Correlations of differentially expressed gap junction connexins cx26, cx30, cx32, cx43 and cx46 with breast cancer progression and prognosis.

BACKGROUND AND AIMS: Connexins and their cell membrane channels contribute to the control of cell proliferation and compartmental functions in breast glands and their deregulation is linked to breast carcinogenesis. Our aim was to correlate connexin expression with tumor progression and prognosis in primary breast cancers. MATERIALS AND METHODS: Meta-analysis of connexin isotype expression data of 1809 and 1899 breast cancers from the Affymetrix and Illumina array platforms, respectively, was performed. Expressed connexins were also monitored at the protein level in tissue microarrays of 127 patients equally representing all tumor grades, using immunofluorescence and multilayer, multichannel digital microscopy. Prognostic correlations were plotted in Kaplan-Meier curves and tested using the log-rank test and cox-regression analysis in univariate and multivariate models. RESULTS: The expression of GJA1/Cx43, GJA3/Cx46 and GJB2/Cx26 and, for the first time, GJA6/Cx30 and GJB1/Cx32 was revealed both in normal human mammary glands and breast carcinomas. Within their subfamilies these connexins can form homo- and heterocellular epithelial channels. In cancer, the array datasets cross-validated each other's prognostic results. In line with the significant correlations found at mRNA level, elevated Cx43 protein levels were linked with significantly improved breast cancer outcome, offering Cx43 protein detection as an independent prognostic marker stronger than vascular invasion or necrosis. As a contrary, elevated Cx30 mRNA and protein levels were associated with a reduced disease outcome offering Cx30 protein detection as an independent prognostic marker outperforming mitotic index and necrosis. Elevated versus low Cx43 protein levels allowed the stratification of grade 2 tumors into good and poor relapse free survival subgroups, respectively. Also, elevated versus low Cx30 levels stratified grade 3 patients into poor and good overall survival subgroups, respectively. CONCLUSION: Differential expression of Cx43 and Cx30 may serve as potential positive and negative prognostic markers, respectively, for a clinically relevant stratification of breast cancers

Prevalence and Predictors of Loss of Wild Type BRCA1 in Estrogen Receptor Positive and Negative BRCA1-Associated Breast Cancers

Introduction: The majority of breast cancers that occur in BRCA1 mutation carriers (BRCA1 carriers) are estrogen receptor-negative (ER-). Therefore, it has been suggested that ER negativity is intrinsic to BRCA1 cancers and reflects the cell of origin of these tumors. However, approximately 20% of breast cancers that develop in BRCA1 carriers are ER-positive (ER+); these cancers are more likely to develop as BRCA1 carriers age, suggesting that they may be incidental and unrelated to BRCA1 deficiency. The purpose of this study was to compare the prevalence of loss of heterozygosity due to loss of wild type (wt) BRCA1 in ER+ and ER- breast cancers that have occurred in BRCA1 carriers and to determine whether age at diagnosis or any pathologic features or biomarkers predict for loss of wt BRCA1 in these breast cancers. Methods: Relative amounts of mutated and wt BRCA1 DNA were measured by quantitative polymerase chain reaction performed on laser capture microdissected cancer cells from 42 ER+ and 35 ER- invasive breast cancers that developed in BRCA1 carriers. BRCA1 gene methylation was determined on all cancers in which sufficient DNA was available. Immunostains for cytokeratins (CK) 5/6, 14, 8 and 18, epidermal growth factor receptor and p53 were performed on paraffin sections from tissue microarrays containing these cancers. Results: Loss of wt BRCA1 was equally frequent in ER+ and ER- BRCA1-associated cancers (81.0% vs 88.6%, respectively; P = 0.53). One of nine cancers tested that retained wt BRCA1 demonstrated BRCA1 gene methylation. Age at diagnosis was not significantly different between first invasive ER+ BRCA1 breast cancers with and without loss of wt BRCA1 (mean age 45.2 years vs 50.1 years, respectively; P = 0.51). ER+ BRCA1 cancers that retained wt BRCA1 were significantly more likely than those that lost wt BRCA1 to have a low mitotic rate (odds ratio (OR), 5.16; 95% CI, 1.91 to ∞). BRCA1 cancers with loss of wt BRCA1 were more likely to express basal cytokeratins CK 5/6 or 14 (OR 4.7; 95% CI, 1.85 to ∞). Conclusions: We found no difference in the prevalence of loss of wt BRCA1 between ER+ and ER- invasive BRCA1-associated breast cancers. Our findings suggest that many of the newer therapies for BRCA1 breast cancers designed to exploit the BRCA1 deficiency in these cancers may also be effective in ER+ cancers that develop in this population

Omecamtiv mecarbil in chronic heart failure with reduced ejection fraction, GALACTIC‐HF: baseline characteristics and comparison with contemporary clinical trials

