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Background: Trastuzumab is indicated in neoadjuvant setting in locally advanced HER2 positive breast cancer patients (Gianni L. Lancet 2010). There is no data on the impact of the use of neoadjuvant Trastuzumab (T) compared to adjuvant T on survival. Patients and methods: From May 2004 to October 2007, 341 stage II-III breast cancer patients were included in a phase II randomized trial and received 4 cycles (c) of epirubicin (75 mg/m2 d1)–cyclophosphamide (750 mg/m2 d1) q 3 w followed by 4 (c) of docetaxel (100 mg/m2 d1) q 3 w. Pts with HER2+++ tumor (120 pts) were randomized to receive or not neoadjuvant T combined with docetaxel. From September 2005, all pts with HER2+cancer received adjuvant T for a total of 18 c (106 pts). All pts with hormone receptors positive tumor received hormonal treatment according to menopausal status (Pierga et al BCRT 2010). We report here overall survival (OS) and disease free survival (DFS) data at 5 year and associated prognostic factors. Results: At a median follow up of 49 months, the median DFS was not reached for the whole population and was statistically superior for the HER2 positive cancer patients treated with chemotherapy plus neoadjuvant T compared to the other groups, p = 0.018. The median OS is not reached for the whole population and is statistically higher in HER2 positive tumor group compared to HER2 negative group (p = 0.00077). For 106 HER2 positive breast cancer patients who had received one year of complete trastuzumab treatment, there was no significant difference in OS and DFS between pts who started T in neoadjuvant setting versus in adjuvant setting. DFS and OS were not significantly influenced by pathological Complete Response rate (pCR) (respectively, p = 0.22 and p = 0.56). At multivariate analysis including 6 factors (age, tumor size, clinical lymph node, ER, PgR), factors which influenced OS were tumor size (p = 0.03) and ER expression (p = 0.06) and for DFS, clinical lymph node status (p = 0.049) and PgR expression (p = 0.046). Conclusion: pCR is not a surrogate of survival in the HER2+subgroup. HER2 positive breast cancer pts receiving trastuzumab have a significant higher OS than those with HER2 negative tumors. OS and DFS do not seem to differ between the neoadjuvant T group and the T adjuvant group. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P1-14-03.

Background: However hormone receptor–positive, early-stage breast cancer is a disease with a long natural history and improved survival with 5-year adjuvant endocrine treatments. Yet late recurrence remains an important issue in adjuvant therapy. Predictors of late recurrence are not yet well characterized. Method: A total of 252 breast primary tumors were selected at the Netherlands Cancer Institute from retrospective series of ER+, HER2− breast cancer patients with a follow-up of at least 10 years. Gene expression analysis was performed using Agilent 4×44K microarrays. In order to identify genes associated to late survival differences, we used the survdiff function implemented in the R package survival and we set the parameter rho to −1 to give weight to the later part of the survival curve. The survdiff function was applied to each probe individually for DMFS time considering the probe as a covariate dichotomized into 2 groups (above and below the median expression across all samples). The parameter “strata” was used to stratify the 140 patients based on additional clinico-pathological parameters (Grade, Diameter, Lymph node status and MammaPrint), in order to find genes that add prognostic value to those parameters already known. This approach uses the distant-metastasis free survival (DMFS) time as a continuous variable. Results: After univariate analysis, MammaPrint, diameter, lymph node status and grade were significantly associated to late DMFS differences (Chi-square test p-values equal to 0.016, 0.004, In order to independently validate the prognostic power of these two genes, we tested their performance in the validation set of treated patients (n = 112) and in three publicly available datasets. In all datasets, the CH25H gene confirmed to be significantly associated to metastasis-free survival time in all tested series. Conclusions: These results might indicate that CH25H is an independent marker of late metastatic relapses. CH25H catalyzes the formation of 25-hydroxycholesterol from cholesterol, leading to repress cholesterol biosynthetic enzymes. In the last years, it is emerging that lipid metabolism plays an important role in breast cancer development and progression. Taken together, these findings make the CH25H gene a potential target for late metastasis control in breast cancer. These results warrant further prospective investigation and functional characterization of CH25H in this setting. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P4-09-05.

Background: Predicting a benefit from trastuzumab in patients with HER2+ breast cancer remains an important goal. Possible mechanisms of resistance include altered receptor antibody interaction, Akt and MAPK pathways, and loss of p27. The objective of this study was to determine the correlation between pMAPkinase (pMAPK), pAKT, p27, IGF-IR protein expression and the benefit of trastuzumab for patients randomized to chemotherapy (CT) alone and CT with trastuzumab. Patients and methods: From May 2004 to October 2007, 120 patients with stage II and III HER2+ breast carcinomas were enrolled in a phase II trial of neoadjuvant chemotherapy (CT) with epirubicin-cyclophosphamide (4 courses) followed by docetaxel ± trastuzumab (T) (4 courses). A complete pathological response (pCR) was defined by the absence of residual invasive carcinoma in the breast and axillary lymph nodes. A tissue microarray was constructed from paraffin-embedded tumor samples collected prior to neoadjuvant chemotherapy. Patients' tumours were scored HER2 3+ immunohistochemically (IHC) or 2+ IHC with HER2 amplification by FISH. Immunohistochemical analysis of pMAPK, pAKT, p27 and IGF-IR was performed on tumor tissue microarrays before CT. The H-score (intensity × %) was evaluated. Specimens were classified as exhibiting high or low expression based on a median value as the cut-off point for each marker. A logistic regression model, including the marker and its interaction with treatment, was used to analyse the markers predictive of a treatment effect on the pCR. The independent predictive value was analysed in a multivariate logistic regression adjusting on the lymph node and ER status. Results: 117/120 (97.5%) patients had sufficient tumor for the analysis. The pCR rate was 19% in the CT arm and 25% in the CT+T arm. The median H-score was: pMAPK = 28, pAKT= 25, p27= 50 and IGF-IR = 15. No significant difference was observed in the pCR rate between the two arms according to pAKT, p27, IGF-IR expression. The pCR rate was higher in CT+T compared to CT alone in patients with high pMAPK expression (OR = 4.7 (0.9–24.2); interaction p = 0.03). No difference was observed in the pCR rate in patients with low pMAPK expression (OR = 0.5 (0.1–1.8). Conclusions: In HER2-positive breast cancers, pMAPK expression evaluated by IHC was significantly associated with a pathological response in the arm with neoadjuvant trastuzumab. High pMAPK expression could be a predictive marker of response to trastuzumab in a CT +T regimen. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P3-06-04.

Background: Cox 2 is frequently over expressed in breast cancers. Celecoxib is a COX-2 inhibitor with anti angiogenic and pro-apoptotic activities. There are few data of anti-COX2 treatment in breast cancers. and no data on the impact of neoadjuvant anti COX 2 agent on survival. Patients and methods/: From May 2004 to October 2007, 340 stage II-III breast cancer patients were included in a phase II randomized trial and received 4 cycles (c) of epirubicin (75 mg/m2)–cyclophosphamide (750 mg/m2) q 3 w followed by 4 (c) of docetaxel (100 mg/m2) q 3 w. Pts with HER2 negative tumors (220 pts) were randomized to receive or not neoadjuvant celecoxib (200 mg bid) combined with docetaxel. All pts with hormone receptors positive tumor received hormonal treatment according to menopausal status (Pierga et al BCRT 2010). We report here overall survival (OS) and disease free survival (DFS) data and prognostic factors analyses at 5 years. Results/: At a median follow up of 49 months, the median DFS and OS are not reached for the whole population and none of them is significantly different between pts who received celecoxib or who did not (p = respectively 0.62 and 0.36). Celecoxib had no impact either on clinical and pathological complete response rate (pCR). DFS is significantly higher in patients who achieved pCR as compared to those who did not (p = 0.017; RR = 0.21 [0.051–0.88], whereas OS is borderline significant [p = 0.07; RR = 0.19 (0.026–1.4)]. Patients with triple negative (TN) tumors (78 pts) achieved worst DFS (p = 0.02) and OS (p Conclusion/: Celecoxib had no influence on pCR, DFS or OS. Despite higher pCR rate triple negative breast cancer patients' subgroup remains with the poorest outcome. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P1-14-18.

