Racial Disparity in Quadruple Negative Breast Cancer: Aggressive Biology and Potential Therapeutic Targeting and Prevention.

Black/African-American (AA) women, relative to their White/European-American (EA) counterparts, experience disproportionately high breast cancer mortality. Central to this survival disparity, Black/AA women have an unequal burden of aggressive breast cancer subtypes, such as triple-negative breast cancer (ER/PR-, HER2-wild type; TNBC). While TNBC has been well characterized, recent studies have identified a highly aggressive androgen receptor (AR)-negative subtype of TNBC, quadruple-negative breast cancer (ER/PR-, HER2-wildtype, AR-; QNBC). Similar to TNBC, QNBC disproportionately impacts Black/AA women and likely plays an important role in the breast cancer survival disparities experienced by Black/AA women. Here, we discuss the racial disparities of QNBC and molecular signaling pathways that may contribute to the aggressive biology of QNBC in Black/AA women. Our immediate goal is to spotlight potential prevention and therapeutic targets for Black/AA QNBC; ultimately our goal is to provide greater insight into reducing the breast cancer survival burden experienced by Black/AA women.

Impact of Federal, State, and Local Housing Policies on Disparities in Cardiovascular Disease in Black/African American Men and Women: From Policy to Pathways to Biology.

Racist and discriminatory federal, state, and local housing policies significantly contribute to disparities in cardiovascular disease incidence and mortality for individuals that self-identify as Black or African American. Here we highlight three key housing policies - "redlining," zoning, and the construction of highways - which have wrought a powerful, sustained, and destructive impact on cardiovascular health in Black/African American communities. Redlining and highway construction policies have restricted access to quality health care, increased exposure to carcinogens such as PM2.5, and increased exposure to extreme heat. At the root of these policy decisions are longstanding, toxic societal factors including racism, segregation, and discrimination, which also serve to perpetuate racial inequities in cardiovascular health. Here, we review these societal and structural factors and then link them with biological processes such as telomere shortening, allostatic load, oxidative stress, and tissue inflammation. Lastly, we focus on the impact of inflammation on the immune system and the molecular mechanisms by which the inflamed immune microenvironment promotes the formation of atherosclerotic plaques. We propose that racial residential segregation and discrimination increases tissue inflammation and cytokine production, resulting in dysregulated immune signaling, which promotes plaque formation and cardiovascular disease. This framework has the power to link structural racism not only to cardiovascular disease, but also to cancer.

Epigenetic Dysregulation of KCNK9 Imprinting and Triple-Negative Breast Cancer.

Genomic imprinting is an inherited form of parent-of-origin specific epigenetic gene regulation that is dysregulated by poor prenatal nutrition and environmental toxins. KCNK9 encodes for TASK3, a pH-regulated potassium channel membrane protein that is overexpressed in 40% of breast cancer. However, KCNK9 gene amplification accounts for increased expression in <10% of these breast cancers. Here, we showed that KCNK9 is imprinted in breast tissue and identified a differentially methylated region (DMR) controlling its imprint status. Hypomethylation at the DMR, coupled with biallelic expression of KCNK9, occurred in 63% of triple-negative breast cancers (TNBC). The association between hypomethylation and TNBC status was highly significant in African-Americans (p = 0.006), but not in Caucasians (p = 0.70). KCNK9 hypomethylation was also found in non-cancerous tissue from 77% of women at high-risk of developing breast cancer. Functional studies demonstrated that the KCNK9 gene product, TASK3, regulates mitochondrial membrane potential and apoptosis-sensitivity. In TNBC cells and non-cancerous mammary epithelial cells from high-risk women, hypomethylation of the KCNK9 DMR predicts for increased TASK3 expression and mitochondrial membrane potential (p < 0.001). This is the first identification of the KCNK9 DMR in mammary epithelial cells and demonstration that its hypomethylation in breast cancer is associated with increases in both mitochondrial membrane potential and apoptosis resistance. The high frequency of hypomethylation of the KCNK9 DMR in TNBC and non-cancerous breast tissue from high-risk women provides evidence that hypomethylation of the KNCK9 DMR/TASK3 overexpression may serve as a marker of risk and a target for prevention of TNBC, particularly in African American women.

