Radiotherapy or Surgery of the Axilla After a Positive Sentinel Node in Breast Cancer: 10-Year Results of the Randomized Controlled EORTC 10981-22023 AMAROS Trial.

PURPOSE: The European Organisation for Research and Treatment of Cancer 10981-22023 AMAROS trial evaluated axillary lymph node dissection (ALND) versus axillary radiotherapy (ART) in patients with cT1-2, node-negative breast cancer and a positive sentinel node (SN) biopsy. At 5 years, both modalities showed excellent and comparable axillary control, with significantly less morbidity after ART. We now report the preplanned 10-year analysis of the axillary recurrence rate (ARR), overall survival (OS), and disease-free survival (DFS), and an updated 5-year analysis of morbidity and quality of life. METHODS: In this open-label multicenter phase III noninferiority trial, 4,806 patients underwent SN biopsy; 1,425 were node-positive and randomly assigned to either ALND (n = 744) or ART (n = 681). RESULTS: Per intention-to-treat analysis, 10-year ARR cumulative incidence was 0.93% (95% CI, 0.18 to 1.68; seven events) after ALND and 1.82% (95% CI, 0.74 to 2.94; 11 events) after ART (hazard ratio [HR], 1.71; 95% CI, 0.67 to 4.39). There were no differences in OS (HR, 1.17; 95% CI, 0.89 to 1.52) or DFS (HR, 1.19; 95% CI, 0.97 to 1.46). ALND was associated with a higher lymphedema rate in updated 5-year analyses (24.5% v 11.9%; P < .001). Quality-of-life scales did not differ by treatment through 5 years. Exploratory analysis showed a 10-year cumulative incidence of second primary cancers of 12.1% (95% CI, 9.6 to 14.9) after ART and 8.3% (95% CI, 6.3 to 10.7) after ALND. CONCLUSION: This 10-year analysis confirms a low ARR after both ART and ALND with no difference in OS, DFS, and locoregional control. Considering less arm morbidity, ART is preferred over ALND for patients with SN-positive cT1-2 breast cancer.

Prevention of lymphoedema after axillary clearance by external compression sleeves PLACE randomised trial results. Effects of high BMI.

Around 25% of women undergoing Axillary Clearance (ANC) develop lymphedema (LE). Intervention with a compression garment is recommended to prevent LE but no randomised evidence exists to support this strategy. METHODS: A randomised trial tested standard management versus application of graduated compression garments (20-24 mmHg) to affected arm, for 1 year. Women with node positive breast cancer (n = 1300) undergoing ANC consented to arm volume measurements and those developing a 4-9% relative arm volume increase (RAVI) (subclinical LE) within 9 months post-surgery were randomised. Primary outcome was proportion of patients developing LE (RAVI > 10%) by 24-months in each group. Secondary endpoints included Quality of life in each group. RESULTS: In total 143 patients were randomised (74 no sleeve: 69 compression sleeve) between October 2010 and November 2015. The lymphoedema rate at 24 months in the 'no sleeve' group was at 41%, similar to the 'sleeve' group (30%: p = 0.32). Thirtytwo patients randomised to the 'no sleeve' group had a sleeve applied within 24 months. Body Mass Index (BMI) at randomisation predicted LE at any time point HR 1.04 (CI 1.01-1.08; p = 0.01). Patients with obesity (BMI > 30) had higher rates of LE in both groups (46%) compared to those with BMI < 30 (24%). No difference between patients was found in either group in changes in QoL. Compression sleeves applied after development of LE improved QoL scores (FACT-B p = 0.007:TOI p = 0.042). CONCLUSION: Early intervention with External Compression garments does not prevent clinical LE, particularly in women with a high BMI > 30. The use of prophylactic garments in subclinical LE (RAVI < 9%) is unwarranted.

Margin status and survival outcomes after breast cancer conservation surgery: prospectively registered systematic review and meta-analysis.

