The stabilisation of HIF-α is central to the transcriptional response of animals to hypoxia, regulating the expression of hundreds of genes including those involved in angiogenesis, metabolism and metastasis. HIF-α is degraded under normoxic conditions by proline hydroxylation, which allows for recognition and ubiquitination by the von-Hippel-Lindau (VHL) E3 ligase complex. The aim of our study was to investigate the posttranslational modification of HIF-1α in tumours, to assess whether there are additional mechanisms besides reduced hydroxylation leading to stability. To this end we optimised antibodies against the proline-hydroxylated forms of HIF-1α for use in formalin fixed paraffin embedded (FFPE) immunohistochemistry to assess effects in tumour cells in vivo. We found that HIF-1α proline-hydroxylated at both VHL binding sites (Pro402 and Pro564), was present in hypoxic regions of a wide range of tumours, tumour xenografts and in moderately hypoxic cells in vitro. Staining for hydroxylated HIF-1α can identify a subset of breast cancer patients with poorer prognosis and may be a better marker than total HIF-1α levels. The expression of unhydroxylated HIF-1α positively correlates with VHL in breast cancer suggesting that VHL may be rate-limiting for HIF degradation. Our conclusions are that the degradation of proline-hydroxylated HIF-1α may be rate-limited in tumours and therefore provides new insights into mechanisms of HIF upregulation. Persistence of proline-hydroxylated HIF-1α in perinecrotic areas suggests there is adequate oxygen to support prolyl hydroxylase domain (PHD) activity and proline-hydroxylated HIF-1α may be the predominant form associated with the poorer prognosis that higher levels of HIF-1α confer.
Patients with newly diagnosed, early stage estrogen receptor positive (ER+) breast cancer often show disease free survival in excess of five years following surgery and systemic adjuvant therapy. An important question is whether diagnostic tumor tissue from the primary lesion offers an accurate molecular portrait of the cancer post recurrence and thus may be used for predictive diagnostic purposes for patients with relapsed, metastatic disease. As the class I phosphatidylinositol 3' kinase (PI3K) pathway is frequently activated in ER+ breast cancer and has been linked to acquired resistance to hormonal therapy, we hypothesized pathway status could evolve over time and treatment. Biomarker analyses were conducted on matched, asynchronous primary and metastatic tumors from 77 patients with ER+ breast cancer. We examined whether PIK3CA and AKT1 alterations or PTEN and Ki67 levels showed differences between primary and metastatic samples. We also sought to look more broadly at gene expression markers reflective of proliferation, molecular subtype, and key receptors and signaling pathways using an mRNA analysis platform developed on the Fluidigm BioMark™ microfluidics system to measure the relative expression of 90 breast cancer related genes in formalin-fixed paraffin-embedded (FFPE) tissue. Application of this panel of biomarker assays to matched tumor pairs showed a high concordance between primary and metastatic tissue, with generally few changes in mutation status, proliferative markers, or gene expression between matched samples. The collection of assays described here has been optimized for FFPE tissue and may have utility in exploratory analyses to identify patient subsets responsive to targeted therapies.
The brain is a common site of metastatic disease in patients with breast cancer, which has few therapeutic options and dismal outcomes. The purpose of our study was to identify common and rare events that underlie breast cancer brain metastasis. We performed deep genomic profiling, which integrated gene copy number, gene expression and DNA methylation datasets on a collection of breast brain metastases. We identified frequent large chromosomal gains in 1q, 5p, 8q, 11q, and 20q and frequent broad-level deletions involving 8p, 17p, 21p and Xq. Frequently amplified and overexpressed genes included ATAD2, BRAF, DERL1, DNMTRB and NEK2A. The ATM, CRYAB and HSPB2 genes were commonly deleted and underexpressed. Knowledge mining revealed enrichment in cell cycle and G2/M transition pathways, which contained AURKA, AURKB and FOXM1. Using the PAM50 breast cancer intrinsic classifier, Luminal B, Her2+/ER negative, and basal-like tumors were identified as the most commonly represented breast cancer subtypes in our brain metastasis cohort. While overall methylation levels were increased in breast cancer brain metastasis, basal-like brain metastases were associated with significantly lower levels of methylation. Integrating DNA methylation data with gene expression revealed defects in cell migration and adhesion due to hypermethylation and downregulation of PENK, EDN3, and ITGAM. Hypomethylation and upregulation of KRT8 likely affects adhesion and permeability. Genomic and epigenomic profiling of breast brain metastasis has provided insight into the somatic events underlying this disease, which have potential in forming the basis of future therapeutic strategies.