Spatiotemporal strategies to identify aggressive biology in precancerous breast biopsies.

Over 90% of breast cancer is cured; yet there remain highly aggressive breast cancers that develop rapidly and are extremely difficult to treat, much less prevent. Breast cancers that rapidly develop between breast image screening are called "interval cancers." The efforts of our team focus on identifying multiscale integrated strategies to identify biologically aggressive precancerous breast lesions. Our goal is to identify spatiotemporal changes that occur prior to development of interval breast cancers. To accomplish this requires integration of new technology. Our team has the ability to perform single cell in situ transcriptional profiling, noncontrast biological imaging, mathematical analysis, and nanoscale evaluation of receptor organization and signaling. These technological innovations allow us to start to identify multidimensional spatial and temporal relationships that drive the transition from biologically aggressive precancer to biologically aggressive interval breast cancer. This article is categorized under: Cancer > Computational Models Cancer > Molecular and Cellular Physiology Cancer > Genetics/Genomics/Epigenetics.

Environmental Exposures during Puberty: Window of Breast Cancer Risk and Epigenetic Damage.

During puberty, a woman's breasts are vulnerable to environmental damage ("window of vulnerability"). Early exposure to environmental carcinogens, endocrine disruptors, and unhealthy foods (refined sugar, processed fats, food additives) are hypothesized to promote molecular damage that increases breast cancer risk. However, prospective human studies are difficult to perform and effective interventions to prevent these early exposures are lacking. It is difficult to prevent environmental exposures during puberty. Specifically, young women are repeatedly exposed to media messaging that promotes unhealthy foods. Young women living in disadvantaged neighborhoods experience additional challenges including a lack of access to healthy food and exposure to contaminated air, water, and soil. The purpose of this review is to gather information on potential exposures during puberty. In future directions, this information will be used to help elementary/middle-school girls to identify and quantitate environmental exposures and develop cost-effective strategies to reduce exposures.

Elevated leptin disrupts epithelial polarity and promotes premalignant alterations in the mammary gland.

Obesity is a highly prevalent and modifiable breast cancer risk factor. While the role of obesity in fueling breast cancer progression is well established, the mechanisms linking obesity to breast cancer initiation are poorly understood. A hallmark of breast cancer initiation is the disruption of apical polarity in mammary glands. Here we show that mice with diet-induced obesity display mislocalization of Par3, a regulator of cellular junctional complexes defining mammary epithelial polarity. We found that epithelial polarity loss also occurs in a 3D coculture system that combines acini with human mammary adipose tissue, and establish that a paracrine effect of the tissue adipokine leptin causes loss of polarity by overactivation of the PI3K/Akt pathway. Leptin sensitizes non-neoplastic cells to proliferative stimuli, causes mitotic spindle misalignment, and expands the pool of cells with stem/progenitor characteristics, which are early steps for cancer initiation. We also found that normal breast tissue samples with high leptin/adiponectin transcript ratio characteristic of obesity have an altered distribution of apical polarity markers. This effect is associated with increased epithelial cell layers. Our results provide a molecular basis for early alterations in epithelial architecture during obesity-mediated cancer initiation.

Engaging Underrepresented Adolescents in Authentic Scientific Settings: Scientist Role Models and Improving Psychosocial Outcomes.

There is a critical need for more effective comprehensive programs to increase the number of underrepresented minority students pursuing scientific careers. Science education often is fragmented, delivered with single-focused approaches - traditional classroom lectures, or hands-on-activities, or conducting research. The current paper examines a comprehensive biomedical research program that integrated classroom teaching, hands-on-activities, conducting a research study, and mentoring from scientists in authentic scientific settings. We assessed short-term psychosocial outcomes and long-term academic outcomes in the participants, largely underrepresented minority high school students. The psychosocial outcomes assessed pre and post program include: knowledge of science pathways, attitudes toward science, self-efficacy in science, and scientific communication skills. Post-program results showed an increasing trend for knowledge of science pathways, attitudes toward science, and self-efficacy in science. Post-program, students also reported significant increases in feeling they had role models in science. A long-term assessment was conducted examining participating students' college attendance and majoring in a STEM field. The long-term assessment showed that 77% of students were attending college, 79% were majoring in STEM, and 75% were planning to pursue additional higher education. Findings provide evidence for the short-term and long-term benefits of a comprehensive biomedical research program conducted in an authentic scientific setting.