OBJECTIVE: To determine if margin involvement is associated with distant recurrence and to determine the required margin to minimise both local recurrence and distant recurrence in early stage invasive breast cancer. DESIGN: Prospectively registered systematic review and meta-analysis of literature. DATA SOURCES: Medline (PubMed), Embase, and Proquest online databases. Unpublished data were sought from study authors. ELIGIBILITY CRITERIA: Eligible studies reported on patients undergoing breast conserving surgery (for stages I-III breast cancer), allowed an estimation of outcomes in relation to margin status, and followed up patients for a minimum of 60 months. Patients with ductal carcinoma in situ only or treated with neoadjuvant chemotherapy or by mastectomy were excluded. Where applicable, margins were categorised as tumour on ink (involved), close margins (no tumour on ink but <2 mm), and negative margins (≥2 mm). RESULTS: 68 studies from 1 January 1980 to 31 December 2021, comprising 112 140 patients with breast cancer, were included. Across all studies, 9.4% (95% confidence interval 6.8% to 12.8%) of patients had involved (tumour on ink) margins and 17.8% (13.0% to 23.9%) had tumour on ink or a close margin. The rate of distant recurrence was 25.4% (14.5% to 40.6%) in patients with tumour on ink, 8.4% (4.4% to 15.5%) in patients with tumour on ink or close, and 7.4% (3.9% to 13.6%) in patients with negative margins. Compared with negative margins, tumour on ink margins were associated with increased distant recurrence (hazard ratio 2.10, 95% confidence interval 1.65 to 2.69, P<0.001) and local recurrence (1.98, 1.66 to 2.36, P<0.001). Close margins were associated with increased distant recurrence (1.38, 1.13 to 1.69, P<0.001) and local recurrence (2.09, 1.39 to 3.13, P<0.001) compared with negative margins, after adjusting for receipt of adjuvant chemotherapy and radiotherapy. In five studies published since 2010, tumour on ink margins were associated with increased distant recurrence (2.41, 1.81 to 3.21, P<0.001) as were tumour on ink and close margins (1.44, 1.22 to 1.71, P<0.001) compared with negative margins. CONCLUSIONS: Involved or close pathological margins after breast conserving surgery for early stage, invasive breast cancer are associated with increased distant recurrence and local recurrence. Surgeons should aim to achieve a minimum clear margin of at least 1 mm. On the basis of current evidence, international guidelines should be revised. SYSTEMATIC REVIEW REGISTRATION: CRD42021232115.

Prognostic and Predictive Value of HER2 Expression in Ductal Carcinoma In Situ: Results from the UK/ANZ DCIS Randomized Trial.

PURPOSE: HER2 is overexpressed more frequently in ductal carcinoma in situ (DCIS) than in invasive breast cancer but its prognostic significance and predictive role for radiotherapy has not been clearly established. We investigated the prognostic and predictive value of HER2 overexpression in DCIS. EXPERIMENTAL DESIGN: HER2 expression was evaluated by IHC using the HercepTest™ in samples from UK/ANZ DCIS trial participants (n = 755) with IHC 3+ expression categorized as HER2 positive for primary analyses. Sensitivity analyses included HER2 categorization as negative (IHC 0,1+), equivocal (IHC 2+), and positive (IHC 3+) and analyses restricted to a nested case-control component where 181 cases (with recurrence) were matched to 362 controls by treatment arm and age. RESULTS: Two-hundred and forty-five (34.4%) of evaluable 713 samples [181 ipsilateral breast events (IBE)] were HER2 positive. HER2 overexpression was associated with significantly increased risk of IBE [HR = 2.29; 95% confidence interval (95% CI), 1.64-3.14; P < 0.0001] and in situ IBE (DCIS-IBE; HR = 2.90; 95% CI, 1.91-4.40; P < 0.0001), but not of invasive IBE (I-IBE; HR = 1.40; 95% CI, 0.81-2.42; P = 0.23; Pheterogeneity = 0.04). Inclusion of HER2 significantly improved [Δχ2 (1d.f.) 12.25; P = 0.0005] a prognostic model of clinicopathological and treatment variables, HER2 being an independent predictor of IBE (multivariate HR = 1.91; 95% CI, 1.33-2.76; P = 0.0004). Radiotherapy benefit in preventing DCIS-IBE was significantly greater (Pheterogeneity = 0.04) in HER2-positive DCIS (HR = 0.16; 95% CI, 0.07-0.41) compared with HER2-negative DCIS (HR = 0.58; 95% CI, 0.28-1.19). CONCLUSIONS: HER2 overexpression is associated with significantly increased risk of in situ recurrence and is also predictive of radiotherapy benefit, with greater reductions in in situ but not invasive recurrences in HER2-positive DCIS.

FAK inhibition alone or in combination with adjuvant therapies reduces cancer stem cell activity.

