIS and IBC (n = 4). Initially, IBC and normal tissue were subjected to whole exome sequencing. Subsequently, targeted sequencing was performed to validate those tumor-specific variants identified by whole exome sequencing. Finally, we analyzed whether those specific variants of the invasive component were also present in the DCIS component. There was a high genomic concordance between synchronous DCIS and IBC (52 out of 92 mutations were present in both components). However, the remaining mutations (40 out of 92) were restricted to the invasive component. The proportion of tumor cells with these mutations was higher in the invasive component compared to the DCIS component in a subset of patients. Our findings support the theory that the progression from DCIS to IBC could be driven by the selection of subclones with specific genetic aberrations. This knowledge improves our understanding of DCIS progression, which may lead to the identification of potential markers of progression and novel therapeutic targets in order to develop a more personalized treatment of patients with DCIS.