Simon G. Royce

Use of Molecular Tumor Characteristics to Prioritize Mismatch Repair Gene Testing in Early-Onset Colorectal Cancer

PURPOSE The relationships between mismatch repair (MMR) protein expression, microsatellite instability (MSI), family history, and germline MMR gene mutation status have not been studied on a population basis. METHODS We studied 131 unselected patients with colorectal cancer diagnosed younger than age 45 years. For the 105 available tumors, MLH1, MSH2, MSH6, and PMS2 protein expression using immunohistochemistry (IHC) and MSI were measured. Germline DNA was screened for hMLH1, hMSH2, hMSH6, and hPMS2 mutations for the following patients: all from families fulfilling the Amsterdam Criteria for hereditary nonpolyposis colorectal cancer (HNPCC); all with tumors that were high MSI, low MSI, or that lacked expression of any MMR protein; and a random sample of 23 with MS-stable tumors expressing all MMR proteins. RESULTS Germline mutations were found in 18 patients (nine hMLH1, four hMSH2, four hMSH6, and one hPMS2); all tumors exhibited loss of MMR protein expression, all but one were high MSI or low MSI, and nine were from a family fulfilling Amsterdam Criteria. Sensitivities of IHC testing, MSI (high or low), and Amsterdam Criteria for MMR gene mutation were 100%, 94%, and 50%, respectively. Corresponding positive predictive values were 69%, 50%, and 75%. CONCLUSIONS Tumor IHC analysis of four MMR proteins and MSI testing provide a highly sensitive strategy for identifying MMR gene mutation-carrying, early-onset colorectal cancer patients, half of whom would have been missed using Amsterdam Criteria alone. Tumor-based approaches for triaging early-onset colorectal cancer patients for MMR gene mutation testing, irrespective of family history, appear to be an efficient screening strategy for HNPCC.

Obesity results in progressive atrial structural and electrical remodeling: implications for atrial fibrillation

BACKGROUND Obesity is associated with atrial fibrillation (AF); however, the mechanisms by which it induces AF are unknown. OBJECTIVE To examine the effect of progressive weight gain on the substrate for AF. METHODS Thirty sheep were studied at baseline, 4 months, and 8 months, following a high-calorie diet. Ten sheep were sampled at each time point for cardiac magnetic resonance imaging and hemodynamic studies. High-density multisite biatrial epicardial mapping was used to quantify effective refractory period, conduction velocity, and conduction heterogeneity index at 4 pacing cycle lengths and AF inducibility. Histology was performed for atrial fibrosis, inflammation, and intramyocardial lipidosis, and molecular analysis was performed for endothelin-A and -B receptors, endothelin-1 peptide, platelet-derived growth factor, transforming growth factor β1, and connective tissue growth factor. RESULTS Increasing weight was associated with increasing left atrial volume (P = .01), fibrosis (P = .02), inflammatory infiltrates (P = .01), and lipidosis (P = .02). While there was no change in the effective refractory period (P = .2), there was a decrease in conduction velocity (P<.001), increase in conduction heterogeneity index (P<.001), and increase in inducible (P = .001) and spontaneous (P = .001) AF. There was an increase in atrial cardiomyocyte endothelin-A and -B receptors (P = .001) and endothelin-1 (P = .03) with an increase in adiposity. In association, there was a significant increase in atrial interstitial and cytoplasmic transforming growth factor β1 (P = .02) and platelet-derived growth factor (P = .02) levels. CONCLUSIONS Obesity is associated with atrial electrostructural remodeling. With progressive obesity, there were changes in atrial size, conduction, histology, and expression of profibrotic mediators. These changes were associated with spontaneous and more persistent AF.