Cancer stem-like cells (CSC) contribute to therapy resistance and recurrence. Focal adhesion kinase (FAK) has a role in CSC regulation. We determined the effect of FAK inhibition on breast CSC activity alone and in combination with adjuvant therapies. FAK inhibition reduced CSC activity and self-renewal across all molecular subtypes in primary human breast cancer samples. Combined FAK and paclitaxel reduced self-renewal in triple negative cell lines. An invasive breast cancer cohort confirmed high FAK expression correlated with increased risk of recurrence and reduced survival. Co-expression of FAK and CSC markers was associated with the poorest prognosis, identifying a high-risk patient population. Combined FAK and paclitaxel treatment reduced tumour size, Ki67, ex-vivo mammospheres and ALDH+ expression in two triple negative patient derived Xenograft (PDX) models. Combined treatment reduced tumour initiation in a limiting dilution re-implantation PDX model. Combined FAK inhibition with adjuvant therapy has the potential to improve breast cancer survival.

Prognostic Value of ER and PgR Expression and the Impact of Multi-clonal Expression for Recurrence in Ductal Carcinoma in situ: Results from the UK/ANZ DCIS Trial.

PURPOSE: The prognostic value of estrogen receptor (ER)/progesterone receptor (PgR) expression in ductal carcinoma in situ (DCIS) is unclear. We observed multi-clonality when evaluating ER/PgR expression in the UK/ANZ DCIS trial, therefore, we investigated the prognostic role of both uni-clonal and multi-clonal ER/PgR expression in DCIS. EXPERIMENTAL DESIGN: Formalin-fixed paraffin embedded tissues were collected from UK/ANZ DCIS trial participants (n = 755), and ER/PgR expression was evaluated by IHC in 181 cases (with recurrence) matched to 362 controls by treatment arm and age. Assays were scored by the Allred method and by a newly devised clonal method-analyses categorizing multi-clonal DCIS as ER/PgR-positive as per current practice (Standard) and as ER/PgR-negative (clonal) were performed. RESULTS: ER expression was multi-clonal in 11% (39/356) of ER-positive (70.6%, 356/504) patients. Ipsilateral breast event (IBE) risk was similarly higher in ER-multi-clonal and ER-negative DCIS as compared with DCIS with uni-clonal ER expression. ER-negative DCIS (clonal) had a higher risk of in situ IBE [OR 4.99; 95% confidence interval (CI), 2.66-9.36; P < 0.0001], but the risk of invasive IBE was not significantly higher (OR 1.72; 95% CI, 0.84-3.53; P = 0.14), P heterogeneity = 0.03. ER was an independent predictor in multivariate analyses (OR 2.66; 95% CI, 1.53-4.61). PgR status did not add to the prognostic information provided by ER. CONCLUSIONS: ER expression is a strong predictor of ipsilateral recurrence risk in DCIS. ER-positive DCIS with distinct ER-negative clones has a recurrence risk similar to ER-negative DCIS. ER should be routinely assessed in DCIS, and ER scoring should take clonality of expression into account.

Rivaroxaban compared to no treatment in ER-negative stage I-III early breast cancer patients (the TIP Trial): study protocol for a phase II preoperative window-of-opportunity study design randomised controlled trial.