Although the prognosis of patients with small (≤1cm) tumors is generally favorable, emerging data suggests that biological behavior varies between intrinsic subtypes in such patients. Furthermore, it still remains unclear whether HER2-positive pT1a-bN0M0 patients could benefit from adjuvant trastuzumab. For further evaluation, we sought to conduct a meta-analysis so as to get a better understanding of the prognosis for HER2-positive pT1a-bN0M0 patients and their survival benefit from adjuvant trastuzumab, accordingly, offering the implications for current practice.
The PubMed database, the online proceedings of the American Society of Clinical Oncology (ASCO) Annual Meetings, the online proceedings of the San Antonio Breast Cancer Symposium, and the CD proceedings of the International St. Gallen Breast Cancer Conference were searched for all relevant studies published before September 2012. Relative risks (RRs) were used to compare the prognosis of different intrinsic subtypes for pT1a-bN0M0 breast cancer. Analyses were also performed to estimate the association between adjuvant trastuzumab and various survival outcomes.
With eight eligible studies identified, this meta-analysis demonstrated a deleterious effect of HER2+ phenotype on disease-free survival (DFS; RR = 3.677, 95% CI 2.606–5.189, p <0.001) and distant disease-free survival (DDFS; RR = 3.824, 95% CI 2.249–6.501, p<0.001) as compared to HR+/HER2- subgroup. However, significant difference failed to be achieved in terms of any endpoint between HER2+ and triple negative breast cancer (TNBC). Besides, a marked improvement in DFS was observed with the addition of trastuzumab for HER2-positive pT1a-bN0M0 patients (RR = 0.323, 95% CI 0.191–0.547, p<0.001).
This meta-analysis clarifies that intrinsic subtypes might be a reliable marker to predict the prognosis in pT1a-bN0M0 breast cancer. Besides, even for such early stage HER2-positive patients, adjuvant trastuzumab might bring significant survival benefit.
Accumulation of hepatic lipid droplet (HLD) is the hallmark pathology of non-alcoholic fatty liver disease (NAFLD). This study examined the effects of soy isoflavones (ISF) and different amounts of soy proteins on the accumulation of HLD, lipid metabolism and related gene expression in rats. Weanling Sprague–Dawley rats were fed diets containing either 20 % casein protein without (D1) or with (D2) supplemental ISF (50 mg/kg diet) or substitution of casein with increasing amounts of alcohol-washed soy protein isolate (SPI, 5, 10, and 20 %; D3, D4, D5) for 90 days. Dietary casein (20 %) induced accumulation of HLD in female, but not in male rats. Both soy proteins and ISF remarkably prevented the formation of HLD. Soy proteins lowered hepatic total cholesterol and triglyceride in a dose-dependent manner. Interestingly, soy proteins but not ISF significantly increased free fatty acids in the liver of the female rats compared to D1. Proteomic analysis showed that at least 3 enzymes involved in lipogenesis were down-regulated and 7 proteins related to fatty acid β-oxidation or lipolysis were up-regulated by soy protein over D1. Additionally, 9 differentially expressed proteins identified were related to amino acid metabolism, 5 to glycolysis and 2 to cholesterol metabolism. Dietary ISF and SPI markedly reduced hepatic-peroxisome-proliferator-activated receptor γ2 (PPARγ2) and fat-specific protein 27 (FSP27) in female rats. Overall, this study has shown that partial or full replacement of dietary casein by soy protein or supplementation with soy ISF can effectively prevent the accumulation of HLD. The potential molecular mechanism(s) involved might be due to suppression of lipogenesis and stimulation of lipolysis and down-regulation of PPARγ2 and FSP27. This suggests that consumption of soy foods or supplements might be a useful strategy for the prevention or treatment of fatty liver diseases.
Electronic supplementary material
The online version of this article (doi:10.1007/s12263-013-0373-3) contains supplementary material, which is available to authorized users.
Soy proteins; Isoflavones; Fatty liver; Lipid metabolism; Gene expression
Dicer, an RNase III-type endonuclease, is the key enzyme involved in RNA interference and microRNA pathways. Aberrant expression of Dicer is reported in several human cancers. Our aim was to assess the prognostic role of Dicer in breast cancer.
The entire series comprised 666 invasive breast cancers (IBCs), 480 DCIS cases (397 associated with IBC and 83 pure DCIS) and 305 lymph node metastases. Cytoplasmic Dicer expression by immunohistochemistry was scored as negative (no staining) and positive (weak, moderate or strong staining).
