Identification of gene networks associated with acute myeloid leukemia by comparative molecular methylation and expression profiling

Around 80% of acute myeloid leukemia (AML) patients achieve a complete remission, however many will relapse and ultimately die of their disease. The association between karyotype and prognosis has been studied extensively and identified patient cohorts as having favourable [e.g. t(8; 21), inv (16)/t(16; 16), t(15; 17)], intermediate [e.g. cytogenetically normal (NK-AML)] or adverse risk [e.g. complex karyotypes]. Previous studies have shown that gene expression profiling signatures can classify the sub-types of AML, although few reports have shown a similar feature by using methylation markers. The global methylation patterns in 19 diagnostic AML samples were investigated using the Methylated CpG Island Amplification Microarray (MCAM) method and CpG island microarrays containing 12,000 CpG sites. The first analysis, comparing favourable and intermediate cytogenetic risk groups, revealed significantly differentially methylated CpG sites (594 CpG islands) between the two subgroups. Mutations in the NPM1 gene occur at a high frequency (40%) within the NK-AML subgroup and are associated with a more favourable prognosis in these patients. A second analysis comparing the NPM1 mutant and wild-type research study subjects again identified distinct methylation profiles between these two subgroups. Network and pathway analysis revealed possible molecular mechanisms associated with the different risk and/or mutation sub-groups. This may result in a better classification of the risk groups, improved monitoring targets, or the identification of novel molecular therapies.

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.

Helix pomatia agglutinin lectin-binding oligosaccharides of aggressive breast cancer

Predicting long-term outcome after breast-cancer diagnosis remains problematic, particularly for patients with clinically small, axillary lymph node- negative tumours, Evidence suggests that the lectin Helix pomatia agglutinin (HPA) identifies oligosaccharides associated with poor-prognosis cancer. Our aim was to identify oligosaccharides that bind HPA in aggressive breast cancers. Breast-cancer cell lines (MCF-7, BT-549 and BT-20) and a cell line From human milk (HBL-100), which showed a range of HPA-binding intensities, were used to extract HPA-binding glycoproteins, Oligosaccharides were released using anhydrous hydrazine and separated on a range of HPLC matrices. We investigated whether HPA-binding oligosaccharides from cell lines were present in human breast-cancer tissues, using 69 breast-cancer specimens from patients with between 5 and 10 years' follow-up. A monosialylated oligosaccharide was over-expressed in the cell line that bound HPA strongly. Further analysis by normal-phase HPLC showed that the 2-aminobenzamide-conjugated oligosaccharide had a hydrodynamic volume of 4.58 glucose units (HPAgly 1), Increased expression of HPAgly 1 was associated with HPA staining of breast-cancer specimens (Student's t-test p = 0.025). Analysis of oligosaccharide levels and disease-free survival after treatment for breast cancer indicated a shorter disease-free interval for patients with elevated levels of HPAgly 1, This is the first time that histochemical lectin staining has been correlated with biochemical mapping of oligosaccharides, Using this approach, we have identified a monosialylated HPA lectin-binding oligosaccharide present in breast-cancer cells grown in vitro which is elevated in breast-cancer specimens that bind the lectin, (C) 2001 Wiley-Liss, Inc.

Expression of osteopontin coregulators in primary colorectal cancer and associated liver metastases

Background: A transcription regulatory complex (TRC) that includes Ets1, Ets2, PEA3 and ß-catenin/T-cell factors regulates osteopontin (OPN) that is implicated in colorectal cancer (CRC) dissemination. The consistency of OPN transcriptional control between primary CRC and metastases is unclear. This study investigates expression and prognostic significance of the OPN–TRC in primary human CRC and associated colorectal liver metastases (CRLM).

Methods: Osteopontin–TRC factors were assayed by digital microscopy in 38 primary CRCs and matched CRLM specimens and assessed against clinical prognosis.

Results: In primary CRC, OPN expression intensity correlated with that of its co-activators, PEA3 (r=0.600; P<0.01), Ets1 (r=0.552; P<0.01), Ets2 (r=0.521; P<0.01) and had prognostic significance. Osteopontin intensity in primary CRC inversely correlated with the interval between diagnosis and resection of CRLM. Overall OPN intensity was lower in CRLM than primary CRC and correlations with co-activators were weaker, for example, Ets1 (P=0.047), PEA3 (P=0.022) or nonsignificant (Ets2). The ratio of OPN expression in CRLM vs primary CRC had prognostic significance.

Conclusion: This study supports transcriptional control of OPN by known coregulators in both primary and secondary CRC. Weaker associations in CRLM suggest involvement of other unknown factors possibly from the liver microenvironment or resulting from additional genetic or epigenetic changes that drive tumour metastatic capability in OPN transcriptional control.