Introduction: Neoadjuvant chemotherapy increases the likelihood that breast conservation therapy for breast cancer patients will be successful. Breast Cancer Index (BCI), a gene expression assay combining HoxB13/IL17BR ratio and Molecular Grade Index (MGI), is prognostic for the risk of distant recurrence and overall survival in tamoxifen-treated and untreated breast cancer patients. It was previously reported that high risk patients, as determined by BCI, had a 10-fold greater probability of pathologic complete response (pCR) with neoadjuvant chemotherapy than low risk patients. The aim of the current study was to examine the relationship between BCI score and the use of breast conservation surgery (BCS) following treatment with neoadjuvant chemotherapy. Material and Methods: A total of 145 women (tumor size T1, T2 and T3) were treated with neoadjuvant cheomtherapy for stage I-III breast cancer. RNA was extracted from FFPE tumor samples and a real-time RT-PCR assay was completed to generate a BCI score and risk group categorization as previously described (Jerevall et al. Br J Cancer 2011). The relationship between BCS, BCI and clinicopathological factors was examined using univariate and multivariate logistic regression. Results: Of the 145 patients (67% ER+, 54% PR+, 57% >50 y old), 48 (33.1%) underwent BCS. BCI categorized 62 (43%) of patients as low, 50 (34%) as intermediate and 33 (23%) as high risk. The rate of BCS for the three BCI risk categories was 15% (low risk), 48% (intermediate risk) and 45% (high risk). In the low risk group, the rate of BCS was 15% corresponding to a NPV of 85%. This is consistent with previous data from the same cohort, where the NPV of BCI for pCR was 98.4% with only one patient in the low risk group achieving pCR. In univariate analysis, pathological tumor size (pT), ER, PR, grade and BCI were predictors of BCS. A higher BCI score was associated with higher likelihood of BCS (odds ratio of 3.90; CI: 1.45−10.49; p=0.0069). In multivariate analysis, pT and BCI remained significantly associated with BCS, while ER status was not (p=0.23). Results were similar in the subset of patients with T1 and T2 tumors (N=97). In this subset, BCI categorized 42% of patients as low, 37% as intermediate and 21% as high risk and the rate of BCS was 22%, 64% and 60%, respectively. In multivariate analysis of this subset, only BCI was significantly associated with BCS. In all patients, the concordance index based on a model with pT alone was 0.695. When BCI was incorporated into the model with pT, the concordance index increased to 0.801 (p= 0.0002). Conclusion: In this study, we have shown that patients with higher BCI scores were associated with a higher likelihood of receiving BCS after neoadjuvant chemotherapy. Addition of BCI to tumor size increased accuracy in predicting likelihood of BCS. BCI along with standard pathological factors may improve estimation of individual probability of BCS after neoadjuvant chemotherapy. This study gives rise to the hypothesis that patients with low BCI should not be eligible for neoadjuvant chemotherapy since the likelihood of breast conservation is low. Further large confirmatory studies are necessary. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P5-13-08.

Background Previous reports have described the use of microarrays to assess the molecular classification of human breast cancers and defined new subgroups based on gene expression that are relevant to patient management through their ability to predict metastatic relapse and survival relapse. However, different mechanisms may be associated with the development of early and late distant metastases. With the hypothesis that tumors may lead to early or distant metastases based on their intrinsic biological initial features, we aimed at defining molecular profiles for several subgroups of patients based on their outcome over time. Material and methods Breast primary tumors were selected from retrospective series of patients with frozen material available. These series include patients of all ages, LN- and LN+; Estrogen or Progesteron-receptor positive, Her2-negative, no adjuvant treatment, with a follow-up of more than 10 years (y) for the control group or distant metastatic relapse as first event (DM) for the study group (n=144). Patients tumors were classified in 4 groups: no relapse at 10 y (M0), DM before 3 y (M0-3, n=30), DM between 3 and 7 y (M3-7), DM after 7 y (M7+). Samples were collected in 2 different institutions (NKI series for identifying the signature and IGR series for validation). Gene expression analysis of breast tumor samples was performed using custom-made Agilent 44K high-density microarrays and hybridized against the Mammaprint® reference pool (MRP). Tumors were also assessed for their Mammaprint® status, wound-healing signature status and their intrinsic subtypes based on the Blueprint® signature. Moreover, we identified the pathway-level activities of the patient groups using PARADIGM. Results and Discussion For the NKI series, A subset of 144 samples was included based on the selection criteria: 57 M0, 31 M0-3, 25 M3-7, 31 M7+. None of the 3 previously mentioned signatures correctly identified M0 vs. M7+ patients. In order to identify a predictive signature of late relapse (after 7y) we considered M0 and M7+ MammaPrint-Low Risk patients and we split them in a training (n=41) and in a test (n=23) sets. A 73-gene signature was able to classify M7+ patients with 75% of sensitivity and 66% of specificity on the test set. DM after 7yr showed significant activation of pathway related to inflammatory response and angiogenesis. Detailed results and validation results on the independent IGR series will be presented at the meeting. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr S1-6.

Background Post-operative chemotherapy in poor responders after NAC is questionable. We evaluated the use of adjuvant Vinorelbin (NVB) + continuous infusion 5FU or Capecitabine (Cap) in women with poor pathological response to preoperative sequential Epirubicin Cyclophosphamide- Docetaxel.(ECT) Materials and methods: From May 2004 till October 2007, 340 patients (pts) with stage II or III breast adenocarcinoma received neoadjuvant ECT +/− Traztuzumab (HER2+), +/− Celecoxib (HER2 negative) in a randomized phase II trial (Pierga, 2010). 53 patients (15.5 %) had pathological complete response (no infiltrative or in situ carcinoma in the breast and lymph nodes). Eleven pts were not operated. 276 pts had grade 3–4 Chevallier residual disease. These poor responders could be optionally offered post surgery chemotherapy (NVB+5-FU or Cap)+/− Traztuzumab.This regimen started concomitantly with post-operative RT. All positive hormonal receptors received hormonotherapy. We attempted to evaluate DFS according to post operative CT in the 276 pts. To take into account the differences between populations, treated and untreated patients were matched (1:1) according to variables with a significant difference between the two groups, analyses were adjusted with all other prognostic factors. Results: Out of 276 pts with 3–4 chevallier's grade after NAC, 198 pts did not receive any CT (group A), 78 received adjuvant CT (group B): NVB + 5-FU (FUN: N=41) or Cap (N=17) + Traztuzumab (21pts, 11 FUN and 10 CapNVB), 28 pts HER2+ in the group A receive Traztuzumab alone after surgery. The main characteristics of these patients differed for the initial lymph nodes status (N0: 15 (19 %) in group A vs 88 (44 %) in group B, p = With a median follow up of 38.9 months, 63/276 pts (22.8 %) recurred, 44 (22 %) in the group A and 19 (24.3 %) in the group B. DFS for the 276 pts is 95% at 12 months (m) and 86% at 24 m, 95% and 87% in group A and 96% and 83% in group B. (logrank test: p= 0.63). We matched the 2 groups (124 pts; 62/group) according to initial lymph nodes status, local response (Chevallier grade 3 or 4) and pN status after NAC. Characteristics of this matched population were similar except for more ER negative in chemo group, 19 pts (30.6 %) and 9 pts (14.5 %) in non chemo arm. In this population, 27 events occurred, with 13 in the chemo group and 14 in the non chemo'one. Treatment effect was non significant in univariate analysis (p = 0.93). In multivariate analysis adjusted for initial tumor size (TNM), age, molecular status (hormonal receptor and HER2 status), histological grade, initial lymph node status, histological response after NAC, treatment effect was non significant (Hazard Ratio = 1 IC95% [0.38; 2.67], p = 0.99). There was no difference in treatment effect according to molecular status (test for interaction p = 0.30).This multivariate analysis was also completed in the 276 pts with similar results. Conclusion: In poor responders patients after Anthracyclins-Taxanes NAC, adjuvant chemotherapy with Vinorelbine- 5-FU or Capecitabine- did not appear to prolong DFS. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P5-18-13.

Background CTC count before a new line of treatment and CTC count early changes under chemotherapy have been reported as an independent prognostic marker in metastatic breast cancer in a recent pooled analysis in 841 pts (Liu M. ASCO 2011). The aim of this study was to build a prognostic tool including CTC and other parameters to assesse its predictive value for progression-free survival (PFS) and overall survival (OS). Methods: Data from the IC 2006–04 study were used. This prospective multicentre study included 267 metastatic breast cancer patients treated by first line chemotherapy with or without targeted therapy, in whom appropriate pre-treatment prognostic variables (age, performance status, number of metastatic sites, disease-free interval, ER, PR and HER2 status, tumor grade, LDH, serum markers, CTC count by CellSearch technique before treatment and before cycle 2) were available for statistical analysis. We constructed a multivariate Cox regression model for PFS and OS prediction. A stepwise selection process was applied to achieve the most informative and parsimonious models. Performance was measured with the C-index statistic. Internal validation was performed using leave-two-out technique. Results: Four nomograms have been obtained, in two clinical settings: at inclusion (before the start of any treatment) taking into account the initial CTC count, and during treatment (before cycle 2) taking into account CTC changes under treatment. Their accuracy was good for PFS and OS prediction, with C-index ranging from 0.72 to 0.88. Internal validations allow considering a good accuracy of the models in an external population. Conclusion: These clinically relevant nomograms are a simple tool for a personalized prognostic assessment including CTC assessment. Validation on independent series of patients are ongoing. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-07-04.