Dicer staining was assessable in 446 IBC, 128 DCIS and 101 lymph node metastases. Expression of Dicer was observed in 33% (145/446) of IBCs, 34% (44/128) of DCIS and 57% (58/101) of lymph node metastases. Dicer expression was increased in nodal metastases compared to primary tumours (p<0.001); and was associated with ER negativity (p<0.001), HER2 positivity (p<0.001), high Ki67 labeling index (p<0.001) and expression of basal-like biomarkers (p = 0.002). Dicer positivity was more frequent in the HER2 overexpressing (p<0.001) and basal-like (p = 0.002) subtypes compared to luminal A subtype. Dicer expression was associated with reduced overall survival (OS) on univariate analysis (p = 0.058) and remained an independent predictor of OS on multivariate analysis (HR 2.84, 95% CI 1.43–5.62, p = 0.003), with nodal status (HR 2.61, 95% CI 1.18–5.80, p = 0.018) and PR (HR 0.28, 95% CI 0.13–0.59, p = 0.001). Further, moderate or strong expression of Dicer was associated with improved disease-free survival in the HER2-overexpressing subtype compared to negative or weak expression (p = 0.038).
Deregulated Dicer expression is associated with aggressive tumour characteristics and is an independent prognostic factor for OS. Our findings suggest that Dicer is an important prognostic marker in breast cancer and that its prognostic role may be subtype specific.
Urinary nucleosides are associated with many types of cancer. In this study, six targeted urinary nucleosides, namely adenosine, cytidine, 3-methylcytidine, 1-methyladenosine, inosine, and 2-deoxyguanosine, were chosen to evaluate their role as biomarkers of four different types of cancer: lung cancer, gastric cancer, colon cancer, and breast cancer. Urine samples were purified using solid-phase extraction (SPE) and then analyzed using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The Mann-Whitney U test and Principal Component Analysis (PCA) were used to compare differences in urinary nucleosides between patients with one of four types of cancer and healthy controls. The diagnostic sensitivity of single nucleosides for different types of cancer ranged from 14% to 69%. In contrast, the diagnostic sensitivity of a set of six nucleosides ranged from 37% to 69%. The false-positive identification rate associated with the set of six nucleosides in urine was less than 2% compared with that of less than 5% for a single nucleoside. Furthermore, combining the set of six urinary nucleosides with carcinoembryonic antigen improved the diagnostic sensitivity for colon cancer. In summary, the study show that a set of six targeted nucleosides is a good diagnostic marker for breast and colon cancers but not for lung and gastric cancers.
Recent studies have highlighted the role of androgen receptor (AR) as a prognostic biomarker of breast cancer. However, its predictive role in disease free survival (DFS) and overall survival (OS) still remains inconclusive. The present study aimed to retrospectively investigate the association between AR and survival outcomes in breast cancer and also identify this association by a meta-analysis of published researches. Clinical data from 109 patients with breast cancer, who underwent surgery at Ruijin Hospital, Shanghai, were retrospectively analyzed for immunohistochemical AR expression measured by tissue microarray. For meta-analysis, articles available in Pubmed on the relationship between AR and breast cancer outcomes were included. Data obtained from both were combined and analyzed. Women with AR positive tumors in the retrospective study had a significantly better DFS (HR 0.24, 95% CI 0.07-0.88) and OS (HR 0.19, 95% CI 0.04-0.85) than women with AR negative ones. Meta-analysis showed that AR expression in breast tumors was an indicator of better DFS (HR 0.52, 95% CI 0.43-0.64). In subgroup analysis, AR could predict DFS outcome in estrogen receptor (ER) positive (HR 0.45, 95% CI 0.34-0.59), ER negative (HR 0.42, 95% CI 0.26-0.67), and triple negative breast cancer (HR 0.40, 95% CI 0.23-0.69). Moreover, in ER positive breast cancer patients, the expression of AR could predict better OS (HR 0.39, 95% CI 0.19-0.82). The present analysis indicated that AR expression was associated with lower risk of recurrence in patients with all breast cancer types and better OS in cases with ER positive.
γ-secretase is a large ubiquitously expressed protease complex composed of four core subunits: presenilin, Aph1, PEN-2, and nicastrin. The function of γ-secretase in the cells is to proteolytically cleave various proteins within their transmembrane domains. Presenilin and Aph1 occur as alternative variants belonging to mutually exclusive γ-secretase complexes and providing the complexes with heterogeneous biochemical and physiological properties. γ-secretase is proposed to have a role in the development and progression of cancer and γ-secretase inhibitors are intensively studied for their probable anti-tumor effects in various types of cancer models. Here, we for the first time determined mRNA expression levels of presenilin-1, presenilin-2, Aph1a, Aph1b, PEN-2, and nicastrin in a set of breast cancer tissue samples (N = 55) by quantitative real-time PCR in order to clarify the clinical significance of the expression of different γ-secretase complex components in breast cancer. We found a high positive correlation between the subunit expression levels implying a common regulation of transcription. Our univariate Kaplan-Meier survival analyses established low expression level of γ-secretase complex as a risk factor for breast cancer specific mortality. The tumors expressing low levels of γ-secretase complex were characterized by high histopathological tumor grade, low or no expression of estrogen and progesterone receptors and consequently high probability to fall into the class of triple negative breast cancer tumors. These results may provide novel tools to further categorize breast cancer tumors, especially the highly aggressive and poorly treatable breast cancer type of triple negative cases, and suggest a significant role for γ-secretase in breast cancer.
