Chromosomal Instability In Breast Cancer Research Articles

Exploring chromosomal instability and clonal heterogeneity in breast cancer.

Chromosomal instability (CIN), characterized by fluctuations in chromosome number or structure within cells, stands out as a hallmark of cancer, enabling tumors to thrive in hostile conditions. CIN serves as a driver of genetic diversity, giving rise to clonal heterogeneity (CH). Emerging evidence points to a potential correlation between CIN, CH, and the prognosis of breast cancer (BC) patients, especially in tumors exhibiting overexpression of the human epidermal growth factor receptor 2 (HER2+). However, our understanding of the role of CIN in other subtypes of BC is limited. Furthermore, it remains unclear whether CIN levels above a certain threshold in BC tumors could adversely affect tumor growth, or if lower to moderate levels of CIN might be associated with a more favorable prognosis for BC patients compared to elevated levels. Elucidating these relationships could significantly influence risk assessment and the formulation of future therapeutic approaches targeting CIN in BC. This study aimed to assess CIN and CH in tumor tissue samples obtained from Colombian patients diagnosed with luminal A, luminal B, HER2+, or triple-negative BC, and compare them with established clinicopathological parameters. The findings of this study indicate that BC patients, exhibit intermediate CIN, high CH, and stable aneuploidy. All these characteristics were found to be related with clinicopathological features. Our results suggest that the identification of CIN, CH and aneuploidy could improve cancer risk stratification, which could help to clarify the prediction of clinical outcomes and guiding personalized therapeutic strategies for patients with different BC subtypes.

Misaligned Chromosomes are a Major Source of Chromosomal Instability in Breast Cancer.

We surveyed the single-cell landscape of mitotic defects that generate CIN in primary and metastatic breast cancer and relevant models. Misaligned chromosomes predominate, and are less immunostimulatory than other chromosome segregation errors.

SIRT7 Downregulation Promotes Breast Cancer Metastasis Via LAP2α-Induced Chromosomal Instability.

Chromosomal instability (CIN) plays an important role in the initiation and progression of carcinomas. However, the regulatory mechanism of metastasis mediated by CIN in breast cancer is not fully understood. Here, we aimed to demonstrate that the deregulation of SIRT7 and lamina-associated polypeptide 2α (LAP2α) critically contributes to CIN-induced metastasis in breast cancer. Expression of SIRT7 and chromosome stability-related genes was examined using western blotting, quantitative real-time PCR, immunohistochemistry, and immunofluorescence; functional significance of SIRT7 was examined using in vitro and in vivo models; and interaction between SIRT7 and LAP2α was assessed by co-inmunoprecipitation (Co-IP) assays. Doxorubicin (DOX) inhibited SIRT7 expression and enhanced CIN in breast cancer cells; SIRT7 deficiency led to CIN in breast cancer cells. Co-IP approach and immunohistochemistry demonstrated that SIRT7 interacted directly and positively with LAP2α and SIRT7 knockdown led to increased ubiquitination-dependent degradation of LAP2α and reduced protein levels of LAP2α, whereas LAP2α knockdown did not affect SIRT7 expression. In vitro and in vivo evidence revealed that SIRT7 promotes breast cancer metastasis through the SIRT7/LAP2α axis. In summary, SIRT7 interacts with LAP2α to regulate CIN and metastasis in breast cancer, and inhibition of SIRT7/LAP2α axis represents a potential therapeutic strategy for preventing breast cancer metastasis.

Centrosome Aberrations as Drivers of Chromosomal Instability in Breast Cancer.

Chromosomal instability (CIN), or the dynamic change in chromosome number and composition, has been observed in cancer for decades. Recently, this phenomenon has been implicated as facilitating the acquisition of cancer hallmarks and enabling the formation of aggressive disease. Hence, CIN has the potential to serve as a therapeutic target for a wide range of cancers. CIN in cancer often occurs as a result of disrupting key regulators of mitotic fidelity and faithful chromosome segregation. As a consequence of their essential roles in mitosis, dysfunctional centrosomes can induce and maintain CIN. Centrosome defects are common in breast cancer, a heterogeneous disease characterized by high CIN. These defects include amplification, structural defects, and loss of primary cilium nucleation. Recent studies have begun to illuminate the ability of centrosome aberrations to instigate genomic flux in breast cancer cells and the tumor evolution associated with aggressive disease and poor patient outcomes. Here, we review the role of CIN in breast cancer, the processes by which centrosome defects contribute to CIN in this disease, and the emerging therapeutic approaches that are being developed to capitalize upon such aberrations.

