Breast cancer genetics: Unsolved questions and open perspectives in an expanding clinical practice

Breast cancer is the most common cause of cancer death in the United Kingdom, with a lifetime risk of one in nine in women. Only 5-10% of all cancers is thought to be due to strongly penetrant inherited predisposing genes, such as BRCA1 and BRCA2. However, other less penetrant genes, including some autosomal recessive genes, are likely to be of etiological importance in other families. This review addresses the current knowledge of breast cancer susceptibility genes and explores the possibilities for future developments. Features of tumor pathology, prognosis, and the scope for targeted treatments in mutation carriers are discussed, and the management of known carriers and those at increased risk for developing breast cancer are evaluated. Genetic testing for cancer susceptibility may become widely available in the future, and has important ethical and management implications. (C) 2004 Wiley-Liss, Inc.

Evaluation of a multi-functional nanocarrier for targeted breast cancer iNOS gene therapy.

The present study determines whether the novel designer biomimetic vector (DBV) can condense anddeliver the cytotoxic iNOS gene to breast cancer cells to achieve a therapeutic effect. We have previouslyshown the benefits of iNOS for cancer gene therapy but the stumbling block to future development hasbeen the delivery system.The DBV was expressed, purified and complexed with the iNOS gene. The particle size and chargewere determined via dynamic light scattering techniques. The toxicity of the DBV/iNOS nanoparticleswas quantified using the cell toxicity and clonogenic assays. Over expression of iNOS was confirmed viaWestern blotting and Griess test.The DBV delivery system fully condensed the iNOS gene with nanoparticles less than 100 nm. Transfectionwith the DBV/iNOS nanoparticles resulted in a maximum of 62% cell killing and less than 20%clonogenicity. INOS overexpression was confirmed and total nitrite levels were in the range of 18M.We report for the first time that the DBV can successfully deliver iNOS and achieve a therapeuticeffect. There is significant cytotoxicity coupled with evidence of a bystander effect. We concludethat the success of the DBV fusion protein in the delivery of iNOS in vitro is worthy of future in vivo experiments.

The Delta Np63 Proteins Are Key Allies of BRCA1 in the Prevention of Basal-Like Breast Cancer

Little is known about the origin of basal-like breast cancers, an aggressive disease that is highly similar to BRCA1-mutant breast cancers. p63 family proteins that are structurally related to the p53 suppressor protein are known to function in stem cell regulation and stratified epithelia development in multiple tissues, and p63 expression may be a marker of basal-like breast cancers. Here we report that Delta Np63 isoforms of p63 are transcriptional targets for positive regulation by BRCA1. Our analyses of breast cancer tissue microarrays and BRCA1-modulated breast cancer cell lines do not support earlier reports that p63 is a marker of basal-like or BRCA1 mutant cancers. Nevertheless, we found that BRCA1 interacts with the specific p63 isoform Delta Np63 gamma along with transcription factor isoforms AP-2 alpha and AP-2 gamma. BRCA1 required Delta Np63 gamma and AP-2 gamma to localize to an intronic enhancer region within the p63 gene to upregulate transcription of the Delta Np63 isoforms. In mammary stem/progenitor cells, siRNA- mediated knockdown of Delta Np63 expression resulted in genomic instability, increased cell proliferation, loss of DNA damage checkpoint control, and impaired growth control. Together, our findings establish that transcriptional upregulation of Delta Np63 proteins is critical for BRCA1 suppressor function and that defects in BRCA1-Delta Np63 signaling are key events in the pathogenesis of basal-like breast cancer. Cancer Res; 71( 5); 1933-44. (c) 2011 AACR.

IGF status is altered by tamoxifen in patients with breast cancer

Aims-An increased concentration of insulin-like growth factor 1 (IGF-1) is an independent risk factor for premenopausal breast cancer. Tamoxifen is thought initially to reduce concentrations of IGF-1 and increase concentrations of the IGF binding proteins. The aim of this study was to compare concentrations of IGF-1, IGF binding protein 1 (IGF-BP1), and IGF-BP3 in patients with breast cancer (n = 14) with those seen in control subjects (n = 23) and to assess the effect of tamoxifen on IGF status in these patients.

Are caregiving responsibilities associated with non-attendance at breast screening?

Background: Previous research showed that deprived individuals are less likely to attend breast screening and those providing intense amounts of informal care tend to be more deprived than non-caregivers. The aim of this study was to examine the relationship between informal caregiving and uptake of breast screening and to determine if socio-economic gradients in screening attendance were explained by caregiving responsibilities.