TGF-β1 is a multi-functional cytokine that plays an important role in breast carcinogenesis. Critical role of TGF-β1 signaling in breast cancer progression is well documented. Some TGF-β1 polymorphisms influence its expression; however, their impact on breast cancer risk is not clear.
We analyzed 1222 samples in a candidate gene-based genetic association study on two distantly located and ethnically divergent case-control groups of Indian women, followed by a population-based genetic epidemiology study analyzing these polymorphisms in other Indian populations. The c.29C>T (Pro10Leu, rs1982073 or rs1800470) and c.74G>C (Arg25Pro, rs1800471) polymorphisms in the TGF-β1 gene were analyzed using direct DNA sequencing, and peripheral level of TGF-β1 were measured by ELISA.
c.29C>T substitution increased breast cancer risk, irrespective of ethnicity and menopausal status. On the other hand, c.74G>C substitution reduced breast cancer risk significantly in the north Indian group (p = 0.0005) and only in the pre-menopausal women. The protective effect of c.74G>C polymorphism may be ethnicity-specific, as no association was seen in south Indian group. The polymorphic status of c.29C>T was comparable among Indo-Europeans, Dravidians, and Tibeto-Burmans. Interestingly, we found that Tibeto-Burmans lack polymorphism at c.74G>C locus as true for the Chinese populations. However, the Brahmins of Nepal (Indo-Europeans) showed polymorphism in 2.08% of alleles. Mean TGF-β1 was significantly elevated in patients in comparison to controls (p<0.001).
c.29C>T and c.74G>C polymorphisms in the TGF-β1 gene significantly affect breast cancer risk, which correlates with elevated TGF-β1 level in the patients. The c.29C>T locus is polymorphic across ethnically different populations, but c.74G>C locus is monomorphic in Tibeto-Burmans and polymorphic in other Indian populations.
Background and Objectives
Many studies have shown a negative association between the consumption of soy products and the risk of some cancers, but little is known about the effect of soy consumption on nasopharyngeal carcinoma. We assessed the association between the consumption of soy products on nasopharyngeal carcinoma risk in Chinese individuals.
This case-control study included 600 (448 males and 152 females) incident cases of nasopharyngeal carcinoma, and an equal number of controls, matched according to gender, age (± 3 y) and household type to the nasopharyngeal carcinoma cases. All subjects were recruited from hospitals in Guangzhou, China. A face-to-face interview was conducted with each study individual to collect general information and habitual dietary intake using a 78-item quantitative food-frequency questionnaire. Odds ratios and their 95% confidence intervals were estimated using conditional logistic regression analyses.
The median intakes of soy foods (in protein) were 0.5/0.5, 1.4/1.7, 2.7/3.3 and 6.1/7.7 (male/female) g/d in the quartiles 1 to 4. Both univariate and multivariate analyses showed no significant association between the consumption of soy proteins or soy isoflavones and the risk of nasopharyngeal carcinoma. The adjusted odds ratios (95% confidence intervals) between extreme quartiles were 0.97 (0.66-1.45) for soy proteins and 0.97 (0.66-1.42) for total isoflavones. Null associations were also observed between intake of the individual isoflavones daidzein, genistein and glycitein and NPC risk, with adjusted odds ratios for the extreme quartiles ranging between 0.73 and 1.23.
Habitual consumption of soy products had no significant effect on the risk of nasopharyngeal carcinoma in Chinese adults with a relatively low intake.
An estimated 1500–3000 invasive Endometrial Stromal Sarcomas (ESS) cases annually occur worldwide. Before 2003, ESS was divided as low and high grade ESS based on mitotic activity. In 2003 the WHO changed the names, excluded mitoses and made nuclear atypia and necrosis the essential diagnostic criteria to distinguish ESS, Low Grade (ESS-LG, recurrence-free survival >90%) and Undifferentiated Endometrial Sarcoma (UES, poor prognosis). We have evaluated in WHO2003 defined ESS-LG whether proliferation biomarkers predict recurrence. Using survival analysis, the prognostic value of classical mitosis counts (Mitotic Activity Index, MAI) in haematoxyllin-eosin (H&E) sections, and immunohistochemical proliferation biomarkers (Ki-67 and PhosphoHistone-3 (PPH3)) were examined in 24 invasive endometrial stromal sarcomas. Three of 24 (12.5%) ESS-LG recurred. The MAI, PPH3 and Ki-67 were all prognostic (P = 0.001, 0.002 and 0.03). MAI values were >3 in the recurrent cases, but never exceeded 10 (the classical threshold for low and high grade). Non-recurrent cases had 0≤MAI≤3. PPH3 and Ki67 counts can be easier to perform than MAI and therefore helpful in the diagnosis of ESS, Low Grade. In conclusion, in this small study of WHO2003 defined ESS-LG, high levels of proliferation as measured by MAI, PPH3 and Ki-67 are predictive of recurrence. Larger studies are required to confirm these results.