Electrophysiological, Electroanatomical, and Structural Remodeling of the Atria as Consequences of Sustained Obesity

BACKGROUND Obesity and atrial fibrillation (AF) are public health issues with significant consequences. OBJECTIVES This study sought to delineate the development of global electrophysiological and structural substrate for AF in sustained obesity. METHODS Ten sheep fed ad libitum calorie-dense diet to induce obesity over 36 weeks were maintained in this state for another 36 weeks; 10 lean sheep with carefully controlled weight served as controls. All sheep underwent electrophysiological and electroanatomic mapping; hemodynamic and imaging assessment (echocardiography and dual-energy x-ray absorptiometry); and histology and molecular evaluation. Evaluation included atrial voltage, conduction velocity (CV), and refractoriness (7 sites, 2 cycle lengths), vulnerability for AF, fatty infiltration, atrial fibrosis, and atrial transforming growth factor (TGF)-β1 expression. RESULTS Compared with age-matched controls, chronically obese sheep demonstrated greater total body fat (p < 0.001); LA volume (p < 0.001); LA pressure (p < 0.001), and PA pressures (p < 0.001); reduced atrial CV (LA p < 0.001) with increased conduction heterogeneity (p < 0.001); increased fractionated electrograms (p < 0.001); decreased posterior LA voltage (p < 0.001) and increased voltage heterogeneity (p < 0.001); no change in the effective refractory period (ERP) (p > 0.8) or ERP heterogeneity (p > 0.3). Obesity was associated with more episodes (p = 0.02), prolongation (p = 0.01), and greater cumulative duration (p = 0.02) of AF. Epicardial fat infiltrated the posterior LA in the obese group (p < 0.001), consistent with reduced endocardial voltage in this region. Atrial fibrosis (p = 0.03) and TGF-β1 protein (p = 0.002) were increased in the obese group. CONCLUSIONS Sustained obesity results in global biatrial endocardial remodeling characterized by LA enlargement, conduction abnormalities, fractionated electrograms, increased profibrotic TGF-β1 expression, interstitial atrial fibrosis, and increased propensity for AF. Obesity was associated with reduced posterior LA endocardial voltage and infiltration of contiguous posterior LA muscle by epicardial fat, representing a unique substrate for AF.

Ethnicity and risk for colorectal cancers showing somatic BRAF V600E mutation or CpG island methylator phenotype

Colorectal cancers arising from serrated polyps are characterized by the CpG island methylator phenotype (CIMP) and somatic mutation (V600E) in the BRAF proto-oncogene. Few epidemiologic studies have investigated risk factors for these tumors. We conducted a cohort study of 41,328 residents of Melbourne, Australia that included 9,939 participants of southern European origin and 31,389 of Anglo-Celtic origin. Colorectal adenocarcinomas were identified from population-based cancer registries. BRAF V600E mutation in tumors was determined using a PCR-based allelic discrimination method. Tumors were classified as CIMP positive when at least three of five markers (RUNX3, CACNA1G, SOCS1, NEUROG1, and IGF2) were methylated according to MethyLight analysis. Hazard ratios (HR) and 95% confidence intervals (95% CI) were estimated by Cox regression with adjustment for risk factors for colorectal cancer. During follow-up, 718 participants were diagnosed with colorectal cancer. CIMP assays were done for 579 and BRAF V600E mutation testing for 582. After adjustment for other risk factors, when compared with people of Anglo-Celtic origin, those of southern European origin had lower incidence of colorectal cancer that had CIMP (HR, 0.32; 95% CI, 0.16-0.67) or BRAF mutations (HR, 0.30; 95% CI, 0.16-0.58) but similar incidence of colorectal cancer without CIMP (HR, 0.86; 95% CI, 0.70-1.05) or BRAF (HR, 0.90; 95% CI, 0.74-1.11). People of southern European origin had lower risk of colorectal cancers with CIMP and BRAF mutation than people of Anglo-Celtic origin, which may in part be due to genetic factors that are less common in people of southern European origin. (Cancer Epidemiol Biomarkers Prev 2008;17(7):1774–80)

The regulation of fibrosis in airway remodeling in asthma.