Centromere 17 copy number gain reflects chromosomal instability in breast cancer

Chromosomal instability (CIN) is known to be associated with prognosis and treatment response in breast cancer. This study was conducted to determine whether copy number gain of centromere 17 (CEP17) reflects CIN, and to evaluate the prognostic and predictive value of CIN in breast cancer. CIN status was determined by summing copy number gains of four centromeric probes (CEP1, CEP8, CEP11, and CEP16) based on fluorescence in situ hybridization and CIN scores were calculated using next generation sequencing data. High CIN was associated with adverse clinicopatholgical parameters of breast cancer. Among them, positive HER2 status, high Ki-67 index and CEP17 copy number gain were found to be independent predictors of high CIN. High CIN was associated with poor clinical outcome of the patients in the whole group, as well as in luminal/HER2-negative and HER2-positive subtypes. CEP17 copy number was significantly higher in the high-CIN-score group than in the low-CIN-score group. A positive linear correlation between the mean CEP17 copy number and the CIN score was found. In conclusion, CEP17 copy number was confirmed as a useful predictor for CIN in breast cancer, and high CIN was revealed as an indicator of poor prognosis in breast cancer.

9PD - Aurora-B mediated snail phosphorylation is essential for mitotic spindle checkpoint and for preventing chromosomal instability in breast cancer

BackgroundSnail is a transcriptional repressor that promotes epithelial-to-mesenchymal transition (EMT). Snail is upregulated in breast cancer and it is associated with chromosomal instability (CIN) and metastasis. The mitotic spindle assembly checkpoint (SAC) is a critical mechanism to prevent CIN. Extensive studies have investigated signaling pathways that involve in protein-protein interactions and post translational modifications. However, it not known how mitosis might be impacted by transcriptional regulation. Due to the fact that Snail is a transcriptional regulator and that Snail is upregulated in breast cancer with CIN, it is critical to understand whether and how abnormally regulated Snail contributes to breast cancer CIN via transcriptional regulation. MethodsWe used siRNA to knockdown Snail in breast caner cell lines to look for a change in SAC. We developed a phosphor-specific antibody to study Aurora-B mediated phosphorylation. Site specific mutagenesis was used to generate phosphorylation site mutants, and exogenous proteins were expressed in cells to look for an effect in SAC and CIN. ResultsWe found that Snail is an essential element that is required for the SAC. Genetic depletion of Snail in breast cancer cells by siRNA led to a defect in activation of SAC (measured by accumulation of Histone Serine 10 phosphorylation in response to spindle damaging agents). Interestingly we also found that the Snail protein level was significantly enhanced in the presence of nocodazole. Further, we found that Snail upregulation was through phosphorylation in an Aurora-B dependent manner. We identified a serine site that can be phosphorylated by Aurora-B. Using a phosphor-specific antibody we have developed, we further proved that the phosphorylation event was initiated in mitosis. Functional studies reveal a critical role for Snail phosphorylation in activation of the SAC and prevention of CIN. Conclusions1. Snail is required for mitotic spindle checkpoint activation; 2. Snail is phosphorylated by Aurora B in mitosis; 3. Aurora-B medicated Snail phosphorylation is required for optinal activation of the spindle checkpoint and prevention of chromosomal instability in breast cancer. Legal entity responsible for the studyThe authors. FundingTianjin Medical University Cancer Institute and Hospital. DisclosureAll authors have declared no conflicts of interest.

Chromosome 17 centromere amplification and chromosomal instability (CIN) in breast cancer: Pathogenic and therapeutic implications.

Chromosomal instability (CIN) is present in variable degrees in a significant percentage (up to 90%) of cancers and often portends adverse outcomes. However, it has not been incorporated in clinical practice as a prognostic marker due to the lack of standardization and proof of clinical utility of assays to measure it, as well as uncertainties regarding optimal cut-offs. Amplification of the centromeric region of chromosome 17 as measured by In Situ Hybridization (ISH) of the CEP17 probe is used clinically as part of the ISH assay for HER2 status determination in breast cancer in cases with intermediate (2+) result of HER2 protein expression by immunohistochemistry. CEP17 amplification concerns the centromeric area and rarely extends beyond it to involve polysomy of the whole chromosome. The association of CEP17 amplification with generalized CIN remains uncertain. Such association, if confirmed, could be an opportunity for a practical and clinically validated test of CIN in breast cancer. This paper explores the association of CIN with centromere 17 amplification and with centromere function in general, as well as the pathophysiology of centromeres/kinetochore function during mitosis that underlies their relationship with CIN in cancer and in breast cancer in particular.

