αE-catenin is a candidate tumor suppressor for the development of E-cadherin-expressing lobular-type breast cancer.

Although mutational inactivation of E-cadherin (CDH1) is the main driver of invasive lobular breast cancer (ILC), approximately 10-15% of all ILCs retain membrane-localized E-cadherin despite the presence of an apparent non-cohesive and invasive lobular growth pattern. Given that ILC is dependent on constitutive actomyosin contraction for tumor development and progression, we used a combination of cell systems and in vivo experiments to investigate the consequences of α-catenin (CTNNA1) loss in the regulation of anchorage independence of non-invasive breast carcinoma. We found that inactivating somatic CTNNA1 mutations in human breast cancer correlated with lobular and mixed ducto-lobular phenotypes. Further, inducible loss of α-catenin in mouse and human E-cadherin-expressing breast cancer cells led to atypical localization of E-cadherin, a rounded cell morphology, and anoikis resistance. Pharmacological inhibition experiments subsequently revealed that, similar to E-cadherin-mutant ILC, anoikis resistance induced by α-catenin loss was dependent on Rho/Rock-dependent actomyosin contractility. Finally, using a transplantation-based conditional mouse model, we demonstrate that inducible inactivation of α-catenin instigates acquisition of lobular features and invasive behavior. We therefore suggest that α-catenin represents a bona fide tumor suppressor for the development of lobular-type breast cancer and as such provides an alternative event to E-cadherin inactivation, adherens junction (AJ) dysfunction, and subsequent constitutive actomyosin contraction. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Intraductal cisplatin treatment in a BRCA-associated breast cancer mouse model attenuates tumor development but leads to systemic tumors in aged female mice.

BRCA deficiency predisposes to the development of invasive breast cancer. In BRCA mutation carriers this risk can increase up to 80%. Currently, bilateral prophylactic mastectomy and prophylactic bilateral salpingo-oophorectomy are the only preventive, albeit radical invasive strategies to prevent breast cancer in BRCA mutation carriers. An alternative non-invasive way to prevent BRCA1-associated breast cancer may be local prophylactic treatment via the nipple. Using a non-invasive intraductal (ID) preclinical intervention strategy, we explored the use of combined cisplatin and poly (ADP)-ribose polymerase 1 (PARP1) inhibition to prevent the development of hereditary breast cancer. We show that ID cisplatin and PARP-inhibition can successfully ablate mammary epithelial cells, and this approach attenuated tumor onset in a mouse model of Brca1-associated breast cancer from 153 to 239 days. Long-term carcinogenicity studies in 150 syngeneic wild-type mice demonstrated that tumor incidence was increased in the ID treated mammary glands by 6.3% due to systemic exposure to cisplatin. Although this was only evident in aged mice (median age = 649 days), we conclude that ID cisplatin treatment only presents a safe and feasible local prevention option if systemic exposure to the chemotherapy used can be avoided.

Corrigendum: p120-catenin prevents multinucleation through control of MKLP1-dependent RhoA activity during cytokinesis.

[This corrects the article DOI: 10.1038/ncomms13874.].

p120-catenin prevents multinucleation through control of MKLP1-dependent RhoA activity during cytokinesis.

Spatiotemporal activation of RhoA and actomyosin contraction underpins cellular adhesion and division. Loss of cell-cell adhesion and chromosomal instability are cardinal events that drive tumour progression. Here, we show that p120-catenin (p120) not only controls cell-cell adhesion, but also acts as a critical regulator of cytokinesis. We find that p120 regulates actomyosin contractility through concomitant binding to RhoA and the centralspindlin component MKLP1, independent of cadherin association. In anaphase, p120 is enriched at the cleavage furrow where it binds MKLP1 to spatially control RhoA GTPase cycling. Binding of p120 to MKLP1 during cytokinesis depends on the N-terminal coiled-coil domain of p120 isoform 1A. Importantly, clinical data show that loss of p120 expression is a common event in breast cancer that strongly correlates with multinucleation and adverse patient survival. In summary, our study identifies p120 loss as a driver event of chromosomal instability in cancer.

DNA promoter hypermethylation in nipple fluid: a potential tool for early breast cancer detection.