Endocrine treatment is the most preferable systemic treatment in metastatic breast cancer patients that have had an estrogen receptor (ER) positive primary tumor or metastatic lesions, however, approximately 20% of these patients do not benefit from the therapy and demonstrate further metastatic progress. One reason for failure of endocrine therapy might be the heterogeneity of ER expression in tumor cells spreading from the primary tumor to distant sites which is reflected in detectable circulating tumor cells (CTCs).
A sensitive and specific staining protocol for ER, keratin 8/18/19, CD45 was established. Peripheral blood from 35 metastatic breast cancer patients with ER-positive primary tumors was tested for the presence of CTCs. Keratin 8/18/19 and DAPI positive but CD45 negative cells were classified as CTCs and evaluated for ER staining. Subsequently, eight individual CTCs from four index patients (2 CTCs per patient) were isolated and underwent whole genome amplification and ESR1 gene mutation analysis.
CTCs were detected in blood of 16 from 35 analyzed patients (46%), with a median of 3 CTCs/7.5 ml. In total, ER-negative CTCs were detected in 11/16 (69%) of the CTC positive cases, including blood samples with only ER-negative CTCs (19%) and samples with both ER-positive and ER-negative CTCs (50%). No correlation was found between the intensity and/or percentage of ER staining in the primary tumor with the number and ER status of CTCs of the same patient. ESR1 gene mutations were not found.
CTCs frequently lack ER expression in metastatic breast cancer patients with ER-positive primary tumors and show a considerable intra-patient heterogeneity, which may reflect a mechanism to escape endocrine therapy. Provided single cell analysis did not support a role of ESR1 mutations in this process.
Acrylamide, a possible human carcinogen, is formed in certain carbohydrate-rich foods processed at high temperature. We evaluated if dietary acrylamide, at doses (0.5, 1.0 or 2.0 mg/kg diet) reflecting upper levels found in human foods, modulated colon tumorigenesis in two rodent models. Male F344 rats were randomized to receive diets without (control) or with acrylamide. 2-weeks later, rats in each group received two weekly subcutaneous injections of either azoxymethane (AOM) or saline, and were killed 20 weeks post-injections; colons were assessed for tumors. Male athymic nude (nu/nu) mice bearing HT-29 human colon adenocarcinoma cells-derived tumor xenografts received diets without (control) or with acrylamide; tumor growth was monitored and mice were killed 4 weeks later. In the F344 rat study, no tumors were found in the colons of the saline-injected rats. However, the colon tumor incidence was 54.2% and 66.7% in the control and the 2 mg/kg acrylamide-treated AOM-injected groups, respectively. While tumor multiplicity was similar across all diet groups, tumor size and burden were higher in the 2 mg/kg acrylamide group compared to the AOM control. These results suggest that acrylamide by itself is not a “complete carcinogen”, but acts as a “co-carcinogen” by exacerbating the effects of AOM. The nude mouse study indicated no differences in the growth of human colon tumor xenografts between acrylamide-treated and control mice, suggesting that acrylamide does not aid in the progression of established tumors. Hence, food-borne acrylamide at levels comparable to those found in human foods is neither an independent carcinogen nor a tumor promoter in the colon. However, our results characterize a potential hazard of acrylamide as a colon co-carcinogen in association with known and possibly other environmental tumor initiators/promoters.
Wnt-5a protein expression in primary tumors from unselected breast cancer patients has revealed a tumor suppressive function of the protein. However, in vitro experiments on human breast cancer cells have reported contradictory results, indicating both a tumor suppressive and promoting functions of Wnt-5a. This could be due to various functions of Wnt-5a in different subgroups of patients. The unselected cohorts analyzed to date for Wnt-5a protein expression contained few premenopausal patients. The aim of the present investigation was to evaluate the prognostic significance of Wnt-5a protein expression in a cohort of premenopausal women with comprehensive data on biomarkers, molecular subtypes and long-term outcome. In a randomized trial of adjuvant tamoxifen versus no adjuvant treatment, 564 premenopausal primary breast cancer patients were included. The median follow-up time was 14 years. A tumor tissue array was constructed and 361 samples were evaluated for Wnt-5a reactivity by immunohistochemistry. The primary end-point was recurrence-free survival. Wnt-5a protein expression was reduced or lost in 146/361 of tumors and correlated to younger age, estrogen receptor (ER) negativity and triple-negative phenotype. Wnt-5a was a prognostic factor in the whole cohort (p = 0.003). In patients with ER-positive tumors, Wnt-5a was an independent positive prognostic marker (HR 0.51 95% CI: 0.33–0.78 p = 0.002) and HER2 a negative prognostic marker (HR 2.84 95% CI: 1.51–5.31, p = 0.001) in a Cox multivariate analysis adjusted for standard prognostic markers and tamoxifen treatment. In the ER-negative subset, Wnt-5a added no prognostic information. In a subgroup analysis, Wnt-5a was significantly associated with better prognosis in patients with Luminal A tumors (p = 0.04). Conclusively, our results suggest that loss of Wnt-5a is a valuable prognostic marker in premenopausal breast cancer patients in particular in patients with ER-positive tumors and out-performed conventional prognostic factors in this subset of patients.
