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Background Breast cancer is the most common invasive cancer among women worldwide. Next-generation sequencing (NGS) has revolutionized the study of cancer across research labs around the globe; however, genomic testing in clinical settings remains limited. Advances in sequencing reliability, pipeline analysis, accumulation of relevant data, and the reduction of costs are rapidly increasing the feasibility of NGS-based clinical decision making. Methods We report the development of MammaSeq, a breast cancer-specific NGS panel, targeting 79 genes and 1369 mutations, optimized for use in primary and metastatic breast cancer. To validate the panel, 46 solid tumors and 14 plasma circulating tumor DNA (ctDNA) samples were sequenced to a mean depth of 2311× and 1820×, respectively. Variants were called using Ion Torrent Suite 4.0 and annotated with cravat CHASM. CNVKit was used to call copy number variants in the solid tumor cohort. The oncoKB Precision Oncology Database was used to identify clinically actionable variants. Droplet digital PCR was used to validate select ctDNA mutations. Results In cohorts of 46 solid tumors and 14 ctDNA samples from patients with advanced breast cancer, we identified 592 and 43 protein-coding mutations. Mutations per sample in the solid tumor cohort ranged from 1 to 128 (median 3), and the ctDNA cohort ranged from 0 to 26 (median 2.5). Copy number analysis in the solid tumor cohort identified 46 amplifications and 35 deletions. We identified 26 clinically actionable variants (levels 1–3) annotated by OncoKB, distributed across 20 out of 46 cases (40%), in the solid tumor cohort. Allele frequencies of ESR1 and FOXA1 mutations correlated with CA.27.29 levels in patient-matched blood draws. Conclusions In solid tumor biopsies and ctDNA, MammaSeq detects clinically actionable mutations (OncoKB levels 1–3) in 22/46 (48%) solid tumors and in 4/14 (29%) of ctDNA samples. MammaSeq is a targeted panel suitable for clinically actionable mutation detection in breast cancer. Electronic supplementary material The online version of this article (10.1186/s13058-019-1102-7) contains supplementary material, which is available to authorized users.

BackgroundBreast cancer is the most common malignancy to spread to the orbit and periorbit, and the invasive lobular carcinoma (ILC) histologic subtype of breast cancer has been reported to form these ophthalmic metastases (OM) more frequently than invasive ductal carcinomas (IDC). We herein report our single academic institution experience with breast cancer OM with respect to anatomical presentation, histology (lobular vs. ductal), treatment, and survival.MethodsWe employed the natural language processing platform, TIES (Text Information Extraction System), to search 2.3 million de-identified patient pathology and radiology records at our institution in order to identify patients with OM secondary to breast cancer. We then compared the resultant cohort, the “OM cohort,” to two other representative metastatic breast cancer patient (MBC) databases from our institution. Histological analysis of selected patients was performed.ResultsOur TIES search and manual refinement ultimately identified 28 patients who were diagnosed with breast cancer between 1995 and 2016 that subsequently developed OM. Median age at diagnosis was 54 (range 28–77) years of age. ER, PR, and HER2 status from the 28 patients with OM did not differ from other patients with MBC from our institution. The relative proportion of patients with ILC was significantly higher in the OM cohort (32.1%) than in other MBC patients in our institution (11.3%,p = 0.007). Median time to first OM in the OM cohort was 46.7 months, and OM were the second most frequent first metastases after bony metastases. After diagnosis of the first distant metastasis of any kind, median survival of patients with ILC (21.4 months) was significantly shorter than that of patients with IDC (55.3 months,p = 0.03). Nine patients developed bilateral OM. We observed a significant co-occurrence of OM and central nervous system metastases (p = 0.0053). The histological analysis revealed an interesting case in which the primary tumor was of a mixed ILC/IDC subtype, while only ILC was present in the OM.ConclusionsOM from breast cancer are illustrative of the difference in metastatic behavior of ILC versus IDC and should be considered when treating patients with ILC, especially in those with complaints of visual acuity changes.