Lack of RUNX3 inactivation in columnar cell lesions of breast

Aims:

Overexpression of endoplasmic reticulum protein 29 regulates mesenchymal-epithelial transition and suppresses xenograft tumor growth of invasive breast cancer cells

Endoplasmic reticulum protein 29 (ERp29) is a novel endoplasmic reticulum ( ER) secretion factor that facilitates the transport of secretory proteins in the early secretory pathway. Recently, it was found to be overexpressed in several cancers; however, little is known regarding its function in breast cancer progression. In this study, we show that the expression of ERp29 was reduced with tumor progression in clinical specimens of breast cancer, and that overexpression of ERp29 resulted in G(0)/G(1) arrest and inhibited cell proliferation in MDA-MB-231 cells. Importantly, overexpression of ERp29 in MDA-MB-231 cells led to a phenotypic change and mesenchymal-epithelial transition (MET) characterized by cytoskeletal reorganization with loss of stress fibers, reduction of fibronectin (FN), reactivation of epithelial cell marker E-cadherin and loss of mesenchymal cell marker vimentin. Knockdown of ERp29 by shRNA in MCF-7 cells reduced E-cadherin, but increased vimentin expression. Furthermore, ERp29 overexpression in MDA-MB-231 and SKBr3 cells decreased cell migration/invasion and reduced cell transformation, whereas silencing of ERp29 in MCF-7 cells enhanced cell aggressive behavior. Significantly, expression of ERp29 in MDA-MB-231 cells suppressed tumor formation in nude mice by repressing the cell proliferative index (Ki-67 positivity). Transcriptional profiling analysis showed that ERp29 acts as a central regulator by upregulating a group of genes with tumor suppressive function, for example, E-cadherin (CDH1), cyclin-dependent kinase inhibitor (CDKN2B) and spleen tyrosine kinase (SYK), and by downregulating a group of genes that regulate cell proliferation (eg, FN, epidermal growth factor receptor ( EGFR) and plasminogen activator receptor ( uPAR)). It is noteworthy that ERp29 significantly attenuated the overall ERK cascade, whereas the ratio of p-ERK1 to p-ERK2 was highly increased. Taken together, our results showed that ERp29 is a novel regulator leading to cell growth arrest and cell transition from a proliferative to a quiescent state, and reprogramming molecular portraits to suppress the tumor growth of MDA-MB-231 breast cancer cells. Laboratory Investigation (2009) 89, 1229-1242; doi: 10.1038/labinvest.2009.87; published online 21 September 2009

Positive association between nuclear Runx2 and oestrogen-progesterone receptor gene expression characterises a biological subtype of breast cancer

Purpose: The runt-related transcription factor, Runx2 may have an oncogenic role in mediating metastatic events in breast cancer, but whether Runx2 has a role in the early phases of breast cancer development is not clear. We examined the expression of Runx2 and its relationship with oestrogen receptor (ER) and progesterone receptor (PR) in breast cancer cell lines and tissues.

RUNX3 inactivation by frequent promoter hypermethylation and protein mislocalization constitute an early event in breast cancer progression

Background We had previously established that inactivation of RUNX3 occurs by frequent promoter hypermethylation and protein mislocalization in invasive ductal carcinomas (IDC) of breast. Here, we hypothesize that inactivation of RUNX3 occurring in ductal carcinoma in situ (DCIS) represent early event in breast carcinogenesis. Methods The study cohort of 40 patients included 17 pure DCIS cases and 23 cases of DCIS with associated IDC (DCIS-IDC). The DCIS and IDC components of mixed cases were manually microdissected to permit separate evaluation. All the 63 samples including 17 pure DCIS, 23 samples each of DCIS and IDC of DCIS-IDC cases were analyzed for RUNX3 protein expression using R3-6E9 monoclonal antibody as well as promoter methylation status by methylation specific PCR. Results Compared to matched normal breast samples (4 of 40, 10%), DCIS (35 of 40, 88%) and IDC (21 of 23, 91%) exhibited significant RUNX3 inactivation (P

Novel breast cancer biomarkers identified by integrative proteomic and gene expression mapping

Proteomic and transcriptomic platforms both play important roles in cancer research, with differing strengths and limitations. Here, we describe a proteo-transcriptomic integrative strategy for discovering novel cancer biomarkers, combining the direct visualization of differentially expressed proteins with the high-throughput scale of gene expression profiling. Using breast cancer as a case example, we generated comprehensive two-dimensional electrophoresis (2DE)/mass spectrometry (MS) proteomic maps of cancer (MCF-7 and HCC-38) and control (CCD-1059Sk) cell lines, identifying 1724 expressed protein spots representing 484 different protein species. The differentially expressed cell-line proteins were then mapped to mRNA transcript databases of cancer cell lines and primary breast tumors to identify candidate biomarkers that were concordantly expressed at the gene expression level. Of the top nine selected biomarker candidates, we reidentified ANX1, a protein previously reported to be differentially expressed in breast cancers and normal tissues, and validated three other novel candidates, CRAB, 6PGL, and CAZ2, as differentially expressed proteins by immunohistochemistry on breast tissue microarrays. In total, close to half (4/9) of our protein biomarker candidates were successfully validated. Our study thus illustrates how the systematic integration of proteomic and transcriptomic data from both cell line and primary tissue samples can prove advantageous for accelerating cancer biomarker discovery.