Mammographic density: intersection of advocacy, science, and clinical practice.

Purpose: Here we aim to review the association between mammographic density, collagen structure and breast cancer risk. Findings: While mammographic density is a strong predictor of breast cancer risk in populations, studies by Boyd show that mammographic density does not predict breast cancer risk in individuals. Mammographic density is affected by age, parity, menopausal status, race/ethnicity, and body mass index (BMI).New studies normalize mammographic density to BMI may provide a more accurate way to compare mammographic density in women of diverse race and ethnicity. Preclinical and tissue-based studies have investigated the role collagen composition and structure in predicting breast cancer risk. There is emerging evidence that collagen structure may activate signaling pathways associated with aggressive breast cancer biology. Summary: Measurement of film mammographic density does not adequately capture the complex signaling that occurs in women with at-risk collagen. New ways to measure at-risk collagen potentially can provide a more accurate view of risk.

Characterization of hair-follicle side population cells in mouse epidermis and skin tumors.

A subset of cells, termed side-population (SP), which have the ability to efflux Hoeschst 33342, have previously been demonstrated to act as a potential method to isolate stem cells. Numerous stem/progenitor cells have been localized in different regions of the mouse hair follicle (HF). The present study identified a SP in the mouse HF expressing the ABCG2 transporter and MTS24 surface marker. These cells are restricted to the upper isthmus of the HF and have previously been described as progenitor cells. Consistent with their SP characteristic, they demonstrated elevated expression of ABCG2 transporter, which participates in the dye efflux. Analysis of tumor epidermal cell lines revealed a correlation between the number of SP keratinocytes and the grade of malignancy, suggesting that the SP may play a role in malignant progression. Consistent with this idea, the present study observed an increased number of cells expressing ABCG2 and MTS24 in chemically induced skin tumors and skin tumor cell lines. This SP does not express the CD34 surface marker detected in the multipotent stem cells of the bulge region of the HF, which have been defined as tumor initiation cells. The present study concluded that a SP with properties of progenitor cells is localized in the upper isthmus of the HF and is important in mouse skin tumor progression.

Src Inhibition Blocks c-Myc Translation and Glucose Metabolism to Prevent the Development of Breast Cancer.

Preventing breast cancer will require the development of targeted strategies that can effectively block disease progression. Tamoxifen and aromatase inhibitors are effective in addressing estrogen receptor-positive (ER(+)) breast cancer development, but estrogen receptor-negative (ER(-)) breast cancer remains an unmet challenge due to gaps in pathobiologic understanding. In this study, we used reverse-phase protein array to identify activation of Src kinase as an early signaling alteration in premalignant breast lesions of women who did not respond to tamoxifen, a widely used ER antagonist for hormonal therapy of breast cancer. Src kinase blockade with the small-molecule inhibitor saracatinib prevented the disorganized three-dimensional growth of ER(-) mammary epithelial cells in vitro and delayed the development of premalignant lesions and tumors in vivo in mouse models developing HER2(+) and ER(-) mammary tumors, extending tumor-free and overall survival. Mechanistic investigations revealed that Src blockade reduced glucose metabolism as a result of an inhibition in ERK1/2-MNK1-eIF4E-mediated cap-dependent translation of c-Myc and transcription of the glucose transporter GLUT1, thereby limiting energy available for cell growth. Taken together, our results provide a sound rationale to target Src pathways in premalignant breast lesions to limit the development of breast cancers.

Triple-negative breast cancer in African-American women: disparities versus biology.

Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype that disproportionately affects BRCA1 mutation carriers and young women of African origin. There is evidence that African-American women with TNBC have worse clinical outcomes than women of European descent. However, it is unclear whether survival differences persist after adjusting for disparities in access to health-care treatment, co-morbid disease and income. It remains controversial whether TNBC in African-American women is a molecularly distinct disease or whether African-American women have a higher incidence of aggressive biology driven by disparities: there is evidence in support of both. Understanding the relative contributions of biology and disparities is essential for improving the poor survival rate of African-American women with TNBC.

Tumor mechanics and metabolic dysfunction.