Cofilin immunolabelling correlates with depth of invasion in gastrointestinal endocrine cell tumors

Gastrointestinal endocrine cell tumors are a heterogeneous population of lesions believed to arise from neuroendocrine cells of the gastrointestinal mucosa. The current classification of these tumors is based on tumor size, microscopic features and clinical evidence of metastasis. Although diagnostic categories generally correlate with prognosis, molecular prognostic markers will be clinically useful adjuncts. Cofilin has been implicated in tumor invasion, and its immunolocalisation was studied in gastrointestinal endocrine cell tumors. The immunolocalisation of cofilin was studied by immunohistochemistry in 34 formalin-fixed, paraffin wax-embedded gastrointestinal endocrine cell tumors using a tissue microarray platform. A significant correlation was found between high cofilin immunolabelling and the depth of invasion (p

Potential roles for the PIM1 kinase in human cancer - A molecular and therapeutic appraisal

In vitro experiments have shown the PIM1 kinase to have diverse biological roles in cell survival, proliferation and differentiation. In humans, PIM1 is often expressed in both normal and transformed cells. The PIM1 kinase is a true oncogene implicated in early transformation and tumour progression in haematopoietic malignancies and prostate carcinomas. it is associated with aggressive subgroups of lymphoma, is a marker of poor prognosis in prostate carcinomas and has been suggested to have a role in hormone insensitivity of prostate malignancies. PIM1 has a possible role in other carcinomas with 6p21 genomic alterations. On one hand, PIM1 (due to its role in malignancy) appears to be a promising target for drug development programmes but, on the other hand, the complexity of its molecular structure has posed challenges in the development of PIM1 inhibitors. In this review we discuss PIM1 expression in human tissues (including some new data from our laboratory), its role in human malignancies, as well as the possibilities and challenges in the development of target therapy for PIM1. (c) 2008 Elsevier Ltd. All rights reserved.

Prognostic significance of thymidylate synthase, dihydropyrimidine dehydrogenase and thymidine phosphorylase protein expression in colorectal cancer patients treated with or without 5-fluorouracil-based chemotherapy

Background: Low tumour expression levels of thymidylate synthase (TS), dihydropyrimidine dehydrogenase (DPD) and thymidine phosphorylase (TP) have been linked with improved outcome for colorectal cancer (CRC) patients treated with 5-fluorouracil (5-FU). It is unclear whether this occurs because such tumours have better prognosis or they are more sensitive to 5-FU treatment.

Tissue microarrays characterise the clinical significance of a VEGF-A protein expression signature in gastrointestinal stromal tumours

A tissue microarray analysis of 22 proteins in gastrointestinal stromal tumours ( GIST), followed by an unsupervised, hierarchical monothetic cluster statistical analysis of the results, allowed us to detect a vascular endothelial growth factor ( VEGF) protein overexpression signature discriminator of prognosis in GIST, and discover novel VEGF-A DNA variants that may have functional significance.

Sequence confirmation of the EWS-WT1 fusion gene transcript in the peritoneal effusion of a patient with desmoplastic small round cell tumor

Desmoplastic small round cell tumor (DSRCT) is a rare undifferentiated neoplasm. The prognosis is poor, even if therapy is instituted promptly. and thus it is important to differentiate it from other histologically and cytologically similar-looking malignancies of the young adult. We present a case of DSRCT in a 17-yr-old male with disseminated peritoneal disease and peritoneal effusion. The cytology sample showed a malignant small round cell tumor, the classical cytological features of DSRCT, and immunohistochemistry performed in the prepared cell block exhibited an antibody expression profile in keeping with DSRCT. Further material front the effusion was prepared for RNA extraction, following which a reverse-transcriptase polymerase chain reaction (RTPCR) and sequencing of the t(l l;22)(p13;q11 or q12) were carried out. The result showed the presence of the reciprocal translocation and thus confirmed the diagnosis of DSRCT. This case shows how molecular techniques (including sequencing) call be applied to cytology in clarifying and confirming certain difficult diagnosis of undifferentiated neoplasms, DSRCT in this particular case. Diagn. Cytopathol. 2003;29:341-343. (C) 2003 Wiley-Liss. Inc.

Dietary intake of fruits and vegetables improves microvascular function in hypertensive subjects in a dose-dependent manner

Background— Observational evidence has consistently linked increased fruit and vegetable consumption with reduced cardiovascular morbidity; however, there is little direct trial evidence to support the concept that fruit and vegetable consumption improves vascular function. This study assessed the dose-dependent effects of a fruit and vegetable intervention on arterial health in subjects with hypertension.