Background Patients with BRCA1-BRCA2 germline mutation who developed a first breast cancer (BC) have a high risk to develop a second BC, especially if aged less than 40 at first diagnosis. Thus, a positive BRCA test may influence the therapeutic management and orients towards non conservative surgery to reduce new BC occurrence. We set up a fast-track process to get BRCA1/2 and sometimes p53 mutation results within the shortest possible interval, in recently diagnosed BC patients whose probability to be mutation carriers was high (> 20%). Methods: Between 05/02/2009 and 03/01/2011, 61 patients diagnosed with localized BC requesting neo-adjuvant or adjuvant chemotherapy and whose personal or familial history fulfilled BRCA1/2 or P53 genetic testing criteria, were proposed genetic counselling. After a first multisciplinary team meeting (MT), genetic testing was proposed and performed according to usual genetic counselling rules. Psychological support was offered at every step of the process. Patients were informed that their test would be analysed as part of a fast-track process and were aware of the potential consequences of a positive test. Results were communicated and discussed during a second MT, when initial therapeutic plan was eventually changed. Results: All women who were referred to the genetic clinic for fasttrack testing accepted the analysis. The mean age at diagnosis was 38 (range 26–55). The whole process (from 1st genetic consultation till disclosure consultation) was completed in a median interval of 99 days (range 19–245). All pts received appointment within 5 days, when results were available. 50% pts postponed for more than 15 days their disclosure appointment (range 0–113 days). One patient (pt) declined result disclosure. 18 pts (30%) were found to be carrier of a deleterious mutation (13 BRCA1, 4 BRCA2, 1 p53). Variants of unknown significance (UV) in BRCA1 or BRCA2 genes were identified in another 4 patients. In 18 cases, surgery type initially proposed was changed because of the presence of the germline mutation: all pts were proposed ipsilateral mastectomy eventually associated to immediate reconstruction as often possible, as well as controlateral mastectomy. Final surgery turned to bilateral mastectomy in 3 patients with deleterious BRCA mutation and in one suspect UV 14 patients with mutation accepted the change, but four BRCA2 and one BRCA1 mutation carriers refused mastectomy or bilateral mastectomy. Radiotherapy was cancelled in a p53 mutation carrier. Conclusion: This data show that genetic testing might be considered as a part of therapeutic decision. Moreover, 22% (4/18) opted for a simultaneous controlateral prophylactic mastectomy. On the other hand, some women did not change their mind with mutation knowledge (4/18, 22%). Psychological impact of this procedure is currently retrospectively assessed by a self-questionnaire, fulfilled by mail. Analyses will be available for the meeting. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P2-13-06.

Background: A 1228-probeset molecular classifier (EHT Dx14) able to discriminate benign breast lesions from breast adenocarcinoma with a 96% performance was previously identified in a panel of 71 fine-needle aspirate (FNA) samples of nodular breast lesions collected at Institut Gustave Roussy (IGR). The microarray which enabled the signature identification was based on the genomewide RNA analysis using the ExonHit's SpliceArray™ technology. The objective of the current study was to validate the performance of the EHT Dx14 molecular classifier on an independent cohort of FNA samples stored at the Centre de Ressources Biologiques at IGR. Patients and methods: The samples were collected from women undergoing investigations for suspicious nodular breast lesions classified BIRAD ACR 4 or 5 on mammogram and/or ultrasonography. The morphological cytological analysis of FNA samples classified the lesions as either benign, malignant or indeterminate. All samples were associated with a definite clinico-pathological diagnosis of the breast lesions. The first part of the study aims at confirming the sensitivity, specificity and overall performance of EHT Dx14 among lesions clearly classified by morphological cytology. The second part aims at establishing its performance among lesions with indeterminate cytological results. Results: In the first part of the study, the EHT Dx14 transcriptomic signature was applied on a set of 94 FNA consecutive samples categorized as benign (n=47) or adenocarcinoma (n=47) on cytology. The benign/malignant status of the lesions was confirmed by subsequent investigations or follow-up. Results of EHT DX-14 assessment of the samples were respectively 97.9% (95%CI: [88.7%-99.9%]) for sensitivity and 91.5% (95%CI: [79.6%-97.6%]) for specificity. Results on indeterminate FNA samples are being investigated in the second part of the study. Conclusion: This study did confirm the high performance of EHT Dx14 signature in identifying malignant from benign breast lesions in clinically suspect lesions. EHT Dx14 may provide adequate biological diagnosis in the absence of capability of morphological evaluation. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P1-07-10.

Purpose: Circulating tumors cells (CTC) have been recently proposed as a new dynamic blood marker whose positivity at baseline is a prognostic factor and whose changes under treatment are correlated with progression-free survival (PFS) in metastatic breast cancer patients. However, serum markers levels are also used for the same purpose, and no clear comparison as been reported to date. Patients and methods: The IC 2006–04 enrolled prospectively 267 metastatic breast cancer patients treated by first line chemotherapy and confirmed that CTC levels are an independent prognostic factor for PFS and Overall survival (OS). A pre-planned endpoint was to compare prospectively the positivity rates and the value of CTC (CellSearch®), of serum tumor markers (CEA, CA 15–3, CYFRA 21.1), and of serum non-tumor markers (LDH, ALP) at baseline and under treatment for PFS prediction, independently from the other known prognostic factors, using univariate analyses and concordance indexes. Results: Table 1 shows the incidence of each of the 6 blood markers. Assessing all the 6 markers retrieved 90% of patients with at least one elevated marker at baseline. Interestingly, a combination of two markers (CA 15–3 and CYFRA 21.1, often used in lung cancer) retrieved 86% of patients with at least one marker elevated at baseline. All 6 markers were correlated with poor performance status, high number of metastatic sites and with each other. Each marker was associated, when elevated at baseline, with a significantly shorter PFS in univariate anlaysis. Serum marker changes during treatment, assessed either between baseline and week 3 or between baseline and week 6–9, were significantly associated with PFS, as reported for CTC. Concordance indexes comparison showed no clear superiority of any of the serum marker or CTC for PFS prediction. Conclusion: In the largest prospective CTC study in metastatic breast breast cancer, we previously reported that CTC count, but not serum markers, is an independent prognostic factor for PFS and overall survival. However, for the purpose of PFS prediction by measuring blood marker changes during treatment, currently available blood-derived markers (CTC and serum markers) had globally similar performances. Besides CEA and CA 15–3, CYFRA 21.1 is commonly elevated in metastatic breast cancer and has a strong prognostic value. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-07-18.

Background Beyond quantitative CTC analysis for outcome assessment, their qualitative analysis could predict drug response and resistance. Classification of CTC and quantification of treatment targets on CTC can be subjective as they are morphologically heterogeneous. Current HER2 fluorescence assessment methods rely on a visual comparison between CTC images and external controls (cells lines) and are poorly reproducible. We report an automated classification and quantification of HER2 expression on CTC in a large cohort of metastatic breast cancer (MBC) patients (pts). Materials and Methods: CTC enumeration and HER2 assessment was performed with the CellSearch® system in pts with MBC included in the prospective IC 2006–04 study (Pierga, Ann Oncol 2011), before first line chemotherapy. The FITC conjugated antibody CB11 was used to identify HER2. For quantitative assessment of HER2 the breast cancer lines SKBR3 and MCF7 with known HER2 status were spiked in blood of normal donors and processed with the CellSearch system. Digital images of Cytokeratin-PE, DAPI, CD45-APC, and HER2−FITC from these samples were stored. An algorithm, previously developed in prostate cancer (Ligthart, AACR 2011), was applied on these images to automate identification of CTC. The fluorescence of the HER2 FITC channel for each CTC as well as the CD45+, DAPI+ leukocytes contained within each sample were also quantified using this algorithm. HER2 fluorescence levels of CTC were compared to HER2 status of status of pts. Results: Among 267 pts included in this study, 103 had at least 1 CTC/7.5 ml and HER2 fluorescence assessment. In these pts, correlation analysis between automated (aCTC) and manual (mCTC) CTC counts showed an excellent correlation (R2 =0.978). HER2 fluorescence of CTC, varied greatly within and between pts, whereas leukocyte fluorescence was more homogeneous. The vast majority of CTC exhibited less fluorescence than SKBR3 cells, even in HER2+ pts. Primary tumor was HER2+ and HER2− in 36 and 58 pts respectively. Percentage of HER2+ CTC was higher in pts with primary HER2+ status. Although a rare event, the presence within a sample of at least 1 CTC whose HER2 expression is superior to SKBR3 cells was also strongly associated with HER2+ primary tumor (N=15/36 vs. 3/58; p Conclusion: This study is the first to assess HER2 staining intensity in each CTC isolated from a large number of MBC pts. This was made possible by the aCTC count that was highly correlated with the mCTC count in our cohort and permitted HER2 quantification. Our study shows that HER2 staining is highly heterogeneous among CTC within each pt. These findings demonstrate the feasibility of real time quantitative assessment of treatment targets on CTC and opens a path towards personalized treatment. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-07-09.