Progressive loss of epidermal growth factor receptor in a subpopulation of breast cancers: implications in target-directed therapeutics

Understanding the molecular etiology and heterogeneity of disease has a direct effect on cancer therapeutics. To identify novel molecular changes associated with breast cancer progression, we conducted phosphoproteomics of the MCF10AT model comprising isogenic, ErbB2- and ErbB3-positive, xenograft-derived cell lines that mimic different stages of breast cancer. Using in vitro animal model and clinical breast samples, our study revealed a marked reduction of epidermal growth factor receptor (EGFR) expression with breast cancer progression. Such diminution of EGFR expression was associated with increased resistance to Gefitinib/Iressa in vitro. Fluorescence in situ hybridization showed that loss of EGFR gene copy number was one of the key mechanisms behind the low/null expression of EGFR in clinical breast tumors. Statistical analysis on the immunohistochemistry data of EGFR expression from 93 matched normal and breast tumor samples showed that (a) diminished EGFR expression could. be detected as early as in the preneoplastic lesion (ductal carcinoma in situ) and this culminated in invasive carcinomas; (b) EGFR expression levels could distinguish between normal tissue versus carcinoma in situ and invasive carcinoma with high statistical significance (P

Targets of genome copy number reduction in primary breast cancers identified by integrative genomics

The identification of specific oncogenes and tumor suppressor genes in regions of recurrent aneuploidy is a major challenge of molecular cancer research. Using both oligonucleotide single-nucleotide polymorphism and mRNA expression arrays, we integrated genomic and transcriptional information to identify and prioritize candidate cancer genes in regions of increased and decreased chromosomal copy number in a cohort of primary breast cancers. Confirming the validity of this approach, several regions of previously-known copy number (CN) alterations in breast cancer could be successfully reidentified. Focusing on regions of decreased CN, we defined a prioritized list of eighteen candidate genes, which included ARPIN, FBNI, and LZTSI, previously shown to be associated with cancers in breast or other tissue types, and novel genes such as P29, MORF4LI, and TBCID5. One such gene, the RUNX3 transcription factor, was selected for further study. We show that RUNX3 is present at reduced CNs in proportion to the rest of the tumor genome and that RUNX3 CN reductions can also be observed in a breast cancer series from a different center. Using tissue microarrays, we demonstrate in an independent cohort of over 120 breast tissues that RUNX3 protein is expressed in normal breast epithelium but not fat and stromal tissue, and widely down-regulated in the majority of breast cancers (> 85%). In vitro, RUNX3 overexpression suppressed the invasive potential of MDA-MB-231 breast cancer cells in a matrigel assay. Our results demonstrate the utility of integrative genomic approaches to identify novel potential cancer-related genes in primary tumors. This article contains Supplementary Material available at http:// www.interscience.wiley.com/jpages/1045-2257/suppmat. (c) 2006 Wiley-Liss, Inc.

Mutational hotspot in Exon 20 of PIK3CA in breast cancer among Singapore Chinese

The recent identification of somatic mutations in the catalytic region of PIK3 (PIK3CA) in breast cancer and demonstration of their oncogenic function has implicated PIK3CA in mammary carcinogenesis. To investigate possible ethnic differences in patterns of PIK3CA mutations in Singaporean Chinese breast cancer and to characterize these in a panel of cell lines, we sequenced exons 9 and 20 in 80 primary tumors, 19 breast cancer cell lines and 7 normal human mammary epithelial cells (HMECs). Searching for novel hotspots of mutation, we sequenced additional exons ( 1, 2, 6, 7, 14 and 18) in 20 primary tumors and 6 breast cancer cell lines. We detected 33 point mutations in 31 of 80 (39%) breast cancers, and 11 mutations in 10 of 19 (53%) breast cancer cell lines. No mutations were detected in normal breast tissue adjacent to the tumor, or in the 6 normal HMECs. The exon 20 A3140G (H1047R) substitution was identified most frequently (22/31, 71%) and showed a significant association with patient age ( p = 0.043) and stage of the disease ( p = 0.025), but not with ER/PR status or histological grade of the tumor. The incidence of point mutations in PIK3CA, the A3140G substitution in particular, in Singapore breast cancers are among the most frequent reported to date for any gene in breast cancer. The results suggest that mutation of PIK3CA might contribute to development of early stage breast cancer and could provide a potent target for early diagnosis and therapy.