Methods and Results— After a 4-week run-in period during which fruit and vegetable intake was limited to 1 portion per day, participants were randomized to consume either 1, 3, or 6 portions daily for the next 8 weeks. Endothelium-dependent and -independent arterial vasodilator responses were assessed by venous occlusion plethysmography in the brachial circulation before and after intervention. Compliance was monitored with serial contemporaneous 4-day food records and by measuring concentrations of circulating dietary biomarkers. A total of 117 volunteers completed the 12-week study. Participants in the 1-, 3-, and 6-portions/d groups reported consuming on average 1.1, 3.2, and 5.6 portions of fruit and vegetables, respectively, and serum concentrations of lutein and ß-cryptoxanthin increased across the groups in a dose-dependent manner. For each 1-portion increase in reported fruit and vegetable consumption, there was a 6.2% improvement in forearm blood flow responses to intra-arterial administration of the endothelium-dependent vasodilator acetylcholine (P=0.03). There was no association between increased fruit and vegetable consumption and vasodilator responses to sodium nitroprusside, an endothelium-independent vasodilator.

Conclusions— The present study illustrates that among hypertensive volunteers, increased fruit and vegetable consumption produces significant improvements in an established marker of endothelial function and cardiovascular prognosis.

Systematic review of reviews of risk factors for intracranial aneurysms

Systematic reviews of systematic reviews identify good quality reviews of earlier studies of medical conditions. This article describes a systematic review of systematic reviews performed to investigate factors that might influence the risk of rupture of an intracranial aneurysm. It exemplifies the technique of this type of research and reports the finding of a specific study. The annual incidence of subarachnoid haemorrhage resulting from the rupture of intracranial aneurysms is estimated to be nine per 100,000. A large proportion of people who have this bleed, will die or remain dependent on the care of others for some time. Reliable knowledge about the risks of subarachnoid haemorrhage in different populations will help in planning, screening and prevention strategies and in predicting the prognosis of individual patients. If the necessary data were available in the identified reviews, an estimate for the numerical relationship between a particular characteristic and the risk of subarachnoid haemorrhage was included in this report. The identification of eligible systematic reviews relied mainly on the two major bibliographic databases of the biomedical literature: PubMed and EMBASE. These were searched in 2006, using specially designed search strategies. Approximately 2,000 records were retrieved and each of these was checked carefully against the eligibility criteria for this systematic review. These criteria required that the report be a systematic review of studies assessing the risk of subarachnoid haemorrhage in patients known to have an unruptured intracranial aneurysm or of studies that had investigated the characteristics of people who experienced a subarachnoid haemorrhage without previously being known to have an unruptured aneurysm. Reports which included more than one systematic review were eligible and each of these reviews was potentially eligible. The quality of each systematic review was assessed. In this review, 16 separate reports were identified, including a total of 46 eligible systematic reviews. These brought together research studies for 24 different risk factors. This has shown that the following factors appear to be associated with a higher risk of subarachnoid haemorrhage: being a woman, older age, posterior circulation aneurysms, larger aneurysms, previous symptoms,

A concept analysis of renal supportive care: the changing world of nephrology

Title. A concept analysis of renal supportive care: the changing world of nephrology

Aim. This paper is a report of a concept analysis of renal supportive care.

Background. Approximately 1.5 million people worldwide are kept alive by renal dialysis. As services are required to support patients who decide not to start or to withdraw from dialysis, the term renal supportive care is emerging. Being similar to the terms palliative care, end-of-life care, terminal care and conservative management, there is a need for conceptual clarity.

Method. Rodgers' evolutionary method was used as the organizing framework for this concept analysis. Data were collected from a review of CINAHL, Medline, PsycINFO, British Nursing Index, International Bibliography of the Social Sciences and ASSIA (1806-2006) using, 'renal' and 'supportive care' as keywords. All articles with an abstract were considered. The World Wide Web was also searched in English utilizing the phrase 'renal supportive care'.

Results. Five attributes of renal supportive care were identified: available from diagnosis to death with an emphasis on honesty regarding prognosis and impact of disease; interdisciplinary approach to care; restorative care; family and carer support and effective, lucid communication to ensure informed choice and clear lines of decision-making.

Conclusion. Renal supportive care is a dynamic and emerging concept relevant, but not limited to, the end phase of life. It suggests a central philosophy underpinning renal service development that allows patients, carers and the multidisciplinary team time to work together to realize complex goals. It has relevance for the renal community and is likely to be integrated increasingly into everyday nephrology practice.

Wnt-β-catenin-Tcf-4 signalling-modulated invasiveness is dependent on osteopontin expression in breast cancer.