Aims: The safety and efficacy of the novel selective cardiac myosin activator, omecamtiv mecarbil, in patients with heart failure with reduced ejection fraction (HFrEF) is tested in the Global Approach to Lowering Adverse Cardiac outcomes Through Improving Contractility in Heart Failure (GALACTIC‐HF) trial. Here we describe the baseline characteristics of participants in GALACTIC‐HF and how these compare with other contemporary trials. Methods and Results: Adults with established HFrEF, New York Heart Association functional class (NYHA) ≥ II, EF ≤35%, elevated natriuretic peptides and either current hospitalization for HF or history of hospitalization/ emergency department visit for HF within a year were randomized to either placebo or omecamtiv mecarbil (pharmacokinetic‐guided dosing: 25, 37.5 or 50 mg bid). 8256 patients [male (79%), non‐white (22%), mean age 65 years] were enrolled with a mean EF 27%, ischemic etiology in 54%, NYHA II 53% and III/IV 47%, and median NT‐proBNP 1971 pg/mL. HF therapies at baseline were among the most effectively employed in contemporary HF trials. GALACTIC‐HF randomized patients representative of recent HF registries and trials with substantial numbers of patients also having characteristics understudied in previous trials including more from North America (n = 1386), enrolled as inpatients (n = 2084), systolic blood pressure < 100 mmHg (n = 1127), estimated glomerular filtration rate < 30 mL/min/1.73 m2 (n = 528), and treated with sacubitril‐valsartan at baseline (n = 1594). Conclusions: GALACTIC‐HF enrolled a well‐treated, high‐risk population from both inpatient and outpatient settings, which will provide a definitive evaluation of the efficacy and safety of this novel therapy, as well as informing its potential future implementation

Abstract 1706: Comparison of automated and manual RNA isolation from stabilized peripheral blood samples (PBL) for expression analysis of colorectal cancer patients

Abstract Background and aims: PBL based molecular diagnostics requires automated sample processing and evaluation of the isolated RNA for RT-PCR analysis. Our aim was to compare the automated PBL RNA isolation on MagNA Pure 96 with conventional manual RNA isolation by the PAXgene Blood RNA kit and to determine the purity, recovery, RNA integrity (RIN) and sensitivity of downstream RT-PCR applications. Materials and Methods: PBL samples were taken from 25 CRC and 19 healthy normal patients into PAXgene Blood RNA Tubes (PreAnalytix). Total RNA was manually isolated using the PAXgene Blood RNA Kit (Qiagen) for 2 blood samples of each patient. In parallel, the MagNA Pure 96 Cellular RNA Kit LV and MagNA Pure 96 instrument (MP96) (Roche) were applied for total RNA extraction from the other 2 blood samples of each patient. Total RNA concentration was measured using a NanoDrop spectrophotometer. RIN was determined on a Bioanalyzer 2100 (Agilent) using the RNA6000 Nano Kit. Reverse transcription was carried out from 1ug total RNA using Transcriptor First Strand cDNA Synthesis Kit (Roche). The expression of 6 CRC markers from previous microarray analysis (ASGR2, CLIP1, ARHGAP18, 1569263_at, C19orf43, MALAT1) was determined by quantitative real-time PCR on LightCycler 480 System (Roche). Results: The MP96 isolation resulted in higher RNA yields (MP96: 5.99±2.04 ug/tube; manual: 5.65±2.75 ug/tube). The purity OD260/280 of manually isolated samples (2.09 ± 0.03) was equal to that of MP96 isolated ones (2.01 ± 0.05), all samples fulfilled the requirement. The OD230/260 was significantly better in case of MP96 isolated samples (1.85 ± 0.23) compared to manually isolated ones (1.52 ± 0.57) (p&amp;lt;0.0001). The RNA integrity was acceptable in both isolation method, however the manual protocol provided more intact RNA with slightly higher RNA integrity numbers (RIN manual: 9,19 ± 0,26; RIN MP96: 8,25 ± 0,35; p&amp;lt;0.0001). No significant RIN differences were detected between prospective and archived samples. Comparison of both extraction methods and evaluation by RT-PCR showed a linear regression (r = 0.98). In real-time PCR experiments, ARHGAP18 (spec.:80%, sens.:75%) and ASGR2 (spec.:70%, sens.:75%) were proved to be the best differentiating individual markers. The CTs of the RNA extracted by MP96 were lower than those of the manual extraction (means higher expression) reflecting the higher mRNA yield/ratio of the MP96 method. Conclusion: Automated RNA isolation from stabilized PBL samples on MP96 device provides a high quality, purity and yield of total RNA. The CTs of the RNA extracted by MP96 were better than those of the manual extraction. The MP96 extraction is a fully automated, fast and reliable method with 96 RNA extractions within 85 min, compared to the equally precise, but time consuming manual extraction by the PAXgene Blood RNA kit. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1706. doi:1538-7445.AM2012-1706</jats:p