African American (AA) women are more likely than European American (EA) women to be diagnosed with early, aggressive breast cancer. Possible differences in innate immune pathways (e.g., inflammatory responses) have received little attention as potential mechanisms underlying this disparity. We evaluated distributions of selected genetic variants in innate immune pathways in AA and EA women, and examined their associations with breast cancer risk within the Women’s Circle of Health Study (WCHS). In stage I of the study (864 AA and 650 EA women) we found that genotype frequencies for 35 of 42 tested SNPs (18 candidate genes) differed between AAs and EAs (corroborated by ancestry informative markers). Among premenopausal AA women, comparing variant allele carriers to non-carriers, reduced breast cancer risk was associated with CXCL5-rs425535 (OR=0.61, P=0.02), while among EA women, there were associations with TNFA-rs1799724 (OR =2.31, P =0.002) and CRP-rs1205 (OR=0.54, P=0.01). For postmenopausal women, IL1B-rs1143627 (OR=1.80, P=0.02) and IL1B-rs16944 (OR=1.85, P =0.02) were associated with risk among EA women, with significant associations for TNFA-rs1799724 limited to estrogen receptor (ER) positive cancers (OR=2.0, P =0.001). However, none of the SNPs retained significance after Bonferroni adjustment for multiple testing at the level of P0.0012 (0.05/42) except for TNFA-rs1799724 in ER positive cancers. In a stage II validation (1,365 AA and 1,307 EA women), we extended evaluations for four SNPs (CCL2-rs4586, CRP-rs1205, CXCL5-rs425535, and IL1RN-rs4251961), which yielded similar results. In summary, distributions of variants in genes involved in innate immune pathways were found to differ between AA and EA populations, and showed differential associations with breast cancer according to menopausal or ER status. These results suggest that immune adaptations suited to ancestral environments may differentially influence breast cancer risk among EA and AA women.
XRCC3 and RAD51 are two important members in homologous recombination repair pathway. This study was performed to detect the expressions of these two molecules in breast cancer and explore their correlations with clinicopathological factors.
Methods and Results
Immunohistochemistry was used to detect protein expressions of XRCC3 and RAD51 in 248 cases of breast cancer tissue and 78 cases of adjacent non-cancerous tissue. Data showed that expressions for both XRCC3 and RAD51 were significantly increased in breast cancer. High XRCC3 expression was associated with large tumor size and positive PR and HER2 status, while high RAD51 expression was associated with axillary lymph node metastasis and positive PR and HER2 status. The result of multivariate analysis demonstrated that HER2, PR and RAD51 were significantly association with XRCC3. And besides XRCC3, axillary lymph node metastasis and PR were significantly correlated with RAD51.
XRCC3 and RAD51 were significantly associated with clinicopathological factors and they might play important roles in the development and progress of breast cancer.
Inherited factors predisposing individuals to breast and ovarian cancer are largely unidentified in a majority of families with hereditary breast and ovarian cancer (HBOC). We aimed to identify germline copy number variations (CNVs) contributing to HBOC susceptibility in the Finnish population.
A cohort of 84 HBOC individuals (negative for BRCA1/2-founder mutations and pre-screened for the most common breast cancer genes) and 36 healthy controls were analysed with a genome-wide SNP array. CNV-affecting genes were further studied by Gene Ontology term enrichment, pathway analyses, and database searches to reveal genes with potential for breast and ovarian cancer predisposition. CNVs that were considered to be important were validated and genotyped in 20 additional HBOC individuals (6 CNVs) and in additional healthy controls (5 CNVs) by qPCR.
An intronic deletion in the EPHA3 receptor tyrosine kinase was enriched in HBOC individuals (12 of 101, 11.9%) compared with controls (27 of 432, 6.3%) (OR = 1.96; P = 0.055). EPHA3 was identified in several enriched molecular functions including receptor activity. Both a novel intronic deletion in the CSMD1 tumor suppressor gene and a homozygous intergenic deletion at 5q15 were identified in 1 of 101 (1.0%) HBOC individuals but were very rare (1 of 436, 0.2% and 1 of 899, 0.1%, respectively) in healthy controls suggesting that these variants confer disease susceptibility.