BACKGROUND: Breast cancer patients are at a four-fold increased risk of developing a venous thromboembolism (VTE), a major cause of death in this group. Conversely, coagulation factors promote tumour growth and metastasis. This has been evidenced in preclinical models, with an inhibitory effect of anticoagulants on cancer growth through proliferative, angiogenic, apoptotic, cancer stem cell and metastatic processes. The extrinsic clotting pathway is also more upregulated in patients in the relatively poorer prognosis oestrogen receptor (ER)-negative breast cancer subgroup, with increased tumour stromal expression of the coagulation factors Tissue Factor and thrombin. Rivaroxaban (Xarelto®, Bayer AG, Leverkusen, Germany) is a direct oral anticoagulant (DOAC). It is a Factor Xa inhibitor that is routinely prescribed for the prevention of stroke in non-valvular atrial fibrillation and for both VTE prophylaxis and treatment. This trial will assess the anti-proliferative and other anti-cancer progression mechanisms of Rivaroxaban in ER-negative early breast cancer patients. METHODS: This UK-based preoperative window-of-opportunity phase II randomised control trial will randomise 88 treatment-naïve early breast cancer patients to receive 20 mg OD Rivaroxaban treatment for 11 to 17 days or no treatment. Treatment will be stopped 24 h (range 18-36 h) prior to surgery or repeat core biopsy. All patients will be followed up for 2 weeks following surgery or repeat core biopsy. The primary endpoint is change in tumour Ki67. Secondary outcome measures include tumour markers of apoptosis and angiogenesis, extrinsic clotting pathway activation and systemic markers of metastasis, tumour load and coagulation. DISCUSSION: Laboratory evidence supports an anti-cancer role for anticoagulants; however, this has failed to translate into survival benefit when trialled in patients with metastatic disease or poor prognosis cancers, such as lung cancer. Subgroup analysis supported a potential survival benefit in better prognosis advanced disease patients. This is the first study to investigate the anti-cancer effects of anticoagulants in early breast cancer. TRIAL REGISTRATION: UK National Research Ethics Service (NRES) approval 15/NW/0406, MHRA Clinical Trials Authorisation 48380/0003/001-0001. The sponsor is Manchester University NHS Foundation Trust, and the trial is co-ordinated by Cancer Research UK Liverpool Cancer Trials Unit (LCTU). EudraCT 2014-004909-33 , registered 27 July 2015. ISRCTN14785273 .

Breast cancer stromal clotting activation (Tissue Factor and thrombin): A pre-invasive phenomena that is prognostic in invasion.

BACKGROUND: Tumor stroma, of which fibroblasts are the most abundant cell, resembles a non-healing wound, where a procoagulant environment creates a permissive milieu for cancer growth. We aimed to determine if tumor expression of coagulation factors (procoagulant phenotype), and systemic hypercoagulability, occur at the preinvasive (ductal carcinoma in situ; DCIS) stage and correlate with breast cancer subtype, disease-free survival (DFS), and overall survival (OS). METHODS: In a prospective cohort of early breast cancer (DCIS, n = 76; invasive, n = 248) tumor, normal breast and plasma were examined. Fibroblast and epithelial expression of Tissue Factor (TF), thrombin, PAR1, PAR2, and plasma thrombin-antithrombin (TAT) and D-dimer were correlated with clinicopathological data, and 5-year survival. RESULTS: Fibroblast expression of TF, thrombin, and PAR1 was increased in DCIS and invasive cancer compared to normal breast fibroblasts (P ≤ .003, all). Fibroblast TF, thrombin, PAR1, and PAR2 was increased in cancers with high Ki67, high grade, ER- (vs ER+), and HER2+ (vs HER2-) (all P < .05). On univariate analysis, fibroblast TF expression was inversely associated with DFS (P = .04) and OS (P = .02). D-dimer was higher in node positive (507 (CI: 411-625) ng/mL, n = 68) vs negative patients (428 (CI: 387-472) ng/mL, n = 171, P = .004) and inversely associated with OS (P = .047). On multivariate analysis, plasma TAT was associated with reduced OS (HR 3.26, CI 1.16-3.1, P = .02), with a high plasma TAT (≥3.2 ng/mL) associated with > 3-fold mortality risk compared to low TAT. CONCLUSION: This demonstrates procoagulant phenotypic changes occur in fibroblasts at the preinvasive stage. Fibroblast procoagulant phenotype is associated with aggressive breast cancer subtypes and reduced survival. Coagulation may be a therapeutic target in breast cancer.

Recruitment to the "Breast-Activity and Healthy Eating After Diagnosis" (B-AHEAD) Randomized Controlled Trial.

Excess weight at breast cancer diagnosis and weight gain during treatment are linked to increased breast cancer specific and all-cause mortality. The Breast-Activity and Healthy Eating After Diagnosis (B-AHEAD) trial tested 2 weight loss diet and exercise programmes versus a control receiving standard written advice during adjuvant treatment. This article identifies differences in characteristics between patients recruited from the main trial site to those of the whole population from that site during the recruitment period and identifies barriers to recruitment. A total of 409 patients with operable breast cancer were recruited within 12 weeks of surgery. We compared demographic and treatment factors between women recruited from the main trial coordinating site (n = 300) to the whole breast cancer population in the center (n = 532). Uptake at the coordinating site was 42%, comparable to treatment trials in the unit (47%). Women recruited were younger (55.9 vs 61.2 years, P < .001), more likely to live in least deprived postcode areas (41.7% vs 31.6%, P = .004), and more likely to have screen-detected cancers (55.3% vs 48.7%, P = .026) than the whole breast cancer population. The good uptake highlights the interest in lifestyle change around the time of diagnosis, a challenging time in the patient pathway, and shows that recruitment at this time is feasible. Barriers to uptake among older women and women with a lower socioeconomic status should be understood and overcome in order to improve recruitment to future lifestyle intervention programs.