Background: Women carrying a germline BRCA1/2 mutation have a lifetime risk of developing breast cancer of 56 to 80%. Prophylactic bilateral mastectomy provides a valid option to reduce breast cancer incidence, but considerably affects the quality of life. There is therefore an urgent need to evaluate medical preventive alternative. The major breast cancer prevention trials using tamoxifen showed an approximately 50% incidence reduction in high risk women. Adjuvant trials comparing aromatase inhibitors (AI) to tamoxifen revealed a higher preventive efficacy of AI for contra-lateral cancer with fewer thrombo-embolic side effects. Patients and Methods: The French federation of cancer centres (“FNCLCC”) has designed a randomized phase III trial to determine the efficacy of letrozole to prevent breast cancer in postmenopausal BRCA1/2 carriers. The “LIBER” study is a double-blind, letrozole versus placebo study involving 32 centers among with 20 are actives. The study opened for recruitment in march 2008. Results: Here we present data reflecting the acceptability of this preventive trial. Twenty-one centers replied to an inquiry. 690 women were eligible. Out of 485 women informed by letter, 217 (44%) came to consultation and 62 (13%) entered the study. The main concerns of women while considering to enter the trial were: the potential side effects, the probability to receive the placebo and the lack of support from other practitioners. Out of 78 enrolled women, 48 (62%) were BRCA1 mutation carriers, 30 (38%) were BRCA2 mutation carriers, 73 (93%) were oophorectomized, 38 (49%) had a previous unilateral cancer, 40 (52%) did not have cancer. 30 (38%) women were less than 50 years old, and 48 (62%) were 50 or over 50 years old. Discussion: For post-menopausal women bearing a BRCA1/2 genetic predisposition, prevention of breast cancer risk by letrozole could provide a precious alternative when bilateral mastectomy is not retained. Among women who received oral information, the acceptability of this randomized double-blind letrozole versus placebo trial is 28%. Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P1-07-02.

Background: Circadian clocks involve 15 specific genes, which control cell cycle, apoptosis, DNA repair and metabolism. Circadian disruption is associated with both an increased risk of breast cancer, and poor survival in patients (pts) with metastatic cancer (IARC#98; Innominato et al. Cancer Res 2009). Objective: to provide first evidence of clinical relevance of clock genes expression for the therapeutic sensitivity of primary breast cancer. Method: Affymetrix Hgu133plus2 microarrays data were derived from 189 primary breast cancers. All the pts had non metastatic disease and were registered in multicenter Phase II trial of neoadjuvant chemotherapy with epirubicin-cyclophosphamide (4 courses) then docetaxel ± herceptin (4 courses). In this study, negative hormonal receptor status and limited tumor size predicted for pathological complete response rate (pCR) (Pierga JY, et al Breast Cancer Res Treat. 2010).The expression data of 15 core clock genes, within the whole tumor transcriptome, were related both to tumor mitotic index (MI) and estrogen receptors (ESR1), and to chemotherapy-induced pCR. Statistical validation involved T-test statistics, with corrections for multiple testing. Results: In the tumor biopsy obtained before treatment onset, both Per2 and Cry2 expressions correlated positively with ESR1 (r≥0.43; P Clock gene expression and pCR Conclusion: Low expressions of three core genes in the negative loop of the molecular circadian clock strongly predicted for the induction of a pathologic complete response of primary breast cancer with neoadjuvant chemotherapy. While circadian clock disruption seems to constitute a poor prognostic factor in cancer patients, our data suggest that the downregulation of core circadian genes in tumors significantly enhances susceptibility to chemotherapy. Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P2-09-26.

Resume Objectif Depuis 2009, l’irradiation partielle acceleree du sein peut etre proposee aux Etats-Unis aux patientes atteintes de cancer du sein unifocal a faible risque de recidive locale en dehors de tout essai clinique. En France, cette modalite therapeutique n’est realisee que dans le cadre d’essais cliniques, jusqu’alors soit par curietherapie (Programme d’action concertee oncogeriatrie (Pac Gerico) de la Federation nationale des centres de lutte contre cancer), par Mammosite ® (Lille) ou par une technique peroperatoire (Montpellier). Nous rapportons ici la premiere experience dosimetrique francaise d’irradiation partielle acceleree conformationnelle tridimensionnelle. Patientes et methodes Depuis octobre 2007, 25 patientes atteintes d’un cancer du sein de stade pT1N0 ont ete incluses dans un essai de phase II d’irradiation partielle acceleree conformationnelle tridimensionnelle a l’institut Gustave-Roussy. La dose totale prescrite etait de 40 Gy, a raison de 4 Gy par fraction et deux fractions par jour pendant cinq jours. La technique d’irradiation consistait en deux mini-faisceaux tangentiels de 6 MV et d’un faisceau d’electrons direct de 6–22 MeV, qui contribuait a 20 % de la dose totale. Resultats Les volumes cibles anatomoclinique et previsionnel moyens etaient respectivement de 15,1 cm 3 (5,2–28,7 cm 3 ) et de 117 cm 3 (52–185 cm 3 ). La couverture du volume cible previsionnel etait satisfaisante avec l’isodose 40 Gy enveloppant en moyenne au moins 99 %. La dose totale moyenne delivree au volume cible previsionnel etait de 41,8 Gy (41–42,4 Gy). L’heterogeneite de la dose delivree n’excedait pas 5 %. Les doses moyennes delivrees au poumon homolateral et au cœur etaient respectivement de 1,6 Gy (1,0–2,3 Gy) et 1,2 Gy (1,0–1,6 Gy). Conclusion Cette modalite d’irradiation partielle acceleree conformationnelle tridimensionnelle de 40 Gy en dix fractions et cinq jours permet une excellente couverture du volume cible previsionnel tout en epargnant considerablement l’irradiation des tissus sains pulmonaires et cardiaques.

Background: National mammography screening program covers populations aged 50-74. We wondered whether breast cancer characteristics and treatment delays were influenced by age and screening method in a community-based breast unit. Patients and Methods: We analyzed complete data of prospectively recorded community operable breast cancer cases in a single regional cancer center between Jan 1st, 2005 and Oct 10th, 2007. Pts were eligible if they i. had a final diagnosis of invasive BC ii. were operated as first trt for BC, iii. had no history of other previous malignancy within the past 5 year period. Logistic regression analyses (SAS) were used to determine factors influencing delays between first description of the lesion and first appointment at breast unit. Results: 1044 pts met the inclusion criteria and had full data available. Age did not appear as a predictor of therapeutic delay, with both elderly and non-elderly pts having a median delay of 19 days between first screening/evidence of disease and 1st appointment at the breast unit. 59% of elderly and 65% of non-elderly pts had respectively had their breast cancer diagnosed through mammographic screening. As expected, mammographic screening as mode of discovery of the cancer was highest, among the 688 pts aged 50-74 (73%). Tumors characteristics and surgical treatment did not differ according to age categories elderly/non elderly as described in Table 1. Conservation rates were high in both populations. Conclusion: Despite absence of national mammographic screening program in elderly populations, patterns of screening and care appear efficient in a community-based setting, with absence of therapeutic delays, similar tumor staging and similar accesss to conservative surgery as compared to younger populations. Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-10-13.

