Rachel E. Ellsworth

Genomic instability in histologically normal breast tissues: implications for carcinogenesis.

Breast cancer is an important contributor to morbidity and mortality in society, but factors that affect the cause of the disease are poorly defined. Genomic instability drives tumorigenic processes in invasive carcinomas and premalignant breast lesions, and might promote the accumulation of genetic alterations in apparently normal tissues before histological abnormalities are detectable. Evidence suggests that genomic changes in breast parenchyma affect the behaviour of epithelial cells, and ultimately might affect tumour growth and progression. Inherent instability in genes that maintain genomic integrity, as well as exogenous chemicals and environmental pollutants, have been implicated in breast-cancer development. Although molecular mechanisms of tumorigenesis are unclear at present, carcinogenic agents could contribute to fields of genomic instability localised to specific areas of the breast. Understanding the functional importance of genomic instability in early carcinogenesis has important implications for improvement of diagnostic and treatment strategies.

Breast Cancer in the Personal Genomics Era

Breast cancer is a heterogeneous disease with a complex etiology that develops from different cellular lineages, progresses along multiple molecular pathways, and demonstrates wide variability in response to treatment. The “standard of care” approach to breast cancer treatment in which all patients receive similar interventions is rapidly being replaced by personalized medicine, based on molecular characteristics of individual patients. Both inherited and somatic genomic variation is providing useful information for customizing treatment regimens for breast cancer to maximize efficacy and minimize adverse side effects. In this article, we review (1) hereditary breast cancer and current use of inherited susceptibility genes in patient management; (2) the potential of newly-identified breast cancer-susceptibility variants for improving risk assessment; (3) advantages and disadvantages of direct-to-consumer testing; (4) molecular characterization of sporadic breast cancer through immunohistochemistry and gene expression profiling and opportunities for personalized prognostics; and (5) pharmacogenomic influences on the effectiveness of current breast cancer treatments. Molecular genomics has the potential to revolutionize clinical practice and improve the lives of women with breast cancer.

Outer breast quadrants demonstrate increased levels of genomic instability.

AbstractBackground: Theory holds that the upper outer quadrant of the breast develops more malignancies because of increased tissue volume. This study evaluated genomic patterns of loss of heterozygosity (LOH) and allelic imbalance (AI) in non-neoplastic tissues from quadrants of diseased breasts following mastectomy to characterize relationships between genomic instability and the propensity for tumor development. Methods: Tissues from breast quadrants were collected from 21 patients with various stages of breast carcinoma. DNA was isolated from non-neoplastic tissues using standard methods and 26 chromosomal regions commonly deleted in breast cancer were examined to assess genomic instability. Results: Genomic instability was observed in breast quadrants from patients with ductal carcinomas in situ and advanced carcinomas. Levels of instability by quadrant were not predictive of primary tumor location (P = .363), but outer quadrants demonstrated significantly higher levels of genomic instability than did inner quadrants (P = .017). Marker D8S511 on chromosome 8p22– 21.3, one of the most frequently altered chromosomal regions in breast cancer, showed a significantly higher level of instability (P = .039) in outer compared with inner quadrants. Conclusions: Non-neoplastic breast tissues often harbor genetic changes that can be important to understanding the local breast environment within which cancer develops. Greater genomic instability in outer quadrants can partially explain the propensity for breast cancers to develop there, rather than simple volume-related concepts. Patterns of field cancerization in the breast appear to be complex and are not a simple function of distance from a developing tumor.

Identification of differentially expressed genes in breast tumors from African American compared with Caucasian women.

Breast tumors from African American women have less favorable pathological characteristics and higher mortality rates than those of Caucasian women. Although socioeconomic status may influence prognosis, biological factors are also likely to contribute to tumor behavior.

Genomic patterns of allelic imbalance in disease free tissue adjacent to primary breast carcinomas.