Tissue Factor promotes breast cancer stem cell activity in vitro.

Cancer stem cells (CSCs) are a subpopulation of cells that can self-renew and initiate tumours. The clotting-initiating protein Tissue Factor (TF) promotes metastasis and may be overexpressed in cancer cells with increased CSC activity. We sought to determine whether TF promotes breast CSC activity in vitro using human breast cancer cell lines. TF expression was compared in anoikis-resistant (CSC-enriched) and unselected cells. In cells sorted into of TF-expressing and TF-negative (FACS), and in cells transfected to knockdown TF (siRNA) and overexpress TF (cDNA), CSC activity was compared by (i) mammosphere forming efficiency (MFE) (ii) holoclone colony formation (Hc) and (iii) ALDH1 activity. TF expression was increased in anoikis-resistant and high ALDH1-activity T47D cells compared to unselected cells. FACS sorted TF-expressing T47Ds and TF-overexpressing MCF7s had increased CSC activity compared to TF-low cells. TF siRNA cells (MDAMB231,T47D) had reduced CSC activity compared to control cells. FVIIa increased MFE and ALDH1 in a dose-dependent manner (MDAMB231, T47D). The effects of FVIIa on MFE were abrogated by TF siRNA (T47D). Breast CSCs (in vitro) demonstrate increased activity when selected for high TF expression, when induced to overexpress TF, and when stimulated (with FVIIa). Targeting the TF pathway in vivo may abrogate CSC activity.

Randomized controlled trial of stereotactic 11-G vacuum-assisted core biopsy for the diagnosis and management of mammographic microcalcification.

OBJECTIVE: To compare the accuracy of 11-G vacuum-assisted biopsy (VAB) with 14-G core needle biopsy (CNB) to diagnose mammographic microcalcification (MM) and effect on surgical outcomes. METHODS: Following ethical approval, VAB and CNB (control) were compared in a randomized prospective study for first-line diagnosis of MM and subsequent surgical outcomes in two breast-screening units. Participants gave written informed consent. Exclusions included comorbidity precluding surgery, prior ipsilateral breast cancer and lesions >40 mm requiring mastectomy as first surgical procedure. The final pathological diagnosis was compared with the initial biopsy result. Quality-of-life (QOL) questionnaires were administered at baseline, 2, 6 and 12 months. 110 participants were required to show a 25% improvement in diagnosis with VAB compared with CNB (90% power). RESULTS: Eligibility was assessed for 787 cases; 129 females recalled from the National Health Service breast screening programme were randomized. Diagnostic accuracy of VAB was 86% and that of CNB was 84%. Using VAB, 2/14 (14.3%) cases upgraded from ductal carcinoma in situ to invasion at surgery and 3/19 (15.8%) using CNB. Following VAB 7/16 (44%) cases required repeat surgery vs 7/24 (29%) after CNB. Both groups recorded significant worsening of functional QOL measures and increased breast pain at follow-up. CONCLUSION: VAB and CNB were equally accurate at diagnosing MM, and no significant differences in surgical outcomes were observed. ADVANCES IN KNOWLEDGE: The first randomized controlled study of VAB for diagnosis of microcalcification using digital mammography showed no difference in diagnostic accuracy of VAB and CNB, or in the proportion of participants needing repeat non-operative biopsy or second therapeutic operation to treat malignancy.

Comparison of multi-frequency bioimpedance with perometry for the early detection and intervention of lymphoedema after axillary node clearance for breast cancer.