Prognostic Factors in Pseudomyxoma Peritonei with Emphasis on the Predictive Role of Peritoneal Cancer Index and Tumor Markers

Background: Pseudomyxoma peritonei (PMP) is a rare peritoneal condition where mucus-secreting tumorous cells progressively produce a thick, gelatin-like substance. The prognosis of patients with PMP is determined by the degree of cellularity within the mucin (low-grade (LAMN) vs. high-grade (HAMN) histologic features) and by the extent of the disease. Methods: Prognostic relevance of tumor markers CA19-9 and CEA, gender, Peritoneal Cancer Index (PCI), and completeness of cytoreduction (CC) after cytoreductive surgery were evaluated on 193 consecutive PMP patients, based on a retrospective analysis of prospectively gathered data from a German tertial referral center. Results: We demonstrated that low PCI, CC0 status, low-grade histology, and female gender were independent positive prognostic factors for both overall survival (OS) and progression-free survival (PFS). Furthermore, LAMN patients with achieved CC0 status show significantly better OS and PFS compared to those with CC1 status (p = 0.0353 and p = 0.0026 respectively). In contrast, the duration and drug of hyperthermic intraperitoneal chemotherapy (HIPEC) were not prognostic in any comparison. Increased CA19-9 and CEA levels were significantly associated with HAMN cases, but also predicted recurrence in patients with low-grade histologies. Conclusion: Our study confirmed the prognostic role of tumor markers and emphasized the importance of CC status and PCI in a large cohort of PMP- and LAMN patients

Prognostic Factors in Pseudomyxoma Peritonei with Emphasis on the Predictive Role of Peritoneal Cancer Index and Tumor Markers

Background: Pseudomyxoma peritonei (PMP) is a rare peritoneal condition where mucus-secreting tumorous cells progressively produce a thick, gelatin-like substance. The prognosis of patients with PMP is determined by the degree of cellularity within the mucin (low-grade (LAMN) vs. high-grade (HAMN) histologic features) and by the extent of the disease. Methods: Prognostic relevance of tumor markers CA19-9 and CEA, gender, Peritoneal Cancer Index (PCI), and completeness of cytoreduction (CC) after cytoreductive surgery were evaluated on 193 consecutive PMP patients, based on a retrospective analysis of prospectively gathered data from a German tertial referral center. Results: We demonstrated that low PCI, CC0 status, low-grade histology, and female gender were independent positive prognostic factors for both overall survival (OS) and progression-free survival (PFS). Furthermore, LAMN patients with achieved CC0 status show significantly better OS and PFS compared to those with CC1 status (p = 0.0353 and p = 0.0026 respectively). In contrast, the duration and drug of hyperthermic intraperitoneal chemotherapy (HIPEC) were not prognostic in any comparison. Increased CA19-9 and CEA levels were significantly associated with HAMN cases, but also predicted recurrence in patients with low-grade histologies. Conclusion: Our study confirmed the prognostic role of tumor markers and emphasized the importance of CC status and PCI in a large cohort of PMP- and LAMN patients.</jats:p

Prognostic potential of ERG (ETS-related gene) expression in prostatic adenocarcinoma

PURPOSE: Following patients after prostatectomy can be expensive and stressful, therefore, a novel and reliable approach to improve stratification is needed both at diagnosis of PCa and following its treatment. We evaluate the association of both ERG and claudin-4, claudin-5, and beta-catenin expression in tumor tissues of patients with organ-confined and advanced prostatic adenocarcinomas. METHODS: A total of 30 patients were included in the study. Nine men who underwent radical prostatectomy for organ-confined (pT2N0M0) cancer (OCC), 10 patients with clinically advanced cancer (CAC), and 11 controls with benign prostatic hypertrophy (BPH). Using immunohistochemistry applied to tissue microarrays, each group was evaluated for beta-catenin, claudin-4, claudin-5, and ERG expression. RESULTS: The expression of ERG was higher in the CAC group when compared to OCC and BPH (p = 0.7684, p = 0.0224, respectively). Among these patients, 5 from the CAC (45 %) and 5 from the OCC group (56 %) stained positively for ERG (p = 1.0). The mean staining score for those with ERG+ advanced cancer was greater than that for the ERG+ organ-confined cancer (p = 0.0209). ERG staining correlated with Gleason score (Pearson's correlation: 0.498, p = 0.0051), but not with serum PSA level (Pearson's correlation: 0.404, p = 0.1202). When analyzing outcome data, high ERG expressing tumors have shown a significantly worse overall survival (p = 0.0084). CONCLUSIONS: Our results of presence or absence of claudin-4 and claudin-5 and ERG staining intensities suggest their potential as prognostic factors for prostate cancer