Mammary stroma plays an important role in facilitating the neoplastic transformation of epithelial cells, modulating integrity of the extracellular matrix, and maintaining genomic stability, but molecular mechanisms by which stroma affects epithelial structure and function are not well-defined. We used laser-assisted microdissection of paraffin-embedded breast tissues from 30 patients with breast disease and a panel of 52 microsatellite markers defining 26 chromosomal regions to characterize genomic patterns of allelic imbalance (AI) in disease-free tissue adjacent to sites of breast disease and to define genomic regions that may contain genes associated with early carcinogenic processes. The mean frequency of AI in histologically normal tissue adjacent to the primary carcinomas (15.4) was significantly higher than that in distant tissue from the same breast (3.7). The pattern of AI across all chromosomal regions differed between the adjacent tissue and primary tumor in every case. Unique AI events, observed only in tumor (15 of informative markers) or only in adjacent cells (10 of informative markers), were far more common than AI events shared between tumor and adjacent cells (~ 4). Levels of AI characteristic of advanced invasive carcinomas were already present in non-invasive ductal carcinomas in situ, and appreciable levels of AI were observed in adjacent non-neoplastic tissue at all pathological stages. Chromosome 11p15.1 showed significantly higher levels of AI in adjacent cells (p < 0.01), suggesting that this region may harbor genes involved in breast cancer development and progression. Our data indicate that genomic instability may be inherently greater in disease-free tissue close to developing tumors, which may have important implications for defining surgical margins and predicting recurrence.

Outcome disparities in African American women with triple negative breast cancer: a comparison of epidemiological and molecular factors between African American and Caucasian women with triple negative breast cancer.

BackgroundAlthough diagnosed less often, breast cancer in African American women (AAW) displays different characteristics compared to breast cancer in Caucasian women (CW), including earlier onset, less favorable clinical outcome, and an aggressive tumor phenotype. These disparities may be attributed to differences in socioeconomic factors such as access to health care, lifestyle, including increased frequency of obesity in AAW, and tumor biology, especially the higher frequency of triple negative breast cancer (TNBC) in young AAW. Improved understanding of the etiology and molecular characteristics of TNBC in AAW is critical to determining whether and how TNBC contributes to survival disparities in AAW.MethodsDemographic, pathological and survival data from AAW (n = 62) and CW (n = 98) with TNBC were analyzed using chi-square analysis, Student’s t-tests, and log-rank tests. Frozen tumor specimens were available from 57 of the TNBC patients (n = 23 AAW; n = 34 CW); RNA was isolated after laser microdissection of tumor cells and was hybridized to HG U133A 2.0 microarrays. Data were analyzed using ANOVA with FDR <0.05, >2-fold difference defining significance.ResultsThe frequency of TNBC compared to all BC was significantly higher in AAW (28%) compared to CW (12%), however, significant survival and pathological differences were not detected between populations. Gene expression analysis revealed the tumors were more similar than different at the molecular level, with only CRYBB2P1, a pseudogene, differentially expressed between populations. Among demographic characteristics, AAW consumed significantly lower amounts of caffeine and alcohol, were less likely to breastfeed and more likely to be obese.ConclusionsThese data suggest that TNBC in AAW is not a unique disease compared to TNBC in CW. Rather, higher frequency of TNBC in AAW may, in part, be attributable to the effects of lifestyle choices. Because these risk factors are modifiable, they provide new opportunities for the development of risk reduction strategies that may decrease mortality by preventing the development of TNBC in AAW.

Molecular heterogeneity in breast cancer: State of the science and implications for patient care.

The identification of extensive genetic heterogeneity in human breast carcinomas poses a significant challenge for designing effective treatment regimens. Significant genomic evolution often occurs during breast cancer progression, creating variability within primary tumors as well as between the primary carcinoma and metastases. Current risk allocations and treatment recommendations for breast cancer patients are based largely on characteristics of the primary tumor; however, genetic differences between disseminated tumor cells and the primary carcinoma may negatively impact treatment efficacy and survival. In this review we (1) present current information about genomic variability within primary breast carcinomas, between primary tumors and regional/distant metastases, among circulating tumor cells (CTCs) and disseminated tumor cells (DTCs), and in cell-free nucleic acids in circulation, and (2) describe how this heterogeneity affects clinical care and outcomes such as recurrence and therapeutic resistance. Understanding the evolution and functional significance of the composite breast cancer genome within each patient is critical for developing effective therapies that can overcome obstacles presented by molecular heterogeneity.