Purpose:The likelihood of menses recovery (MR) is largely variable in premenopausal patients (pts) receiving adjuvant chemotherapy for BC. Quantifying this probability for each single patient could impact discussion of chemotherapy side effects and better individualize fertility counseling.We performed a pooled analysis from PACS04 and PACS05 randomized trials aiming to develop a nomogram to estimate the probability of menses recovery at 6 and 18 months (mos) after the end of adjuvant chemotherapy (CT) for premenopausal pts with early BC. Patients and Methods: The analyzed population consisted of 1683 pts who were premenopausal and ≤ 50 (out of 4524 enrolled in both trials). In PACS05 node-negative BC pts were randomized to 4 or 6 cycles of FE100C (standard arm); in PACS04 node-positive pts were randomized to 6 cycles of FE100C or 6 cycles of Epirubicin 75mg/m2 and Docetaxel 75 mg/m2 (ED75). Endocrine therapy (ET) (Tamoxifen) x 5 years was mandatory for ER+ BC. Variables significantly associated with MR in the univariate analysis (P Results: Pts' characteristics were: median age 43 (22-50), median body mass index (BMI) at baseline 22.6 (15.6-54.7), at the end of chemotherapy 22.8 (15.8-58.6). ED75 was administrated to 517 (30.7%), while 802 (47.7%) received 6FE100C, 364 (21.6) 4FE100C. Trastuzumab was given to 122 (7.2%), ET to 1229 (73%) pts. CT-induced amenorrhea was observed in 1407 (83.6%) pts. Factors associated to MR were assessed on 1210 pts (excluding pts who recovered menses during CT or of whom date of recovery was not specified). At a median follow-up of 90 mos, 28.2% (342/1210) of pts had recovered menstrual cycles: 11% (133/1210) at 6 mos and 24.3% (294/1210) at 18 mos. Multivariate analysis showed that younger age, higher BMI at the end of CT, non-alkylating agents and absence of ET were independently associated to MR. Table 1 Multivariate Cox regression analysis of menses recoveryVariablesHR (95%CI)P valueAge1.49 (1.16-1.93)< 0.002Age2*0.99 [0.98-0.99] Nomogram concordance-index was 0.749 and 0.750 for predicting MR at 6 and 18 mos respectively. A better calibration was observed at 18 mos, comparing nomogram predictions with the actual probability of MR in the 1210 women. Conclusion:Our analysis confirmed the possibility of developing a user-friendly nomogram for predicting menses recovery after adjuvant chemotherapy. As next step, we will externally validate our nomogram on CANTO premenopausal population, one of the biggest national cohorts aiming to assess the long-term impact of cancer treatments toxicities (UNICANCER NCT01993498 - http://etudecanto.org/). Citation Format: Pistilli B, Mazouni C, Zingarello A, Faron M, Saghatchian M, Grynberg M, Spielmann M, Kerbrat P, Roché H, Lorgis V, Bachelot T, Campone M, Levy C, Goncalves A, Lesur A, Veyrat C, Vanlemmens L, Lemonnier J, Delaloge S. MAAT: Menses after adjuvant treatment. Prediction of menses recovery after chemotherapy for early breast cancer (BC) by using a nomogram model in UNICANCER PACS04 and PACS05 trials [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr PD7-06.

Resume La radiotherapie locoregionale est un outil therapeutique majeur dans la prise en charge des cancers du sein. Les importantes avancees enregistrees dans les annees 1990 dans ce domaine, conduisant a des indications d'irradiation locoregionale de plus en plus larges, font cependant emerger trois questions : comment comprendre et lever la radioresistance de certaines tumeurs mammaires ? Comment reconnaitre les femmes susceptibles de beneficier reellement de l'irradiation et ainsi envisager une desescalade therapeutique dans les autres categories ? Comment identifier les sujets hypersensibles a l'irradiation, a haut risque de toxicite a long terme, et adapter les techniques ou les indications dans ces populations ? Les progres majeurs de la genetique des tumeurs du sein dans les dernieres annees devraient aider les cliniciens a resoudre ces importantes questions. Dans cet article sont revues les principales bases moleculaires de la radiosensibilite des cancers du sein, qu'elles soient constitutionnelles ( BRCA1 , BRCA2 , ATM …) ou somatiques ( p53 , recepteurs a tyrosine kinase, acteurs du cycle cellulaire, des voies de transduction du signal, de l'apoptose, de la reparation de l'ADN …). Les methodes recentes d'exploration du patrimoine genetique constitutionnel ou tumoral (profils d'expression ADN, ARN ou proteines) sont egalement discutees comme outils majeurs de la cancerologie mammaire des prochaines annees.

Background: High number of new drugs are targeting molecular alterations that occur in a few number of patients. Molecular screening that consists in the assessment of single molecular alteration during the screening phase of the therapeutic trial is not suitable. Indeed, this modality of screening is associated with a high rate of screen failure due to the low incidence of the molecular alteration. In order to optimize molecular screening, we have launched a clinical trial that aims at performing array CGH and hot spot mutations in patients with metastatic breast cancer. Patients and Methods. The trial plans to include 400 patients with metastatic breast cancer and included the first patient in may 2011. This trial is being sponsored by French federation of Cancer centers (UNICANCER) and involves 20 centers. Biopsy is being performed on metastatic site. Both frozen and FFPE samples are obtained. DNA extraction is being performed in the investigation center after control for the % of cancer cells. Array CGH and PIK3CA/AKT mutations (SANGE method) are being performed in four genomic platforms. A pilot study that included 106 patients has shown the feasibility of such technologies in the context of daily practice. Data from array CGH are being sent to a bioinformatician who forward the results on both pre-selected targets, DNA instability and some targets of interest selected based on log2(ratio) and function. Recommendations for trials are then being sent to each investigator. There is no limitation for previous lines and it is recommended to perform biopsy in patients who do not present progressive disease. This trial is being funded by French NCI (750 000 euros) Conclusion: SAFIR01 is a trial that aims at using high throughput technologies in order to drive patients with molecular alterations to specific therapeutic trials. Further amendments are being planned including implementation of high throughput sequencing and performance of functional testing. Further trial will compare this high throughput approach to standard methods for target identification. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr OT1-03-02.

Background: Women with germline BRCA1or BRCA2 (BRCA1/2) mutations are an extreme risk population for developing breast cancer, with a life-time risk of 56–80%. Prophylactic mastectomy provides a valid option to reduce such risk, but it considerably affects the quality of life. Medical prevention by aromatase inhibitor that has been recently shown to have preventive effect may thus be an alternative. LIBER is an ongoing, double-blind, randomized phase III trial to evaluate the efficacy of five-years letrozole versus placebo to decrease breast cancer incidence in post-menopausal BRCA1/2 mutation carriers (NCT00673335). We present data on the uptake of this trial. Methods: To evaluate theoretical feasibility, we compared inclusion criteria of women in the LIBER trial (n=113) to characteristics of women entered in the prospective ongoing national GENEPSO cohort (n=1505). Uptake was evaluated through a survey sent to all active centres, with responses obtained from 17 of the 20 (85%) centres. Results: According to characteristics of the women included in GENEPSO cohort (n=1505) and the survey, approximately one third of BRCA1/2 mutation carriers are eligible for this study. From november 2009 to may 2010, 534 women eligible from chart review have been informed by mail of the trial and were invited to an oral information by participating centres. Of them, 44% of women came to the dedicated medical visit. Uptake of drug prevention trial was 32 % of orally informed women and 15 % of overall eligible women. Main reasons of refusal were: potential side effects, probability to receive the placebo and lack of support from the women's physicians. Prior prophylactic oophorectomy and history ***of previous unilateral breast cancer were more frequent in women enrolled in the ONCO-03/LIBER trial than in the French cohort (93% versus 60% and 50 % versus 39 %, respectively), suggesting a higher motivation for medical prevention in these subgroups of patients. Conclusion: One third of women with a BRCA1/2 mutation are eligible to the ONCO-03/LIBER prevention trial. 32 % of orally informed women and, only 15% of overall eligible women entered the trial. To reach accrual objective (n=308), a greater information of the trial should be offered to women with BRCA1/2 mutation and the trial has been proposed to other countries. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr OT2-04-03.

Background: Personalized medicine is a rising paradigm in cancer care. The identification of pathways involved in carcinogenesis along with the development of targeted therapies has revolutionized cancer treatment. There is increasing availability of technologies that can interrogate the genomic landscape of the tumor; however, it is still uncertain whether such platforms are used in clinical practice. Methods: We conducted a 28-item survey to investigate the current use of tumor molecular sequencing in the management of breast cancer patients. A link to the online survey was communicated via various platforms such as the European Society for Medical Oncology (ESMO) and European School of Oncology (ESO) newsletter, and via a dedicated mailing by the Breast International Group (BIG) and other academic groups. Descriptive statistical analysis and Fisher's exact tests were applied to explore potential association between the demographic characteristics and responses. Results: A total of 211 physicians from 35 countries participated to the study between the 9 March and 3 June 2015, with 92% fully completed questionnaires. The mean age of the participants was 45 years (range 27-77). The majority of responders were medical oncologists (88%), practicing in Europe (69%) and working in academic institutions (66%). 62% (130/211) of participants had never requested tumor molecular sequencing for breast cancer patients. Working in academic institutions and having more time allocated to research were associated with the use of tumor molecular sequencing (p = 0.007 and 0.009, respectively). For the 81 participants that used tumor molecular sequencing in the past (Table 1), there was a significant association between accessibility and frequency of use (p=0.02). 92% (181/211) of participants claimed that they would probably use tumor molecular sequencing more often if it was more accessible. Lack of funding and lack of access to the technology were the main reasons for poor endorsement. 89% of participants believe that tumor molecular sequencing will play a major role in the management of breast cancer patients in the future. Current weak evidence and poor access to matched targeted therapy are the main concerns against a wider use of these platforms in clinical practice. Table 1. Summary of replies from the 81 participants that used tumor molecular sequencing for breast cancer patientsQuestionResponseN (%)In what percentage of your breast cancer patients has tumor molecular sequencing been performed at least once?≤5%55 (68%) >5%26 (32%)How often do the results lead to enrollment in a clinical trial?≤10%53 (65%) >10%28 (35%)How confident are you in interpreting tumor sequencing results?Not at all/A little17 (21%) Somewhat/Highly64 (79%)Do you consider molecular sequencing platforms accessible?Not at all/A little45 (55%) Somewhat/Highly36 (45%) Conclusion: Our survey indicates that molecular sequencing platforms are sometimes used, albeit not widely in guiding management of breast cancer patients. Poor accessibility may contribute to the low frequency of use, but lack of evidence and poor access to matched targeted therapy are also major concerns. Citation Format: Gingras I, Sonnenblick A, Dolci S, de Azambuja E, Paesmans M, Delaloge S, Piccart MJ, Sotiriou C, Michail I, Azim Jr HA. The role of precision medicine in "real-life" management of breast cancer patients: A survey assessing the current use and attitudes towards tumor molecular sequencing in clinical practice. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P6-04-13.