This study reveals new information regarding the germline CNVs that likely contribute to HBOC susceptibility in Finland. This information may be used to facilitate the genetic counselling of HBOC individuals but the preliminary results warrant additional studies of a larger study group.
The forkhead box transcription factor Foxo3a has been implicated to play a critical role in various cancers by suppressing tumor growth. Recent studies have identified Foxo3a as a key regulator of Estrogen Receptor-α (ERα). In the present study, we examined the expression of Foxo3a, and investigated its clinical significance and correlation with ER and prognostic role in patients with breast cancer. Immunohistochemical analysis was performed on tumors from 70 breast cancer patients. Interpretable Foxo3a expression was analyzed along with major clinicopathologic variables, and a comparison was made with corresponding 5-year clinical follow-up data. Foxo3a protein expression correlated with ER positivity (P<0.001), histologic grade (1, 2) (P = 0.002), axillary lymph node negativity (P<0.001) and TNM stage (1, 2) (P<0.001). Moreover, the Kaplan-Meier survival curves of the study population showed that a high expression level of Foxo3a was significantly correlated with long-term survival (P<0.0001). In a multivariate analysis, Foxo3a expression was identified as a favorable independent prognostic factor in overall survival (P = 0.038). In conclusion, our results indicated that Foxo3a expression is a favorable prognostic marker in breast cancer. In addition, Foxo3a staining could potentially be used in patient stratification in conjunction with other prognostic markers.
A few studies have evaluated the association between diet and mammographic breast density (MBD) and results are inconsistent. MBD, a well-recognized risk factor for breast cancer, has been proposed as a marker of cumulative exposure to hormones and growth factors. Diets with a high glycemic index (GI) or glycemic load (GL) may increase breast cancer risk, via an effect on the insulin-like growth factor axis. We have investigated the association between carbohydrate intake, GI, GL and MBD in a prospective study. We identified a large series of women, in the frame of the EPIC-Florence cohort, with a mammogram taken five years after enrolment, when detailed information on dietary and lifestyle habits and anthropometric measurements had been collected. Mammograms have been retrieved (1,668, 83%) and MBD assessed according to Wolfe’s classification. We compared women with high MBD (P2+DY Wolfe’s categories) with those with low MBD (N1+P1) through logistic models adjusted for age, education, body mass index, menopause, number of children, breast feeding, physical activity, non-alcohol energy, fibers, saturated fat and alcohol. A direct association between GL and high MBD emerged in the highest quintile of intake in comparison with the lowest quintile (OR = 1.73, 95%CI 1.13–2.67, p for trend = 0.048) while no association with glycemic index was evident. These results were confirmed after exclusion of women reporting to be on a diet or affected with diabetes, and when Hormone Replacement Therapy at the date of mammographic examination used to assess MBD was considered. The effect was particularly evident among leaner women, although no interaction was found. A positive association was suggested for increasing simple sugar and total carbohydrates intakes limited to the highest quintiles. In this Italian population we observed an association between glycemic load, total and rapidly absorbed carbohydrates and high MBD. These novel results warrant further investigations.
Recently Echinoderm microtubule-associated protein-like 4- anaplastic lymphoma kinase (EML4-ALK) fusion gene has become an important biomarker for ALK tyrosine kinase inhibitor (crizotinib) treatment in NSCLC. However, the best detection method and the significance of EML4-ALK variant types remain uncertain.
Reverse transcriptase-polymerase chain reaction (RT-PCR), fluorescence in Situ hybridization (FISH) and Immunohistochemical (IHC) stain were performed on tumor tissues of 312 NSCLC patients for detection of ALK rearrangements. Mutation analyses for EGFR and KRAS genes were also performed.
Thirteen of the 312 patients (4.17%) had ALK rearrangements detected by RT-PCR. If RT-PCR data was used as the gold standard, FISH tests had a low sensitivity (58.33%), but very good specificity (99.32%). IHC stain had better sensitivity (91.67%) than FISH, but lower specificity (79.52%), when the cut off was IHC2+. All of the 8 patients with high abundance of EML4-ALK positive cells in tumor tissues (assessed by the signal intensities of the RT-PCR product), were also have high expression of ALK protein (IHC3+), and positive for FISH, except one failed in FISH. Variants 3a+3b (4/5, 80%) of EML4-ALK fusion gene were more common to have high abundance of EML4-ALK positive cells in tumor tissues than variant 1 (1/3, 33.3%). Meta-analysis of the published data of 2273 NSCLC patients revealed that variant 3 (23/44, 52.3%) was the most common type in Chinese population, while variant 1 (28/37, 75.7%) was most common in Caucasian.