Amplification of HER2 is a marker for global genomic instability

BackgroundGenomic alterations of the proto-oncogene c-erbB-2 (HER-2/neu) are associated with aggressive behavior and poor prognosis in patients with breast cancer. The variable clinical outcomes seen in patients with similar HER2 status, given similar treatments, suggests that the effects of amplification of HER2 can be influenced by other genetic changes. To assess the broader genomic implications of structural changes at the HER2 locus, we investigated relationships between genomic instability and HER2 status in patients with invasive breast cancer.MethodsHER2 status was determined using the PathVysion® assay. DNA was extracted after laser microdissection from the 181 paraffin-embedded HER2 amplified (n = 39) or HER2 negative (n = 142) tumor specimens with sufficient tumor available to perform molecular analysis. Allelic imbalance (AI) was assessed using a panel of microsatellite markers representing 26 chromosomal regions commonly altered in breast cancer. Student t-tests and partial correlations were used to investigate relationships between genomic instability and HER2 status.ResultsThe frequency of AI was significantly higher (P < 0.005) in HER2 amplified (27%) compared to HER2 negative tumors (19%). Samples with HER2 amplification showed significantly higher levels of AI (P < 0.05) at chromosomes 11q23, 16q22-q24 and 18q21. Partial correlations including ER status and tumor grade supported associations between HER2 status and alterations at 11q13.1, 16q22-q24 and 18q21.ConclusionThe poor prognosis associated with HER2 amplification may be attributed to global genomic instability as cells with high frequencies of chromosomal alterations have been associated with increased cellular proliferation and aggressive behavior. In addition, high levels of DNA damage may render tumor cells refractory to treatment. In addition, specific alterations at chromosomes 11q13, 16q22-q24, and 18q21, all of which have been associated with aggressive tumor behavior, may serve as genetic modifiers to HER2 amplification. These data not only improve our understanding of HER in breast pathogenesis but may allow more accurate risk profiles and better treatment options to be developed.

Management of Incidental Findings in the Era of Next-generation Sequencing

Next-generation sequencing (NGS) technologies allow for the generation of whole exome or whole genome sequencing data, which can be used to identify novel genetic alterations associated with defined phenotypes or to expedite discovery of functional variants for improved patient care. Because this robust technology has the ability to identify all mutations within a genome, incidental findings (IF)- genetic alterations associated with conditions or diseases unrelated to the patient’s present condition for which current tests are being performed- may have important clinical ramifications. The current debate among genetic scientists and clinicians focuses on the following questions: 1) should any IF be disclosed to patients, and 2) which IF should be disclosed – actionable mutations, variants of unknown significance, or all IF? Policies for disclosure of IF are being developed for when and how to convey these findings and whether adults, minors, or individuals unable to provide consent have the right to refuse receipt of IF. In this review, we detail current NGS technology platforms, discuss pressing issues regarding disclosure of IF, and how IF are currently being handled in prenatal, pediatric, and adult patients.

Timing of Critical Genetic Changes in Human Breast Disease

BackgroundBreast cancer development has been characterized as a nonobligatory sequence of histological changes from normal epithelium through invasive malignancy. Although genetic alterations are thought to accumulate stochastically during tumorigenesis, little is known about the timing of critical mutations. This study examined allelic imbalance (AI) in tissue samples representing a continuum of breast cancer development to examine the evolution of genomic instability.MethodsLaser-microdissected DNA samples were collected from histologically normal breast specimens (n = 25), atypical ductal hyperplasia (ADH, n = 16), ductal carcinoma-in-situ (DCIS, n = 37), and stage I to III invasive carcinomas (n = 72). Fifty-two microsatellite markers representing 26 chromosomal regions commonly deleted in breast cancer were used to assess patterns of AI.ResultsAI frequencies were <5% in histologically normal and ADH specimens, 20% in DCIS lesions, and approximately 25% in invasive tumors. Mann-Whitney tests showed (1) that levels of AI in ADH samples did not differ significantly from those in histologically normal tissues and (2) that AI frequencies in DCIS lesions were not significantly different from those in invasive carcinomas. ADH and DCIS samples, however, differed significantly (P < .0001).ConclusionsDCIS lesions contain levels of genomic instability that are characteristic of advanced invasive tumors, and this suggests that the biology of a developing carcinoma may already be predetermined by the in situ stage. Observations that levels of AI in ADH lesions are similar to those in disease-free tissues provide a genomic rationale for why prevention strategies at the ADH level are successful and why cases with ADH involving surgical margins do not require further resection.