Mucinous (colloid) breast carcinoma accounts for 1-6% of all breast cancer. It comprises pure mucinous tumours and mixed infiltrating ductal carcinomas with a mucinous component. As this latter mixed form has a worse prognosis than pure colloid carcinoma, making this diagnosis on fine needle aspiration cytology (FNAC) might influence the choice of treatment. We report a consecutive series of 22 cases consisting of 17 mixed and five pure mucinous carcinomas diagnosed by cytology and verified on histopathology. Patients underwent FNAC at the one-stop clinic of our institution during a 7-year period of time. Cytological findings were evaluated by a semi-quantitative method and included percentage of smear surface occupied by mucin, shape of cell groupings, size and outline of tumour nuclei as well as presence or absence of nucleolus. Three of five pure mucinous carcinomas displayed at least two of the following features: abundant mucin, small nuclei and/or regular nuclear outlines. Sparse mucin, large nuclei, irregular nuclear outlines or the presence of nucleoli were found in 7 out of 17 mixed mucinous carcinomas but not in pure tumours. Cytopathological identification of patients with pure mucinous carcinomas may be performed in a limited number of cases.

Since 2009, accelerated partial breast irradiation (APBI) in North America has been allowed to be used for selected group of patients outside a clinical trial according to the ASTRO consensus statement. In France, accelerated partial breast irradiation is still considered investigational, several clinical trials have been conducted using either intraoperative (Montpellier) or Mammosite(®) (Lille) or brachytherapy modality (PAC GERICO/FNCLCC). Here, we report the original dosimetric results of this technique.Since October 2007, Institut Gustave-Roussy has initiated a phase II trial using 3D-conformal accelerated partial breast irradiation (40 Gy in 10 fractions BID in 1 week). Twenty-five patients with pT1N0 breast cancer were enrolled and were treated by two minitangent photons beams (6MV) and an "en face" electron beam (6-22 MeV).The mean clinical target volume and planning target volume were respectively 15.1cm(3) (range: 5.2-28.7 cm(3)) and 117 cm(3) (range: 52-185 cm(3)). The planning target volume coverage was adequate with at least a mean of 99% of the volume encompassed by the isodose 40 Gy. The mean dose to the planning target volume was 41.8 Gy (range: 41-42.4 Gy). Dose inhomogeneity did not exceed 5%. Mean doses to the ipsilateral lung and heart were 1.6 Gy (range: 1.0-2.3 Gy) and 1.2 Gy (range: 1.0-1.6 Gy), respectively.The 3D conformal accelerated partial breast irradiation using two minitangent and "en face" electron beams using a total dose of 40 Gy in 10 fractions BID over 5 days achieves appropriate planning target volume coverage and offers significant normal-tissue sparing (heart, lung). Longer follow-up is needed to evaluate the tissue tolerance to this radiation dose.

CA15-3, a peptide derived from MUC-1, an hormonally-regulated protein, is the most widely used serum marker of breast cancer. CA15-3 level increases at the metastatic phase in 50-80% breast cancer patients. Although rise of CA15-3 precede symptoms of metastasis by a mean time of 2-9 months, current international guidelines do not recommend its routine use for screening for metastases because of moderate sensitivity and absence of clinical impact. We conducted a retrospective study among all patients with metastatic breast cancer seen by three senior breast oncologists during a 4-month period. We evaluated correlation of CA15-3 level at the time of metastatic relapse with ER, PgR and Her2 expressions, tumor type, size and nodal status at initial diagnosis, and sites of metastases. CA15-3 was increased in 168/272 patients (62%) at diagnosis of metastases. ER/PgR positivity was strongly correlated with elevated CA15-3 at this time (P0.0001). CA 15-3 was elevated in 69% of the cases of HR+ Her2- primary tumors at time of metastatic relapse. It was elevated in 56% of HR+ Her2+++, 46% of HR- Her2+++ cases and only in 41% of triple-negative cases (P = 0.003). these data confirm that CA 15-3 is very variably elevated at time of metastatic relapse of breast cancer, and this is dependant on HR status.

When a former breast cancer patient asks for pregnancy, several questions have to be considered. The physician has to answer and counsel clearly and honestly his/her patient regarding the possibilities, risks and specificities associated with a pregnancy in this case. The individual risk of relapse should be considered, together with individual predictive factors and theoretical benefits of specific prolonged treatments such as hormone therapy. Both risks associated with the pregnancy itself and with the modification of treatment schemes have to be evaluated. Risks related to Assisted reproductive techniques when indicated, should also be considered and explained. Potential foetal risks should be explained too. The aim of this review is to summarize the current knowledge and then available tools that the physician can use to help the couple making decision regarding subsequent pregnancy.

This study aimed to evaluate the role of specimen radiography in predicting margin status for non-palpable breast malignancies.We retrospectively reviewed the clinical and pathological data together with specimen radiographs of 164 women with ductal carcinoma in situ, who were referred to our centre between January 1997 and December 2000. In all cases microcalcifications were discovered on mammography. Lesions were localized preoperatively using a guide-wire. Specimen radiography findings and clinicopathological data were correlated with pathological findings.Findings comprised 122 pure ductal carcinomas in situ (74%) and 42 mixed carcinomas, both infiltrating and in situ (26%). On the specimen radiographs, the lesions were close (1mm) to one edge of the lumpectomy in 34 (21%) cases. Histologically, there were 103 positive resection margins (1mm, 63%) and only 61 negative margins (or =1mm, 37%). On univariate analysis, factors associated with positive resection margins were found to be distance from microcalcifications to edge of lesion on specimen radiographs, and radiological multifocality. On multivariate analysis (logistic regression), a radiological margin5mm and multifocality were the only risk factors for close histological margins. Radiological margins were not associated with surgical findings.Our results demonstrate that there is a correlation between specimen radiographs and histological results. The clinical relevance of this should be evaluated in a prospective study.

Local-regional radiation therapy is one of the major therapeutic means in the management of breast cancer. Three questions however arise from the important advances achieved in this domain in the past years. The first question concerns the possibilities to identify and overcome the radioresistance of a subset of tumours. The second question is how to recognize women likely to benefit from adjuvant radiation therapy, and therefore to diminish treatment indications in other groups. Finally, the third question is how to identify subjects at high risk for long term injury following breast irradiation, in order to adapt techniques and indications in such populations. The major advances of breast cancer molecular genetics in the past years should provide clinicians with tools to answer these important questions. In this paper, we review the molecular germline (BRCA1, BRCA2, ATM, ...) and somatic (p53, tyrosine kinase receptors, as well as actors of cell cycle, signal transduction, apoptosis, DNA repair ...) main bases of breast cancer radiosensitivity. Recent methods of exploration of the genetic background of both the host and the tumours (gene and protein expression profiles) are also reviewed as major tools of breast cancer management in the next few years.