Desmosplasia is a characteristic of most solid tumors and leads to fibrosis through abnormal extracellular matrix (ECM) deposition, remodeling, and posttranslational modifications. The resulting stiff tumor stroma not only compromises vascular integrity to induce hypoxia and impede drug delivery, but also promotes aggressiveness by potentiating the activity of key growth, invasion, and survival pathways. Intriguingly, many of the protumorigenic signaling pathways that are mechanically activated by ECM stiffness also promote glucose uptake and aerobic glycolysis, and an altered metabolism is a recognized hallmark of cancer. Indeed, emerging evidence suggests that metabolic alterations and an abnormal ECM may cooperatively drive cancer cell aggression and treatment resistance. Accordingly, improved methods to monitor tissue mechanics and metabolism promise to improve diagnostics and treatments to ameliorate ECM stiffening and elevated mechanosignaling may improve patient outcome. Here we discuss the interplay between ECM mechanics and metabolism in tumor biology and suggest that monitoring these processes and targeting their regulatory pathways may improve diagnostics, therapy, and the prevention of malignant transformation.

Skp2 deficiency inhibits chemical skin tumorigenesis independent of p27(Kip1) accumulation.

S-phase kinase-associated protein 2 (Skp2) functions as the receptor component of the Skp-Cullin-F-box complex and is implicated in the degradation of several cell cycle regulators, such as p21(Cip1), p27(Kip1), p57(Kip2), and cyclin E. Numerous studies in human and experimental tumors have demonstrated low p27(Kip1) levels and elevated Skp2 expression. However, a direct association between the inverse correlation of Skp2 and p27(Kip1) with tumorigenesis has not been demonstrated. Herein, we provide evidence that skin tumorigenesis is inhibited in Skp2(-/-) mice. An analysis of mouse keratinocytes indicates that increased p27(Kip1) levels in Skp2(-/-) epidermis cause reduced cell proliferation that is alleviated in the epidermis from Skp2(-/-)/p27(-/-) compound mice. In contrast, we establish that a p27(Kip1) deficiency does not overturn the reduced skin tumorigenesis experienced by Skp2(-/-) mice. In addition, Skp2(-/-) epidermis exhibits an accumulation of p53-cofactor CBP/p300 that is associated with elevated apoptosis in hair follicles and decreased skin tumorigenesis. We conclude that p27(Kip1) accumulation is responsible for the hypoplasia observed in normal tissues of Skp2(-/-) mice but does not have a preponderant function in reducing skin tumorigenesis.

Evidence for phenotypic plasticity in aggressive triple-negative breast cancer: human biology is recapitulated by a novel model system.

Breast cancers with a basal-like gene signature are primarily triple-negative, frequently metastatic, and carry a poor prognosis. Basal-like breast cancers are enriched for markers of breast cancer stem cells as well as markers of epithelial-mesenchymal transition (EMT). While EMT is generally thought to be important in the process of metastasis, in vivo evidence of EMT in human disease remains rare. Here we report a novel model of human triple-negative breast cancer, the DKAT cell line, which was isolated from an aggressive, treatment-resistant triple-negative breast cancer that demonstrated morphological and biochemical evidence suggestive of phenotypic plasticity in the patient. The DKAT cell line displays a basal-like phenotype in vitro when cultured in serum-free media, and undergoes phenotypic changes consistent with EMT/MET in response to serum-containing media, a unique property among the breast cancer cell lines we tested. This EMT is marked by increased expression of the transcription factor Zeb1, and Zeb1 is required for the enhanced migratory ability of DKAT cells in the mesenchymal state. DKAT cells also express progenitor-cell markers, and single DKAT cells are able to generate tumorspheres containing both epithelial and mesenchymal cell types. In vivo, as few as ten DKAT cells are capable of forming xenograft tumors which display a range of epithelial and mesenchymal phenotypes. The DKAT model provides a novel model to study the molecular mechanisms regulating phenotypic plasticity and the aggressive biology of triple-negative breast cancers.

Combined effect of cyclin D3 expression and abrogation of cyclin D1 prevent mouse skin tumor development.