Background Mutations in the estrogen receptor alpha (ERα) 1 gene (ESR1) are frequently detected in ER+ metastatic breast cancer, and there is increasing evidence that these mutations confer endocrine resistance in breast cancer patients with advanced disease. However, their functional role is not well-understood, at least in part due to a lack of ESR1 mutant models. Here, we describe the generation and characterization of genome-edited T47D and MCF7 breast cancer cell lines with the two most common ESR1 mutations, Y537S and D538G. Methods Genome editing was performed using CRISPR and adeno-associated virus (AAV) technologies to knock-in ESR1 mutations into T47D and MCF7 cell lines, respectively. Various techniques were utilized to assess the activity of mutant ER, including transactivation, growth and chromatin-immunoprecipitation (ChIP) assays. The level of endocrine resistance was tested in mutant cells using a number of selective estrogen receptor modulators (SERMs) and degraders (SERDs). RNA sequencing (RNA-seq) was employed to study gene targets of mutant ER. Results Cells with ESR1 mutations displayed ligand-independent ER activity, and were resistant to several SERMs and SERDs, with cell line and mutation-specific differences with respect to magnitude of effect. The SERD AZ9496 showed increased efficacy compared to other drugs tested. Wild-type and mutant cell co-cultures demonstrated a unique evolution of mutant cells under estrogen deprivation and tamoxifen treatment. Transcriptome analysis confirmed ligand-independent regulation of ERα target genes by mutant ERα, but also identified novel target genes, some of which are involved in metastasis-associated phenotypes. Despite significant overlap in the ligand-independent genes between Y537S and D538G, the number of mutant ERα-target genes shared between the two cell lines was limited, suggesting context-dependent activity of the mutant receptor. Some genes and phenotypes were unique to one mutation within a given cell line, suggesting a mutation-specific effect. Conclusions Taken together, ESR1 mutations in genome-edited breast cancer cell lines confer ligand-independent growth and endocrine resistance. These biologically relevant models can be used for further mechanistic and translational studies, including context-specific and mutation site-specific analysis of the ESR1 mutations. Electronic supplementary material The online version of this article (doi:10.1186/s13058-017-0851-4) contains supplementary material, which is available to authorized users.

The continuing advancement of sequencing technologies has made the systematic identification of all driving somatic events in cancer a possibility. In the June 2012 issue of Nature, five papers show some significant headway in this endeavor, each a herculean effort of genome sequencing, and transcriptome and copy number analysis resulting in data on thousands of breast cancers. Integrating these massive datasets, the authors were able to further subdivide breast cancer and identify a number of novel driver genes. While the studies represent a leap forward in describing the genomics of breast cancer, and clearly highlight the tremendous diversity between tumors, the studies only scrape the surface of molecular changes in breast tumors, with more granularity to come from the study of epigenomics, single cell sequencing, and so on. The immediate importance of the data to clinical care is currently unknown, and will depend upon detailed identification and functional analysis of driver mutations.

Insulin-like growth factors (IGFs) are potent mitogens and survival factors. In the mammary gland, IGFs stimulate proliferation, differentiation, and survival during numerous developmental stages; IGF signaling is required for puberty-dependent ductal outgrowth, stimulates lobuloalveolar development during pregnancy, and is reduced or absent during apoptosis-driven involution. Much of our knowledge of IGF action in the mammary gland in vivo comes from knockout or transgenic models. However, very few of these studies have examined the consequence of these gene alterations on IGF signaling in vivo. We have recently shown that intravenous injection of IGF-I stimulates IGF-IR and IRS phosphorylation in the mammary gland, and we are currently assessing the effect of targeted gene deletion of overexpression of IGF signaling components on downstream signaling in the mammary gland in vivo. Many years of research have shown that the proliferative and survival functions of the IGFs are not only important in mammary gland development, but are also strongly involved in mammary cancer. Early work using breast cancer cell lines in vitro showed that IGFs could increase cell growth and survival; in particular, that IGFs could block the effects of chemotherapy. We have recently shown that breast cancer cell lines grown as xenografts in vivo are also sensitive to intravenous IGF stimulation, and several IGF-IR inhibitors have been shown in the past year to block MCF-7 xenograft growth. IGF-IR and its downstream signaling intermediate IRS-1 can transform fibroblasts in vitro. To date there is no evidence for their transforming ability in vivo using transgenic mice. We have recently characterized mice that overexpress IGF-IR, IRS-1, or IRS-2 in the mammary gland, using mouse mammary tumor virus directed overexpression. We have found that overexpression of a constitutively active IGF-IR in the mammary gland disrupts normal development, such that female mice are unable to lactate, and that mice rapidly develop mammary tumors. Interestingly, overexpression of IRS-1 or IRS-2 also causes mammary tumorigenesis, albeit with a longer time to tumor formation than dominant active IGF-IR. These are the first mouse models showing that IGF-IR or IRS overexpression leads to tumorigenesis in vivo. We are currently examining the pathways required for IGF-IR and IRS-mediated tumorigenesis in the mammary gland.

The San Antonio Breast Cancer Symposium is the largest annual meeting devoted solely to breast cancer research. The late William L McGuire's vision for this meeting was to stimulate 'translational research', many years before this term became popular. In this way, the San Antonio Breast Cancer Symposium represents a forum in which basic and clinical researchers present their research side by side. Each year sees the continued evolution of our understanding of the basic mechanisms of breast cancer initiation and progression, and the clinical application of this knowledge. Major topics of discussion at the symposium this year were the cell cycle, new evolving concepts of estrogen receptor action, breast cancer stem cells, new predictive and prognostic markers (including microarray studies), and continued exploration of the mechanisms of drug resistance. This report will summarize preclinical and translational highlights from the meeting.