Abstract 3376: Early onset of chromosomal instability in breast preneoplasia detected by single-cell genomics

Abstract Chromosomal rearrangements and copy number aberrations are a major mechanism for driver gene alterations in breast cancer. However, the origins of chromosomal instability during breast tumorigenesis are poorly understood. We have recently shown that oncogene expression in normal mammary epithelial cells induces replication stress and a DNA damage response (DDR). The double-strand break sensor Mre11 is required for the oncogene-induced DDR, and suppression of Mre11 is sufficient to promote the development of Her2/Neu-driven breast cancer in a transgenic mouse model. Here we have applied single-cell genomics to primary mammary epithelial cells from this mouse model and demonstrate a profound effect of oncogene expression on chromosomal instability in preneoplastic cells, which is further augmented in Mre11-mutant cells. Her2/Neu-induced chromosomal breakpoints are highly enriched in the vicinity of large genes, suggesting a possible role for gene transcription and/or R-loops in the origin of oncogene-induced genomic fragility. Mre11 hypomorphism does not affect the number of chromosomal breakpoints but alters the genomic aberration signature to one that is over-represented by larger-size deletions. These findings indicate that chromosomal instability is an early event during Her2/Neu-initiated breast neoplasia. Furthermore, DDR perturbation—in the form of Mre11 hypomorphism—is not required for oncogene-induced genomic instability but rather alters the pattern of observed copy number aberrations and promotes the proliferative expansion of genomically unstable clones. Thus, single-cell genomics applied to murine models of breast preneoplasia reveals insights into the origins of chromosomal instability in breast cancer that may inform novel strategies to prognosticate or prevent the progression of high-risk premalignant breast lesions. Citation Format: Katerina Fagan-Solis, Dennis A. Simpson, Luciano Martelotto, Jorge S. Reis-Filho, John H. Petrini, Gaorav P. Gupta. Early onset of chromosomal instability in breast preneoplasia detected by single-cell genomics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3376.

Expression of regulators of mitotic fidelity are associated with intercellular heterogeneity and chromosomal instability in primary breast cancer.

Regulators of transition through mitosis such as SURVIVIN and Aurora kinase A (AURKA) have been previously implicated in the initiation of chromosomal instability (CIN), a driver of intratumour heterogeneity. We investigate the relationship between protein expression of these genes and directly quantified CIN, and their prognostic utility in breast cancer. The expression of SURVIVIN and AURKA was determined by immunohistochemistry in a cohort of 426 patients with primary breast cancer. The association between protein expression and histopathological characteristics, clinical outcome and CIN status, as determined by centromeric FISH and defined by modal centromere deviation, was analysed. Significantly poorer clinical outcome was observed in patients with high AURKA expression levels. Expression of SURVIVIN was elevated in ER-negative relative to ER-positive breast cancer. Both AURKA and SURVIVIN increased expression were significantly associated with breast cancer grade. There was a significant association between increased CIN and both increased AURKA and SURVIVIN expression. AURKA gene amplification was also associated with increased CIN. To our knowledge this is the largest study assessing CIN status in parallel with the expression of the mitotic regulators AURKA and SURVIVIN. These data suggest that elevated expression of AURKA and SURVIVIN, together with AURKA gene amplification, are associated with increased CIN in breast cancer, and may be used as a proxy for CIN in breast cancer samples in the absence of more advanced molecular measurements.

Patterns and incidence of chromosomal instability and their prognostic relevance in breast cancer subtypes

One of the hallmarks of human solid tumors is chromosomal instability (CIN). We studied global patterns as well as individual levels of CIN and determined the prognostic relevance among breast cancer subtypes. For this, we used single nucleotide polymorphism copy number data of 313 primary lymph-node negative breast cancers. The level of CIN for individual samples was determined by counting the total number of chromosomal segments showing a gain or loss per specimen. Hierarchical clustering resulted in four groups showing distinct patterns of abnormalities, predominantly characterized by 1q gain, 8q gain, 1q&8q gain, or no gain of these loci. Estrogen receptor (ER)-positive and ER-negative samples showed an uneven distribution (statistically significant) across the cluster-groups, as did the molecular subtypes and triple-negative tumors (negative for estrogen-, progesterone-, and her2/neu-receptor). The CIN-score was significantly higher in ER-negative and triple-negative samples. Among luminal cancers, luminal B had a higher CIN-score than luminal A. The CIN-score was significantly associated with prognosis, measured by the time to distant metastasis, in ER-positive, luminal B, and her2/neu subtypes, but not in ER-negative patients. Our study points to a multifaceted role for CIN in breast cancer. Within ER-negative samples, CIN is likely related to the onset but other factors govern the progression of the disease. In contrast, CIN is clearly associated with progression in ER-positive, luminal B, and her2/neu subtypes; thus, assessing CIN in these subtypes may contribute to personalized patient management.