Tamoxifen estrogenic action in the uterus induces several uterine diseases, benign and/or malignant ones. The risk of endometrial adenocarcinoma is multiplied by two to three in post-menopausal women. It is mainly linked with the doses and the length of the treatment. However, the global benefit of that drug is not questioned anymore. What matters now though is to find the best way to follow patients on tamoxifen. As a matter of fact, there is no such thing as a consensus that would include specific tests, nor a surveillance protocol in women on tamoxifen. Most teams do not propose any special follow-up. Some patients already show uterine anomalies prior to the beginning of tamoxifen treatment. A yearly gynecologic examination, together with a cervico-vaginal smear, is enough when there are no specific endometrial adenocarcinoma risk factors, nor anomalies detected during the pre-therapeutical evaluation, nor clinical symptomatology. In case of risk factors, or cervical stenosis, or again initial abnormalities though, a yearly transvaginal sonography may be proposed. There is no need for other exploratory examinations if the results are satisfying. In case of symptoms, anomalies in the cervico-vaginal smears, intra-uterine liquid retention with a stenosed cervix, or suspicious endometrial thickness, then an endometrial sampling must be carried out. MRI could be of interest in asymptomatic patients with unclear ultrasonography images. Follow-up must be continued after interruption of tamoxifen. It is important to inform patients about the additional risks of developing an endometrial cancer because of tamoxifen, while still being reassuring. Besides, it is absolutely necessary to recommend them to take quickly medical advice in case of gynecologic symptoms.

Background: After the SWOG0500 trial, the CirCe01 trial (NCT01349842) is the second randomized attempt to demonstrate that patients who received a first cycle of chemotherapy and whose CTC count did not drop should be switched off this chemotherapy. The clinical setting used here is a population with high chemoresistance prevalence, i.e. patients starting a third line chemotherapy; the CTC test will be also repeated at the beginning of every new chemotherapy line in patients randomized in the CTC arm (i.e. to evaluate the 3rd, 4th, 5th … lines). In the CTC arm, it has been hypothesized that many patients with chemoresistant tumor will experience repeated early chemotherapy changes (e.g. 3 chemotherapy regimens tested in 9 weeks), giving support to the discontinuation of inefficient, toxic and costly chemotherapies and the start of palliative care. It is believed, for this subgroup of patients, that such de-escalating management will benefit both patients and healthcare systems. For some other patients, it is also expected that they will benefit from the early chemotherapy turn-around and find earlier an efficient therapy among all those tested. Methods: The main inclusion criteria are metastatic breast cancer patients starting a third line of chemotherapy and an elevated CTC count at baseline. 304 metastatic breast cancer patients will be randomized between the standard arm, in which the treatment management is made following the current standard of care, and the CTC-driven arm, in which the “response” after every first cycle of any new chemotherapy line is assessed by CTC count made before the second cycle. Chemotherapies which did not lead to a significant reduction of CTC count are discontinued, and the patient might be offered another chemotherapy regimen (which will be also evaluated by CTC). The medical primary endpoint of the trial is the overall survival, whereas a medico-economic study as a co-primary endpoint. Several secondary endpoints are pre-planned, including progression-free survival, quality of life, anxiety/depression and comparison with serum markers. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr OT3-4-05.

The hormonal status of 95% of the women treated for epithelial ovarian carcinoma is menopausal, either naturally or after treatment. This raises the important question of the hormonal replacement therapy (HRT) among these patients. Several retrospective studies have explored the potential positive or negative influence of HRT on the genesis of ovarian adenocarcinoma. Although somehow contradictory, these studies taken all together fail to show any favouring nor protective role of HRT. In vitro, estrogens have been shown to induce the proliferation of ovarian cancer cell lines. On the opposite, progesterone and antiestrogens have antiproliferative effects. Both types of effects are mediated by intracellular steroid hormone receptors (ER, PgR). Although high dose progesterone derivatives and antiestrogens have been shown to obtain therapeutic responses in patients carrying advanced ovarian carcinoma, response rates were usually less than 20%, and no clinico-biological correlation (with ER, PgR status) could be demonstrated. There is therefore no evidence for a clinical significance of the presence of hormonal receptors in these tumors. A single retrospective study explored the possible influence of HRT on the prognosis of patients treated for ovarian carcinoma, and did not demonstrate any deleterious effect. This review of recent epidemiological, biological and clinical data fails to find any argument against the prescription of HRT in patients treated for ovarian adenocarcinoma, in the absence of other contra-indications.

Background: Breast cancer (BC) during pregnancy (BCP) is a rare situation that requires multi-disciplinary management. The objectives of this study were to assess the tumor characteristics, clinical course and outcome of such patients (pts). Methods: French hospitals were invited to collect retrospective clinical, treatment and follow-up data of BCP managed between 2000 and 2006. Pts with histologically confirmed M0, invasive BC and pregnant at diagnosis were included. Pts whose pregnancy was interrupted were not eligible. Survival times were calculated from the date of diagnosis. Results: 100 BCP pts were identified. Median age was 32 years (24-42). Median gestational age at diagnosis was 25 weeks (3-38). 84% and 13% had palpable breast axillary mass respectively. Clinical stages were 1T0, 21T1, 39T2, 28T3, 6 T4A-C, 2 T4D, 3 TX, 56 N0, 39 N1, 5 NX. Histological analysis identified 85 ductal carcinomas, 4 lobular, 11 others. The histopathological grades was G1 in 4,3%, G2 in 29%, and G3 in 66,7%. Tumor subtype was luminal A in 3%, luminal B in 37% (24HER2-, 13HER2+), luminal undetermined in 6%, triple-negative in 45,9%, Her2 + in 21,3%, and not classified in 2% (HR -, HER 2 unknown). Median time interval between first observation and biopsy was 31 days (0-337), respectively 40 days (0-337) and 15 days (0-172) when the first observation was made by patients or physicians. Median time interval between pathologic diagnosis and treatment was 18 days (0-295). Treatment was initiated after pregnancy for 42 pts, with median time of 18 days after delivery. 97 pts received chemotherapy with a median number of 6 cycles (4-11), 92 with anthracyclin, 44 with taxanes. 53 chemotherapy were administered in neo adjuvant setting among which 25 during pregnancy), and 44 in adjuvant setting (23 during pregnancy). 98 pts underwent surgery (34 during pregnancy), with 57 conservations and 41 mastectomies, 93 pts received radiotherapy and 43 hormone therapy after pregnancy. 10 pts received Trastuzumab. The mean gestational age at delivery was 35 weeks (22-45). All children were alive, with a median weight of 2735 g at birth (550-3740). The 5-year Overall Survival rate is 83% (95%CI 74-89), while Disease Free Survival is 53% (95%CI 43-63). First recurrence site was metastasis in 28, locoregional in 13, controlateral in 9 and other cancers in 2. Conclusion: Biopsy and treatment intervals remain long among this population. In this large series BCP, there is an excess of triple-negative breast cancer. The 5-year OS rate is higher than previously reported but with DFS is lower. BCP remains an aggressive entity despite adapted treatment. Multivariate analysis will be presented. A comparison of this BCP population to matched controls is ongoing. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P6-06-07.

Background: Rare cancers sometimes present clonal evolution. Genomic characterisation of these rare entities could lead to identify new genes involved in the progression of more frequent cancers. In the present study, we have performed whole exome sequencing of three rare and aggressive histological subtypes in order to identify new targets that could subsequently be validated in common breast cancers. Methods: Whole exome sequencing was applied in ten pleiomorphic lobular carcinoma, seven micropapillary BC and eight metaplastic BC. Each cancer sample was matched with normal DNA from the same patient. Overall 50 samples were therefore sequenced. Tumor samples were selected to present >30% cancer cells. Histological diagnosis was confirmed by a BC pathologist. Whole exome sequencing (WES) was performed by BGI (Bejing Genomic Institute, Hong-Kong, China) using the Illumina Hiseq2000 platform (90bp paired-end reads, depth ≥80x). Genes recurrently mutated (with somatic mutations in >2 cases) were selected for validation. The validation by IonTorrent target sequencing was performed on additional formalin-fixed paraffin-embedded (FFPE) tumor samples (>30% of tumor cells). Results: Complete results of both WES and targeted resequencing validation are available for pleiomorphic lobular carcinoma. Nine of the 10 DNA pairs were eligible for WES. Genes already implicated in BC were significantly mutated: PIK3CA (3/9), TP53 (3/9) and CDH1 (3/9). Novel significantly mutated genes were identified: PYGM (2/9), EMR1 (2/9), ALDH1A2 (2/9), and DCLK1 (2/9). The validation using target gene sequencing was performed on the same 9 cases and on 19 additional confirmed pleiomorphic lobular BC cases (n = 28). For 8 out of the 9 cases with WES data, the results were reproducible on FFPE samples. Validation set confirmed recurrent mutations on PYGM genes (25%). PYGM is involved in glycogen metabolism. Further analyses on gene expression array revealed that PYGM is downregulated in most of the common BC samples, as compared to normal tissue, suggesting a role for this gene in the metabolism of common BC. WES data are available for micropapillary and metaplastic BC. HSPA8 (coding for a Heat shock protein) was found mutated in 3/7 of micropapillary samples. Validation on additional FFPE samples is ongoing and will be presented. WES of metaplastic BC revealed a high frequency of TP53 and PIK3CA mutations (50% in both cases). Conclusions: A novel mutated gene was identified in PLBC. PYGM is a gene involved in glycogen metabolism and was found downregulated in most of the common breast cancer. WES of micropapillary cancers suggests mutations on HSPA8, a gene encoding for a chaperon protein. Validation is ongoing. Deeper sequencing is ongoing for metaplastic BC that may help identifying new mutated genes in this multiclonal disease. Results will be presented. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr PD4-4.