Fibrosis is one of the key pathological features of airway remodeling in asthma. In the normal airway the amount of collagen and other extracellular matrix components is kept in equilibrium by regulation of synthesis and degradation. In asthma this homeostasis is disrupted due to genetic and environmental factors. In the airways of patients with the disease there is increased extracellular matrix deposition, particularly in the reticular basement membrane region, lamina propria and submucosa. Fibrosis is important as it can occur early in the pathogenesis of asthma, be associated with severity and resistant to therapy. In this review we will discuss current knowledge of relaxin and other key regulators of fibrosis in the airway including TGFβ, Smad2/3 and matrix metalloproteinases. As fibrosis is not directly targeted or effectively treated by current asthma drugs including corticosteroids, characterization of airway fibrosis and how it is regulated will be essential for the development of novel therapies for asthma.

Dietary Sulforaphane in Cancer Chemoprevention: The Role of Epigenetic Regulation and HDAC Inhibition

SIGNIFICANCE Sulforaphane, produced by the hydrolytic conversion of glucoraphanin after ingestion of cruciferous vegetables, particularly broccoli and broccoli sprouts, has been extensively studied due to its apparent health-promoting properties in disease and limited toxicity in normal tissue. Recent Studies: Recent identification of a sub-population of tumor cells with stem cell-like self-renewal capacity that may be responsible for relapse, metastasis, and resistance, as a potential target of the dietary compound, may be an important aspect of sulforaphane chemoprevention. Evidence also suggests that sulforaphane may target the epigenetic alterations observed in specific cancers, reversing aberrant changes in gene transcription through mechanisms of histone deacetylase inhibition, global demethylation, and microRNA modulation. CRITICAL ISSUES In this review, we discuss the biochemical and biological properties of sulforaphane with a particular emphasis on the anticancer properties of the dietary compound. Sulforaphane possesses the capacity to intervene in multistage carcinogenesis through the modulation and/or regulation of important cellular mechanisms. The inhibition of phase I enzymes that are responsible for the activation of pro-carcinogens, and the induction of phase II enzymes that are critical in mutagen elimination are well-characterized chemopreventive properties. Furthermore, sulforaphane mediates a number of anticancer pathways, including the activation of apoptosis, induction of cell cycle arrest, and inhibition of NFκB. FUTURE DIRECTIONS Further characterization of the chemopreventive properties of sulforaphane and its capacity to be selectively toxic to malignant cells are warranted to potentially establish the clinical utility of the dietary compound as an anti-cancer compound alone, and in combination with clinically relevant therapeutic and management strategies.

Serelaxin Is a More Efficacious Antifibrotic Than Enalapril in an Experimental Model of Heart Disease

Relaxin is a naturally occurring peptide hormone that mediates systemic hemodynamic and renal adaptive changes during pregnancy and abrogates aberrant scar tissue formation (fibrosis) in diverse pathogeneses. However, its efficacy relative to renin–angiotensin system blockade, the most effective antifibrotic strategy currently available, is not known. We compared the individual versus combined antifibrotic effects of serelaxin (a recombinant form of human gene-2 relaxin) and the angiotensin-converting enzyme inhibitor enalapril, in preventative (started before injury) and therapeutic (treatment of established fibrosis) strategies, in a mouse model of isoprenaline-induced cardiac injury (at 17 days). Changes in systolic blood pressure, organ hypertrophy, and tissue remodeling/fibrosis were assessed. Pretreatment with serelaxin (0.5 mg/kg per day via subcutaneous administration) alone reduced cardiac fibrosis to a greater extent than enalapril (200 mg/L via drinking water; equivalent to 48 mg/kg per day) alone (P<0.05 versus enalapril alone). Additionally, the combined effects of serelaxin and enalapril reduced cardiac fibrosis by at least 2-fold compared with enalapril alone, when administered preventatively or therapeutically; by suppressing transforming growth factor-&bgr;1 expression and phosphorylation of Smad2 (an intracellular regulator of transforming growth factor-&bgr;1 activity; both P<0.05 versus enalapril alone) to a greater extent. The effects of serelaxin were independent of blood pressure, while enalapril lowered systolic blood pressure in the model studied. These findings suggest that serelaxin alone and in combination with an angiotensin-converting enzyme inhibitor more effectively ameliorates fibrosis than angiotensin-converting enzyme inhibition alone in the diseased heart, in a clinically relevant experimental scenario.