INTRODUCTION: Nipple fluid aspiration provides direct non-invasive sampling of fluid from the mammary ductal system, where the majority of breast cancers originate. DNA promoter hypermethylation ("methylation") occurs early and at high frequency in breast carcinogenesis, bearing the potential as a biomarker for cancer detection at its earliest stages. We assessed methylation in nipple fluid from breasts of healthy women, of women with sporadic breast cancer and of their contralateral breasts. Our goal was to investigate whether nipple fluid can be used as a reliable methylation biomarker source. MATERIALS AND METHODS: Methylation levels of 13 genes were analysed by quantitative multiplex-methylation specific PCR (QM-MSP) in nipple fluid samples from breasts of healthy women, and from the affected and contralateral breasts of breast cancer patients. RESULTS: Methylation analysis of the low-volume nipple fluid samples was feasible. Despite the generally low methylation levels, cancerous and healthy breasts nipple fluid could be discriminated with an area under the receiver operating characteristic curve (AUC) of 0.64 (p<0.01) based on a multivariate model including AKR1B1, ALX1, RASSF1A and TM6SF1. Within-patient differences between cancerous and contralateral nipple fluid samples were less prominent. CONCLUSIONS: Cancerous nipple fluid contains increased levels of methylation of tumor suppressor genes that potentially could serve as a biomarker for early breast cancer detection.

Validation of DNA promoter hypermethylation biomarkers in breast cancer--a short report.

PURPOSE: DNA promoter hypermethylation of tumor suppressor genes is known to occur early in cancer development, including breast cancer. To improve early breast cancer detection, we aimed to investigate whether the identification of DNA promoter hypermethylation might be of added value. METHODS: The methylation status of a panel of 19 candidate genes (AKR1B1, ALX1, ARHGEF7, FZD10, GHSR, GPX7, GREM1, GSTP1, HOXD1, KL, LHX2, MAL, MGMT, NDRG2, RASGRF2, SFRP1, SFRP2, TM6SF1 and TMEFF2) was determined in formalin-fixed paraffin-embedded normal breast and breast cancer tissue samples using gel-based methylation-specific PCR (MSP). RESULTS: The promoters of the AKR1B1, ALX1, GHSR, GREM1, RASGRF2, SFRP2, TM6SF1 and TMEFF2 genes were found to be significantly differentially methylated in normal versus malignant breast tissues. Based on sensitivity, specificity and logistic regression analyses the best performing genes for detecting breast cancer were identified. Through multivariate analyses, we found that AKR1B1 and TM6SF1 could detect breast cancer with an area under the curve (AUC) of 0.986 in a receiver operating characteristic (ROC) assessment. CONCLUSIONS: Based on our data, we conclude that AKR1B1 and TM6SF1 may serve as candidate methylation biomarkers for early breast cancer detection. Further studies are underway to evaluate the methylation status of these genes in body fluids, including nipple aspirates and blood.

Clonal intratumor heterogeneity of promoter hypermethylation in breast cancer by MS-MLPA.

Intratumor heterogeneity may lead to sampling bias and may present major challenges to personalized medicine and biomarker development. Despite many studies investigating genetic heterogeneity, epigenetic intratumor heterogeneity of promoter hypermethylation has only rarely been examined in breast cancer. To examine clonal intratumor heterogeneity of promotor hypermethylation, we performed methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) for 24 established tumor-suppressor genes on multiple spatially separated samples obtained from 21 primary breast carcinomas. Multiregion analysis was performed, representing at least two and a maximum of five tumor blocks per patient and four areas per tumor block. Methylation results were heterogeneous at one or more genetic loci between different tumor regions in 95% of breast carcinomas. The most heterogeneous loci in decreasing frequency were RASSF1A (62%), CDKN2B (43%), APC (38%), GSTP1 (33%), CDH13 (24%), DAPK1 (19%), and CDKN1B (5%). Heterogeneity lead to a methylation status change in at least one locus in 65% of the tumors. For most genes, the relative contribution of between-patients and between-block variability to the total variation in methylation results was similar. Regardless of the gene, contribution of within-block variability was of little importance. In conclusion, although most variation in methylation status is present between individual breast cancers, clonal epigenetic heterogeneity is seen within most primary breast carcinomas, indicating that methylation results from a single random sample may not be representative of the whole tumor.