The importance of early detection of lymphoedema by arm volume measurements before surgery and repeated measurements after surgery in women undergoing axillary node clearance (ANC) in order to enable early intervention is recognised. A prospective multi-centre study was performed which studied the difference between multi-frequency bioimpedance electrical analysis (BIS) and perometer arm measurement in predicting the development of lymphoedema. Women undergoing ANC underwent pre-operative and regular post-operative measurements of arm volume by both methods. The primary endpoint is the incidence of lymphoedema (≥10 % arm volume increase compared to contralateral arm by perometer) at 2 and 5 years after ANC. The threshold for intervention in lymphoedema was also assessed. Out of 964 patients recruited, 612 had minimum 6 months follow-up data. Using 1-month post-operative measurements as baseline, perometer detected 31 patients with lymphoedema by 6 months (BIS detected 53). By 6 months, 89 % of those with no lymphoedema reported at least one symptom. There was moderate correlation between perometer and BIS at 3 months (r = 0.40) and 6 months (r = 0.60), with a sensitivity of 73 % and specificity of 84 %. Univariate and multivariate analyses revealed a threshold for early intervention of ≥5 to <10 % (p = 0.03). Threshold for early intervention to prevent progression to lymphoedema is ≥5 to <10 % but symptoms alone do not predict lymphoedema. The modest correlation between methods at 6 months indicates arm volume measurements remain gold standard, although longer term follow-up is required.

Is axillary lymph node clearance required in node-positive breast cancer?

Although the majority of patients with breast cancer have clinically negative axillary nodes at preoperative assessment, around 15-20% of these women will have metastatic disease within the lymph nodes at operative sentinel node biopsy, and additional selective treatment to the axilla might be required. Local treatment to the axilla can include axillary node clearance or axillary radiotherapy. The recent results of the American College of Surgeons Oncology Group Z0011 trial suggested that some women would be safe from recurrence without further axillary treatment if they have less than three involved sentinel nodes, with no extracapsular spread. We review the evidence base for management of the axilla after detection of a positive sentinel node, discuss the evidence for why micrometastatic disease requires systemic but not axillary therapy, and present data suggesting that axillary irradiation for macrometastases gives equivalent control to axillary node clearance, but causes less morbidity such as lymphoedema. Ongoing trials will confirm whether any further therapy can be omitted for all patients with low volume, sentinel-node macrometastases.

Focal adhesion kinase and Wnt signaling regulate human ductal carcinoma in situ stem cell activity and response to radiotherapy.

Cancer stem cells (CSCs) can avoid or efficiently repair DNA damage from radio and chemotherapy, which suggests they play a role in disease recurrence. Twenty percentage of patients treated with surgery and radiotherapy for ductal carcinoma in situ (DCIS) of the breast recur and our previous data show that high grade DCIS have increased numbers of CSCs. Here, we investigate the role of focal adhesion kinase (FAK) and Wnt pathways in DCIS stem cells and their capacity to survive irradiation. Using DCIS cell lines and patient samples, we demonstrate that CSC-enriched populations are relatively radioresistant and possess high FAK activity. Immunohistochemical studies of active FAK in DCIS tissue show high expression was associated with a shorter median time to recurrence. Treatment with a FAK inhibitor or FAK siRNA in nonadherent and three-dimensional matrigel culture reduced mammosphere formation, and potentiated the effect of 2 Gy irradiation. Moreover, inhibition of FAK in vitro and in vivo decreased self-renewal capacity, levels of Wnt3a and B-Catenin revealing a novel FAK-Wnt axis regulating DCIS stem cell activity. Overall, these data establish that the FAK-Wnt axis is a promising target to eradicate self-renewal capacity and progression of human breast cancers.

Radiotherapy or surgery of the axilla after a positive sentinel node in breast cancer (EORTC 10981-22023 AMAROS): a randomised, multicentre, open-label, phase 3 non-inferiority trial.