Among the three detection methods, RT-PCR could detect not only the presence of EML4-ALK fusion gene and their variant types, but also the abundance of EML4-ALK positive cells in NSCLC tumor tissues. The latter two factors might affect the treatment response to anti-ALK inhibitor. Including RT-PCR as a diagnostic test for ALK inhibitor treatment in the prospective clinical trials is recommended.
Many studies have reported the prognostic predictive value of CD166 as a cancer stem cell marker in cancers of the digestive system; however, its predictive value remains controversial. Here, we investigate the correlation between CD166 positivity in digestive system cancers and clinicopathological features using meta-analysis.
A comprehensive search in PubMed and ISI Web of Science through March of 2013 was performed. Only articles containing CD166 antigen immunohistochemical staining in cancers of the digestive system were included,including pancreatic cancer, esophageal cancer, gastric cancer and colorectal cancer. Data comparing 3- and 5-year overall survival along with other clinicopathological features were collected.
Nine studies with 2553 patients who met the inclusion criteria were included for the analysis. The median rate of CD166 immunohistochemical staining expression was 56% (25.4%–76.3%). In colorectal cancer specifically, the results of a fixed-effects model indicated that CD166-positive expression was an independent marker associated with a smaller tumor burden (T category; RR = 0.93, 95%, CI: 0.88–0.98) but worse spread to nearby lymph nodes (N category; RR = 1.17, 95% CI: 1.05–1.30). The 5-year overall survival rate was showed relationship with cytoplasmic positive staining of CD166 (RR = 1.47 95% 1.21–1.79), but no significant association was found in the pool or any other stratified analysis with 3- or 5- year overall survival rate.
Based on the published studies, different cellular location of CD166 has distinct prognostic value and cytoplasmic positive expression is associated with worse prognosis outcome. Besides, our results also find CD166 expression indicate advanced T category and N-positive status in colorectal cancer specifically.
Human arylamine N-acetyltransferase 1 (hNAT1) has become an attractive potential biomarker for estrogen-receptor-positive breast cancers. We describe here the mechanism of action of a selective non-covalent colorimetric biosensor for the recognition of hNAT1 and its murine homologue, mNat2, over their respective isoenzymes, leading to new opportunities in diagnosis. On interaction with the enzyme, the naphthoquinone probe undergoes an instantaneous and striking visible color change from red to blue. Spectroscopic, chemical, molecular modelling and biochemical studies reported here show that the color change is mediated by selective recognition between the conjugate base of the sulfonamide group within the probe and the conjugate acid of the arginine residue within the active site of both hNAT1 and mNat2. This represents a new mechanism for selective biomarker sensing and may be exploited as a general approach to the specific detection of biomarkers in disease.
Epithelial-mesenchymal transition (EMT) plays a crucial role in the progression and aggressiveness of colorectal carcinoma. E-cadherin is the best-characterized molecular marker of EMT, but its prognostic significance for patients with CRC remains inconclusive.
Eligible studies were searched from the PubMed, Embase and Web of Science databases. Correlation between E-cadherin expression and clinicopathological features and prognosis was analyzed. Subgroup analysis was also performed according to study location, number of patients, quality score of studies and cut-off value.
A total of 27 studies comprising 4244 cases met the inclusion criteria. Meta-analysis suggested that downregulated E-cadherin expression had an unfavorable impact on overall survival (OS) of CRC (n = 2730 in 14 studies; HR = 2.27, 95%CI: 1.63–3.17; Z = 4.83; P = 0.000). Subgroup analysis indicated that low E-cadherin expression was significantly associated with worse OS in Asian patients (n = 1054 in 9 studies; HR = 2.86, 95%CI: 2.13–3.7, Z = 7.11; P = 0.000) but not in European patients (n = 1552 in 4 studies; HR = 1.14, 95%CI: 0.95–1.35, Z = 1.39; P = 0.165). In addition, reduced E-cadherin expression indicated an unfavorable OS only when the cut off value of low E-cadherin expression was >50% (n = 512 in 4 studies; HR = 2.08, 95%CI 1.45–2.94, Z = 4.05; P = 0.000). Downregulated E-cadherin expression was greatly related with differentiation grade, Dukes' stages, lymphnode status and metastasis. The pooled OR was 0.36(95%CI: 0.19–0.7, Z = 3.03, P = 0.002), 0.34(95%CI: 0.21–0.55, Z = 6.61, P = 0.000), 0.49(95%CI: 0.32–0.74, Z = 3.02, P = 0.002) and 0.45(95%CI: 0.22–0.91, Z = 3.43, P = 0.001), respectively.
This study showed that low or absent E-cadherin expression detected by immunohistochemistry served as a valuable prognostic factor of CRC. However, downregulated E-cadherin expression seemed to be associated with worse prognosis in Asian CRC patients but not in European CRC patients. Additionally, this meta-analysis suggested that the negative threshold of E-cadherin should be >50% when we detected its expression in the immunohistochemistry stain.