FBXW7 and DNA copy number instability

SKP1-cullin-F-box protein (SCF) type ubiquitin ligases degrade proteins controlling the G1/S transition. Deficiency for FBXW7 (also known as hCDC4), which encodes the F-box protein of the SCF type ubiquitin ligase is associated with genomic instability. Here, we investigated the association of FBXW7 deficiency with chromosomal instability in breast cancer. We screened 49 tumors previously profiled by array CGH for mutations in conserved regions of FBXW7, but found no mutations. Copy number loss of FBXW7, however was associated with enhanced genomic instability in the Complex breast tumor subtype, but instability may not be due to FBXW7 haploinsufficiency, since transcript levels were not reduced in tumors with loss of the locus, whereas reduced expression was observed for other neighboring genes involved in maintenance of genome stability. We also investigated whether cells deficient for FBXW7 showed enhanced instability by challenging cells with methotrexate and assessing numbers of genomic alterations arising in resistant cells. Although methotrexate resistant colonies formed at high frequencies in HCT116 FBXW7+/- and HCT116 FBXW7-/- cells compared to parental HCT116, few copy number alterations were detected in the resistant cells. Taken together these studies suggest that FBXW7 deficiency is unlikely to contribute to the extensive copy number aberrations associated with breast and possibly other tumor types.

Patterns of chromosomal instability in breast cancer

INTRODUCTION: Chromosomal instability may contribute to breast cancer progression and heterogeneity. Gene-expression profiling clustered breast tumors into subclasses IA, predominantly HER2-positive; IB, ER-negative/HER2-negative “basal-like”; IIA, low-grade ER-positive; and IIB, high-grade ER-positive. We hypothesized that chromosomal instability differs among these subclasses, and assessed tumor ploidy and frequency of allelic imbalance.

Centrosome Amplification and the Origin of Chromosomal Instability in Breast Cancer

The development and progression of aggressive breast cancer is characterized by genomic instability leading to multiple genetic defects, phenotypic diversity, chemoresistance, and poor outcome. Centrosome abnormalities have been implicated in the origin of chromosomal instability through the development of multipolar mitotic spindles. Breast tumor centrosomes display characteristic structural abnormalities, termed centrosome amplification , including: increase in centrosome number and volume, accumulation of excess pericentriolar material, supernumerary centrioles, and inappropriate phosphorylation of centrosome proteins. In addition, breast tumor centrosomes also show functional abnormalities characterized by inappropriate centrosome duplication during the cell cycle and nucleation of unusually large microtubule arrays. These observations have important implications for understanding the mechanisms underlying genomic instability and loss of cell polarity in cancer. This review focuses on the coordination of the centrosome, DNA, and cell cycles in normal cells and their deregulation resulting in centrosome amplification and chromosomal instability in the development and progression of breast cancer.

Variable Levels of Chromosomal Instability and Mitotic Spindle Checkpoint Defects in Breast Cancer

Cytogenetic analyses have revealed that many aneuploid breast cancers have cell-to-cell variations of chromosome copy numbers, suggesting that these neoplasms have instability of chromosome numbers. To directly test for possible chromosomal instability in this disease, we used fluorescent in situ hybridization to monitor copy numbers of multiple chromosomes in cultures of replicating breast cancer-derived cell lines and nonmalignant breast epithelial cells. While most (7 of 9) breast cancer cell lines tested are highly unstable with regard to chromosome copy numbers, others (2 of 9 cell lines) have a moderate level of instability that is higher than the "background" level of normal mammary epithelial cells and MCF-10A cells, but significantly less than that seen in the highly unstable breast cancer cell lines. To evaluate the potential role of a defective mitotic spindle checkpoint as a cause of this chromosomal instability, we used flow cytometry to monitor the response of cells to nocodazole-induced mitotic spindle damage. All cell lines with high levels of chromosomal instability have defective mitotic spindle checkpoints, whereas the cell lines with moderate levels of chromosomal instability (and the stable normal mammary cells and MCF10A cells) arrest in G(2) when challenged with nocodazole. Notably, the extent of mitotic spindle checkpoint deficiency and chromosome numerical instability in these cells is unrelated to the presence or absence of p53 mutations. Our results provide direct evidence for chromosomal instability in breast cancer and show that this instability occurs at variable levels among cells from different cancers, perhaps reflecting different functional classes of chromosomal instability. High levels of chromosomal instability are likely related to defective mitotic checkpoints but not to p53 mutations.