BACKGROUND: If treatment of the axilla is indicated in patients with breast cancer who have a positive sentinel node, axillary lymph node dissection is the present standard. Although axillary lymph node dissection provides excellent regional control, it is associated with harmful side-effects. We aimed to assess whether axillary radiotherapy provides comparable regional control with fewer side-effects. METHODS: Patients with T1-2 primary breast cancer and no palpable lymphadenopathy were enrolled in the randomised, multicentre, open-label, phase 3 non-inferiority EORTC 10981-22023 AMAROS trial. Patients were randomly assigned (1:1) by a computer-generated allocation schedule to receive either axillary lymph node dissection or axillary radiotherapy in case of a positive sentinel node, stratified by institution. The primary endpoint was non-inferiority of 5-year axillary recurrence, considered to be not more than 4% for the axillary radiotherapy group compared with an expected 2% in the axillary lymph node dissection group. Analyses were by intention to treat and per protocol. The AMAROS trial is registered with ClinicalTrials.gov, number NCT00014612. FINDINGS: Between Feb 19, 2001, and April 29, 2010, 4823 patients were enrolled at 34 centres from nine European countries, of whom 4806 were eligible for randomisation. 2402 patients were randomly assigned to receive axillary lymph node dissection and 2404 to receive axillary radiotherapy. Of the 1425 patients with a positive sentinel node, 744 had been randomly assigned to axillary lymph node dissection and 681 to axillary radiotherapy; these patients constituted the intention-to-treat population. Median follow-up was 6·1 years (IQR 4·1-8·0) for the patients with positive sentinel lymph nodes. In the axillary lymph node dissection group, 220 (33%) of 672 patients who underwent axillary lymph node dissection had additional positive nodes. Axillary recurrence occurred in four of 744 patients in the axillary lymph node dissection group and seven of 681 in the axillary radiotherapy group. 5-year axillary recurrence was 0·43% (95% CI 0·00-0·92) after axillary lymph node dissection versus 1·19% (0·31-2·08) after axillary radiotherapy. The planned non-inferiority test was underpowered because of the low number of events. The one-sided 95% CI for the underpowered non-inferiority test on the hazard ratio was 0·00-5·27, with a non-inferiority margin of 2. Lymphoedema in the ipsilateral arm was noted significantly more often after axillary lymph node dissection than after axillary radiotherapy at 1 year, 3 years, and 5 years. INTERPRETATION: Axillary lymph node dissection and axillary radiotherapy after a positive sentinel node provide excellent and comparable axillary control for patients with T1-2 primary breast cancer and no palpable lymphadenopathy. Axillary radiotherapy results in significantly less morbidity. FUNDING: EORTC Charitable Trust.

Lapatinib inhibits stem/progenitor proliferation in preclinical in vitro models of ductal carcinoma in situ (DCIS).

Breast-conserving surgery for ductal carcinoma in situ (DCIS) is often combined with irradiation, reducing recurrence rates to 20% within 10 years; however, there is no change in overall survival. Evidence in the invasive breast indicates that breast cancer stem cells (CSCs) are radiotherapy-resistant and are capable of re-initiating a tumor recurrence; hence, targeting CSCs in high risk DCIS patient may improve survival. HER2 is overexpressed in 20% of DCIS and is known to be highly active in breast CSCs; we therefore investigated the effect of Lapatinib on DCIS CSC activity using 2 in vitro culture systems. Two DCIS cell lines DCIS.com (HER2 normal) and SUM225 (HER2 overexpressed) as well as DCIS cells from patient samples (n = 18) were cultured as mammospheres to assess CSC activity and in differentiated 3D-matrigel culture to determine effects within the non-CSCs. Mammosphere formation was reduced regardless of HER2 status, although this was more marked within the HER2-positive samples. When grown as differentiated DCIS acini in 3D-matrigel culture, Lapatinib only reduced acini size in the HER2-positive samples via decreased proliferation. Further investigation revealed lapatinib did not reduce self-renewal activity in the CSC population, but their proliferation was decreased regardless of HER2 status. In conclusion we show Lapatinib can reduce DCIS CSC activity, suggesting that the use of Lapatinib in high-risk DCIS patients has the potential to reduce recurrence and the progression of DCIS to invasive disease.

Critical research gaps and translational priorities for the successful prevention and treatment of breast cancer.