We have previously demonstrated that ras-mediated skin tumorigenesis depends on signaling pathways that act preferentially through cyclin D1 and D2. Interestingly, the expression of cyclin D3 inhibits skin tumor development, an observation that conflicts with the oncogenic role of D-type cyclins in the mouse epidermis. Here, we show that simultaneous up and downregulation of particular members of the D-type cyclin family is a valuable approach to reduce skin tumorigenesis. We developed the K5D3/cyclin D1(-/-) compound mouse, which overexpresses cyclin D3 but lacks expression of cyclin D1 in the skin. Similar to K5D3 transgenic mice, keratinocytes from K5D3/cyclin D1(-/-) compound mice show a significant reduction of cyclin D2 levels. Therefore, this model allows us to determine the effect of cyclin D3 expression when combined with reduced or absent expression of the remaining two members of the D-type cyclin family in mouse epidermis. Our data show that induced expression of cyclin D3 compensates for the reduced level of cyclin D1 and D2, resulting in normal keratinocyte proliferation. However, simultaneous ablation of cyclin D1 and downregulation of cyclin D2 via cyclin D3 expression resulted in a robust reduction in ras-mediated skin tumorigenesis. We conclude that modulation of the levels of particular members of the D-type cyclin family could be useful to inhibit tumor development and, in particular, ras-mediated tumorigenesis.

Skp2 is necessary for Myc-induced keratinocyte proliferation but dispensable for Myc oncogenic activity in the oral epithelium.

The proto-oncogene c-Myc encodes a transcription factor that is implicated in the regulation of cellular proliferation, differentiation, and apoptosis. Myc accelerates the rate of cell proliferation, at least in part, through its ability to down-regulate the expression of the cell cycle inhibitor p27(Kip1). Moreover, p27(Kip1) protein levels are regulated by ubiquitin-mediated turnover, leading to destruction by the E3 ubiquitin ligase SCF(Skp2). Therefore, we hypothesize that a lack of Skp2 expression should lead to increased p27(Kip1) levels and further inhibition of Myc-mediated proliferation and tumorigenesis. Myc expression in epithelial tissues of transgenic mice (K5-Myc) led to increased keratinocyte proliferation and the development of spontaneous tumors within the oral cavity. We generated K5-Myc-transgenic mice in an Skp2-null background. Consistent with our hypothesis, we found that Myc-mediated keratinocyte hyperproliferation was abolished by the loss of Skp2. However, Skp2 ablation did not affect Myc-driven tumorigenesis because the incidence, latency, and degree of differentiation of oral tumors were identical between K5-Myc/Skp2(+/+) and K5-Myc/Skp2(-/-) mice. Altogether, these findings suggest that Skp2 and p27(Kip1) are critical for Myc-driven keratinocyte proliferation; however, Myc-mediated tumorigenesis in the oral epithelium is independent of the Skp2-p27(Kip1) axis.

Unexpected reduction of skin tumorigenesis on expression of cyclin-dependent kinase 6 in mouse epidermis.

Cyclin-dependent kinases (CDKs) 4 and 6 are important regulators of the G(1) phase of the cell cycle, share 71% amino acid identity, and are expressed ubiquitously. As a result, it was assumed that each of these kinases plays a redundant role regulating normal and neoplastic proliferation. In previous reports, we have described the effects of CDK4 expression in transgenic mice, including the development of epidermal hyperplasia and increased malignant progression to squamous cell carcinoma. To study the role of CDK6 in epithelial growth and tumorigenesis, we generated transgenic mice carrying the CDK6 gene under the keratin 5 promoter (K5CDK6). Similar to K5CDK4 mice, epidermal proliferation increased substantially in K5CDK6 mice; however, no hyperplasia was observed. CDK6 overexpression also triggered keratinocyte apoptosis in interfollicular and follicular epidermis as a compensatory mechanism to override aberrant proliferation. Unexpectedly, CDK6 overexpression results in decreased skin tumor development compared with wild-type siblings. The inhibition in skin tumorigenesis was similar to that previously reported in K5-cyclin D3 mice. Furthermore, biochemical analysis of the K5CDK6 epidermis showed preferential complex formation between CDK6 and cyclin D3, suggesting that this particular complex plays an important role in tumor restraint. These studies provide in vivo evidence that CDK4 and CDK6 play a similar role as a mediator of keratinocyte proliferation but differ in apoptosis activation and skin tumor development.