Background:We have previously demonstrated that Tcf-4 regulates osteopontin (OPN) in rat breast epithelial cells, Rama37. In this report, we have examined the importance of this regulation in human breast cancer.Methods:The regulatory roles of Tcf-4 on cell invasion and OPN expression were investigated. The mRNA expression of Tcf-4 and OPN, and survival of breast cancer patients were correlated.Results:Tcf-4 enhanced cell invasion in both MCF10AT and MDA MB 231 breast cancer cells by transcriptionally activating OPN expression. Osteopontin was activated by Wnt signalling in MDA MB 231 cells. Paradoxical results on Tcf-4-regulated OPN expression in MCF10AT (activation) and Rama37 (repression) cells were shown to be a result of differential Wnt signalling competency in MCF10AT and Rama37 cells. High levels of OPN and Tcf-4 mRNA expression were significantly associated with survival in breast cancer patients. Most importantly, Tcf-4-positive patients had a poorer prognosis when OPN was overexpressed, while OPN-negative patients had a better prognosis when Tcf-4 was overexpressed.Conclusion:Our results suggest that Tcf-4 can act as a repressor or activator of breast cancer progression by regulating OPN expression in a Wnt-dependent manner and that Tcf-4 and OPN together may be a novel prognostic indicator for breast cancer progression.

Concomitant and antecedent deep venous thrombosis and cancer survival in male US veterans

Survival is reportedly worse in patients with cancer concurrently diagnosed with deep venous thrombosis. However, information on specific malignancies is limited. From a cohort study of male US veterans we identified incident cancer cases (n aEuroS== aEuroS412 008) and compared survival patterns among those with versus without a history of deep venous thrombosis. Using Cox proportional hazard models, we estimated hazard ratios (HRs) and 95%% confidence intervals as measures of the relative risk of dying. Individuals with (versus without) a concomitant deep venous thrombosis and cancer diagnosis had a higher risk of dying (HR aEuroS== aEuroS1.38; 1.28--1.49). The most prominent excess mortality (HR aEuroS== aEuroS1.29--2.55) was observed among patients diagnosed with deep venous thrombosis at the time of diagnosis of lung, gastric, prostate, bladder, or kidney cancer. Increased risk of dying was also found among cancer patients diagnosed with deep venous thrombosis 1 year (HR aEuroS== aEuroS1.14; 1.07--1.22), 1--5 years (HR aEuroS== aEuroS1.14; 1.10--1.19), and > 5 years (HR aEuroS== aEuroS1.27; 1.23--1.31) before cancer; this was true for most cancer sites (HR aEuroS== aEuroS1.17--1.64). In summary, antecedent deep venous thrombosis confers a worse prognosis upon cancer patients. Advanced stage at diagnosis, treatment effects, lifestyle factors, and comorbidity could explain differences by cancer site and time frame between a prior deep venous thrombosis diagnosis and cancer outcome.

BRCA1 transcriptionally regulates genes associated with the basal breast cancer phenotype

Background Ten to twenty per cent of breast tumours exhibit a basallike genetic profile and these tumours carry a poor prognosis. Breast tumours which contain germline mutations for BRCA1 commonly exhibit a molecular profile similar to basal breast tumours. BRCA1 is a tumour suppressor gene which is mutated in up to 5–10% of breast cancer cases and is involved in multiple cellular processes including DNA damage control, cell cycle checkpoint control, apoptosis, ubiquitination and transcriptional regulation.

Methods Microarray-based profiling was carried out using the HCC1937EV and HCC1937BR breast cancer cell lines. Basal gene and protein expression levels were analysed by qRT-PCR and western blotting. ChIP analyses were performed and demonstrated that BRCA1 regulates basal gene expression through a transcriptional mechanism involving c-myc.

Results We have previously carried out microarray-based expression profiling to examine differences in gene expression when BRCA1 is reconstituted in BRCA1 mutated HCC1937 breast cancer cells. We observed that p-cadherin and the cytokeratin 5 and cytokeratin 17 genes, which are strongly correlated with the basal phenotype, are differentially expressed when BRCA1 is reconstituted. In addition, qRT-PCR and ChIP analysis of BRCA1 reconstituted cells show that BRCA1 represses the expression of these basal genes by a transcriptional mechanism. Furthermore, abrogation of endogenous BRCA1 protein in the T47D cell line using siRNA results in reexpression of these basal genes, suggesting that BRCA1 expression levels may be important in basal gene expression. We have also demonstrated that BRCA1 is physically associated with the promoter regions of basal genes through an association with c-myc. Consequently, we have confirmed that siRNA inhibition of c-myc in T47D cells results in re-expression of these genes.

Conclusions Our results suggest that BRCA1 is involved in the transcriptional regulation of genes associated with the basal phenotype and that BRCA1 controls basal gene expression through a transcriptional mechanism involving c-myc. Further work is now concentrating on defining the relationship between BRCA1 and basal gene expression and how this may affect clinical responses to breast cancer chemotherapy.