INTRODUCTION: Breast cancer remains a significant scientific, clinical and societal challenge. This gap analysis has reviewed and critically assessed enduring issues and new challenges emerging from recent research, and proposes strategies for translating solutions into practice. METHODS: More than 100 internationally recognised specialist breast cancer scientists, clinicians and healthcare professionals collaborated to address nine thematic areas: genetics, epigenetics and epidemiology; molecular pathology and cell biology; hormonal influences and endocrine therapy; imaging, detection and screening; current/novel therapies and biomarkers; drug resistance; metastasis, angiogenesis, circulating tumour cells, cancer 'stem' cells; risk and prevention; living with and managing breast cancer and its treatment. The groups developed summary papers through an iterative process which, following further appraisal from experts and patients, were melded into this summary account. RESULTS: The 10 major gaps identified were: (1) understanding the functions and contextual interactions of genetic and epigenetic changes in normal breast development and during malignant transformation; (2) how to implement sustainable lifestyle changes (diet, exercise and weight) and chemopreventive strategies; (3) the need for tailored screening approaches including clinically actionable tests; (4) enhancing knowledge of molecular drivers behind breast cancer subtypes, progression and metastasis; (5) understanding the molecular mechanisms of tumour heterogeneity, dormancy, de novo or acquired resistance and how to target key nodes in these dynamic processes; (6) developing validated markers for chemosensitivity and radiosensitivity; (7) understanding the optimal duration, sequencing and rational combinations of treatment for improved personalised therapy; (8) validating multimodality imaging biomarkers for minimally invasive diagnosis and monitoring of responses in primary and metastatic disease; (9) developing interventions and support to improve the survivorship experience; (10) a continuing need for clinical material for translational research derived from normal breast, blood, primary, relapsed, metastatic and drug-resistant cancers with expert bioinformatics support to maximise its utility. The proposed infrastructural enablers include enhanced resources to support clinically relevant in vitro and in vivo tumour models; improved access to appropriate, fully annotated clinical samples; extended biomarker discovery, validation and standardisation; and facilitated cross-discipline working. CONCLUSIONS: With resources to conduct further high-quality targeted research focusing on the gaps identified, increased knowledge translating into improved clinical care should be achievable within five years.

Targeting IL-8 signalling to inhibit breast cancer stem cell activity.

Although survival from breast cancer has improved significantly over the past 20 years, disease recurrence remains a significant clinical problem. The concept of stem-like cells in cancer has been gaining currency over the last decade or so, since evidence for stem cell activity in human leukaemia and solid tumours, including breast cancer, was first published. Evidence indicates that this sub-population of cells, known as cancer stem-like cells (CSCs), is responsible for driving tumour formation and disease progression. In breast cancer, there is good evidence that CSCs are intrinsically resistant to conventional chemo-, radio- and endocrine therapies. By evading the effects of these treatments, CSCs are held culpable for disease recurrence. Hence, in order to improve treatment there is a need to develop CSC-targeted therapies. Interleukin-8 (IL-8), an inflammatory cytokine, is upregulated in breast cancer and associated with poor prognostic factors. Accumulating evidence demonstrates that IL-8, through its receptors CXCR1/2, is an important regulator of breast CSC activity. Inhibiting CXCR1/2 signalling has proved efficacious in pre-clinical models of breast cancer providing a good rationale for targeting CXCR1/2 clinically. Here, we discuss the role of IL-8 in breast CSC regulation and development of novel therapies to target CXCR1/2 signalling in breast cancer.

Combined inhibition of ErbB1/2 and Notch receptors effectively targets breast ductal carcinoma in situ (DCIS) stem/progenitor cell activity regardless of ErbB2 status.

Pathways involved in DCIS stem and progenitor signalling are poorly understood yet are critical to understand DCIS biology and to develop new therapies. Notch and ErbB1/2 receptor signalling cross talk has been demonstrated in invasive breast cancer, but their role in DCIS stem and progenitor cells has not been investigated. We have utilised 2 DCIS cell lines, MCF10DCIS.com (ErbB2-normal) and SUM225 (ErbB2-overexpressing) and 7 human primary DCIS samples were cultured in 3D matrigel and as mammospheres in the presence, absence or combination of the Notch inhibitor, DAPT, and ErbB1/2 inhibitors, lapatinib or gefitinib. Western blotting was applied to assess downstream signalling. In this study we demonstrate that DAPT reduced acini size and mammosphere formation in MCF10DCIS.com whereas there was no effect in SUM225. Lapatinb reduced acini size and mammosphere formation in SUM225, whereas mammosphere formation and Notch1 activity were increased in MCF10DCIS.com. Combined DAPT/lapatinib treatment was more effective at reducing acini size in both DCIS cell lines. Mammosphere formation in cell lines and human primary DCIS was reduced further by DAPT/lapatinib or DAPT/gefitinib regardless of ErbB2 receptor status. Our pre-clinical human models of DCIS demonstrate that Notch and ErbB1/2 both play a role in DCIS acini growth and stem cell activity. We report for the first time that cross talk between the two pathways in DCIS occurs regardless of ErbB2 receptor status and inhibition of Notch and ErbB1/2 was more efficacious than either alone. These data provide further understanding of DCIS biology and suggest treatment strategies combining Notch and ErbB1/2 inhibitors should be investigated regardless of ErbB2 receptor status.