Background: The native fluorescence spectra of human cancerous and normal breast tissues under the selected excitation of a 405 nm light should differ according to their different content of NAD(P)H, FAD, collagen or retinol and derived fluorophores. Nodea Medical has developed an in situ probe, Probea®, installed in a 25-Gauge needle, which is able to measure endogenous tissue fluorescence under 405 nm specific wavelength. Tissues are excited by this blue light; their back-production of fluorescence (around 480 nm) is caught and immediately analyzed. The objectives of the present preclinical study are to evaluate the ability of the Probea® probe to distinguish benign versus malignant breast lesions among freshly harvested ex-vivo surgical samples, and to evaluate the fluorescence behaviour of adjacent and remote normal tissues among cancer specimens. Methods: Fresh breast samples coming out of the operating room were harvested and analyzed prior to pathological fixation. Surgical specimens included mastectomies, lumpectomies, and plastic surgeries in different malignant or non-malignant situations. For each of them, the probe was placed within the main target surgical lesion, as well as within surrounding normal epithelial and fat tissues, and remote normal tissues where available (mastectomies or reduction mammoplasties). We aimed at measuring different parameters of the endogenous fluorescence at each measurement point: maximum intensity, peak wavelength, level of signal stability, growth or spontaneous decay in a fixed position. Fluorescence signals harvested were compared to the detailed pathological analyses of each area explored within the surgical samples. Results : 49 fresh breast samples were harvested and analyzed, of which 40 were from patients with cancer and 9 from patients without cancer. The presence of a fluorescence signal within the main lesion was overall strongly associated with a cancer diagnosis (N = 40/49 malignant lesions versus 0/9 benign lesions, p = 0.001). The positive predictive value of the test in this preliminary series is 98%, whereas the negative predictive value is 100% analyzing the maximum fluorescence intensity with a cut-off of 1000. Interestingly, normal adjacent fat tissues surrounding and remote from breast cancer lesions also produced fluorescence which could also be considered as criteria to define malignancy. Complete analyses of multiple parameters are ongoing. Conclusion: In this initial preclinical study, the Probea® probe has shown an elective ability to distinguish benign and malignant lesions. Particularly, its negative predictive value appears very promising. A clinical study to confirm these findings is planned. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P4-03-06.

Background: The IC 2006–04 study included prospectively 267 metastatic breast cancer patients before the start of first line chemotherapy. We previously reported and confirmed the prognostic and predictive of CTC counts before the start of treatment and after 3–4 & 6–9 weeks of treatment (Pierga, JY SABCS 2010, Ann Oncol 2012). We report here the results of the last CTC count performed at tumor progression (i.e. end of first line chemotherapy). Methods: Patients were included in the study from 06/07 to 9/09. CTC were counted by the CellSearch® system at tumor progression (clinical and/or radiological progression). Results: 211 patients experienced a tumor progression at time of final data analysis and the planned CTC count has been performed in 61 patients. High CTC count (≥5 CTC/7.5ml) at progression was observed in 22 patients (36%) and was associated with elevated Cyfra 21–1 and ALP and was not associated with elevated CEA, CA 15–3, LDH, the elapsed progression-free survival, the onset of new metastasis. No CTC was detected at tumor progression in the 5 patients with HER2+ disease, in line with our previous results. The prognostic markers for overall survival at tumor progression were high CTC count (median overall survival 8 vs 19 months, p = .0006) and elevated Cyfra 21–1 (p = .001). Other serum markers (CEA, CA15-3, ALP, LDH) and tumor subtypes had no prognostic value at univariate analysis. At multivariate analysis, the two prognostic factors for overall survival were high CTC count (RR = 2.3 p = 0.03) and elevated Cyfra 21–1 (RR = 3.6 p = 0.02). Conclusion: In the prospective IC 2006–04 study, among the different markers tested at tumor progression during the first line chemotherapy for metastatic breast cancer, high CTC count and elevated Cyfra 21–1 levels were the only two independent prognostic markers for overall survival. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P2-01-13.

Background: Circulating tumor cells (CTCs) isolated by CellSearch® from peripheral blood of metastatic breast patients have been shown as strong independent prognostic factor for progression-free and overall survival. With this technique, the REMAGUS02 prospective multicentric study was the first to report that CTC detection (≥1CTC/7.5ml) before and/or after neoadjuvant chemotherapy was independently associated with distant metastasis-free survival (DMFS, 18 months follow-up, Pierga CCR 2008) and with overall survival (OS, 36 months follow-up, Bidard Ann Oncol 2010). Patients and Methods: In 115 non-metastatic patients diagnosed with large operable or locally advanced breast cancer, we prospectively detected CTC using the CellSearch system before and after neoadjuvant chemotherapy in the REMAGUS02 trial (Pierga, BCRT 2010). For this report, survival analyses were performed at a median follow-up of 70 months. Results: After a median follow-up of 70 months, 20 distant metastatic relapses and 14 deaths have been observed among the 115 patients included. CTC detection before chemotherapy (in 23% of patients) is an independent prognostic factor for both DMFS [p = 0.03, relative risk (RR)=3.2] and OS [p = 0.05, RR = 3.7] in multivariate analyses, together with triple negative tumor status. At long-term, CTC detection after chemotherapy (17%) had no clear prognostic significance (p = 0.15 and 0.22, respectively). Complete pathological response as well as tumour size and all other variables tested did not appear as predictive in this model. Conclusions: Baseline CTC detection is a new independent prognostic factor in the neoadjuvant setting, and is, in this trial, superior to pathological tumor response to predict the survival of non-metastatic breast cancer patients. We confirm therefore our previous results reported with shorter follow-up. Supported by PHRC AOM/2OO2/02117, Pfizer inc., Roche, sanofi-aventis. ISRCTN10059974 Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P2-01-04.

During SABCS 2009, Western France Observatory of Cancer has presented “ALOPERS” results from more than one hundred patients with persisting alopecia or suboptimal hair regrowth after chemotherapy for early breast cancer. Docetaxel 75–100 mg/msq was concerned for majority of patients. 43% of patients experienced alopecia more than 24 months after their last chemotherapy infusion. Optimal information of patients about alopecia and persisting alopecia appears to be mandatory before treatment: 47% of patients undergo a psychological shock during hair loss. Morevover, BCIRG 001 study (TAC versus FAC) led to the conclusion that docetaxel 75 mg/msq is responsible for persisting alopecia for 3% of patients. So, by extrapolating, in France, each year, docetaxel could induce a persisting alopecia in 300 patients. To confirm this figure, “ERALOP” study is in progress to evaluate precisely the incidence of persisting alopecia after docetaxel in early breast cancer patients and then theses data will be communicated to ANSM. It must be noticed that 8% of patients refused to be treated because of the risk of persistant alopecia (Trueb et al, 2010). That's why it is a worldwide public health problem, with personal and societal implications. As alopecia is universal after FEC100 regimen, the cooling scalp has been supposed to be very effective during docetaxel 1 hour infusion and to allow a better hair regrowth but with a high risk in term of tolerance. Since 2012, a prospective clinical trial “ALOPREV” has begun in Western France to evaluate tolerance of cooling scalp, after 3–4 courses of FEC100 regimen and during 3–4 courses of docetaxel 100 mg/msq, for patients treated for early breast cancer. So, ALOPREV is designed as a safety trial. First, cooling scalp major toxicities (headache, neck pain, discomfort associated with cold, sinusitis) are evaluated during each docetaxel course by the patient. Then, auto-questionnary about hair loss and regrowth, nails disorders and others toxicities due to docetaxel and questionnary about quality of life (EORTC QLQ-C30 and BR-23 scales) is completed by the patients before, during and after treatment. Patients will be followed-up 12 months and final toxicities will be reported by physicians. According to our daily practice, it will demonstrate that 60% of patients could tolerate two consecutive cooling scalps during docetaxel infusion, and with statistical considerations (alpha-beta risks and 10% of ineligibility). So 160 patients must be included during inclusion period (18 months). 10 public or private care centers have been opened (45 investigators). This trial implicates particularly nurses teams and all patients will sign an informed consent before FEC100 courses. ALOPREV started two months ago and 25 patients are already enrolled. In conclusion, cooling scalp is known to be safe, manageable and partially effective before hair loss. ALOPREV will evaluate safety and efficacy to avoid persisting alopecia after hair loss. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr OT3-3-04.