Histone deacetylases and their inhibitors: new implications for asthma and chronic respiratory conditions.

Purpose of reviewThe importance of the role of histone deacetylase enzymes in the pathogenesis of asthma and chronic respiratory diseases is increasingly being recognized. Similarly, the potential clinical utility of histone deacetylase enzymes in the treatment of disease is emerging. In this review, the role of histone deacetylases (HDAC) and their inhibitors in asthma and chronic obstructive pulmonary disease is discussed. Recent findingsHDAC are able to catalyze the hydrolysis of acetyl groups on lysine residues of histones, causing the condensation and coiling of chromosomal DNA around histones, and therefore regulating gene expression. Histone deacetylase inhibitors act specifically or broadly on HDAC and also on nonhistone targets. Some have been used in the oncology therapeutic field for some years, but it is only more recently that they have been suggested in the treatment of asthma and other inflammatory lung diseases. SummaryImportant developments have been made in the understanding of histone deacetylase expression in normal and diseased airways and pulmonary tissue as well as effects of histone deacetylase inhibitors on structural and inflammatory cells in the lung, including cell proliferation, differentiation and apoptosis and senescence. Many of these discoveries may have implications in addressing airway inflammation, airway remodeling and airway hyperresponsiveness in asthma.

Relaxin reverses airway remodeling and airway dysfunction in allergic airways disease.

Mice deficient in the antifibrotic hormone relaxin develop structural changes in the airway that resemble airway remodeling, and demonstrate exaggerated remodeling changes in models of allergic airways disease (AAD). Relaxin expression in asthma has not been previously studied. We evaluated the efficacy of relaxin in the treatment of established airway remodeling in a mouse model of AAD. Relaxin expression in mouse AAD was also examined by immunohistochemistry and real-time PCR. BALB/c mice with established AAD were treated with relaxin or vehicle control (sc for 14 d), and effects on airway remodeling, airway inflammation, and airway hyperresponsiveness (AHR) were assessed. Relaxin expression was significantly reduced in the airways of mice with AAD compared with controls. Recombinant relaxin treatment in a mouse model of AAD reversed collagen deposition and epithelial thickening, and significantly improved AHR (all P < 0.05 vs. vehicle control), but did not influence airway inflammation or goblet cell hyperplasia. Relaxin treatment was associated with increased matrix metalloproteinase-2 levels, suggesting a possible mechanism for its antifibrotic effects. Endogenous relaxin expression is decreased in murine AAD, whereas exogenous relaxin represents a novel treatment capable of reversing established airway remodeling and AHR.

Novel therapeutic strategies for lung disorders associated with airway remodelling and fibrosis

Inflammatory cell infiltration, cytokine release, epithelial damage, airway/lung remodelling and fibrosis are central features of inflammatory lung disorders, which include asthma, chronic obstructive pulmonary disease, acute respiratory distress syndrome and idiopathic pulmonary fibrosis. Although the lung has some ability to repair itself from acute injury, in the presence of ongoing pathological stimuli and/or insults that lead to chronic disease, it no longer retains the capacity to heal, resulting in fibrosis, the final common pathway that causes an irreversible loss of lung function. Despite inflammation, genetic predisposition/factors, epithelial-mesenchymal transition and mechanotransduction being able to independently contribute to airway remodelling and fibrosis, current therapies for inflammatory lung diseases are limited by their ability to only target the inflammatory component of the disease without having any marked effects on remodelling (epithelial damage and fibrosis) that can cause lung dysfunction independently of inflammation. Furthermore, as subsets of patients suffering from these diseases are resistant to currently available therapies (such as corticosteroids), novel therapeutic approaches are required to combat all aspects of disease pathology. This review discusses emerging therapeutic approaches, such as trefoil factors, relaxin, histone deacetylase inhibitors and stem cells, amongst others that have been able to target airway inflammation and airway remodelling while improving related lung dysfunction. A better understanding of the mode of action of these therapies and their possible combined effects may lead to the identification of their clinical potential in the setting of lung disease, either as adjunct or alternative therapies to currently available treatments.