Assessing proliferation, cell-cycle arrest and apoptotic end points in human buccal punch biopsies for use as pharmacodynamic biomarkers in drug development

Easily accessible normal tissues expressing the same molecular site(s) of drug action as malignant tissue offer an enhanced potential for early proof of anticancer drug mechanism and estimation of the biologically effective dose. Studies were undertaken in healthy male volunteers to assess the tolerability of single and multiple (four in 24 h) 3 mm punch biopsies of the buccal mucosa, and to determine the feasibility of detecting and quantifying a range of proliferation, cell-cycle arrest and apoptosis markers by immunohistochemistry (IHC) for use as potential pharmacodynamic (PD) end points. The biopsy procedure was well tolerated with 100% of volunteers stating that they would undergo single (n=10) and multiple (n=12) biopsies again. Total retinoblastoma protein (pRb), phosphorylated pRb (phospho-pRb), total p27, phosphorylated p27 (phospho-p27), phosphorylated-histone H3 (phospho-HH3), p21, p53, Cyclin A, Cyclin E, Ki67 all produced good signal detection, but M30, cleaved caspase 3 and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling did not. Total pRb, phospho-pRb, total p27 and phospho-p27 were quantified further in a multiple biopsy study to allow components of variability to be addressed to inform future sizing decisions on intervention studies. Neither site of biopsy within the oral cavity, nor the nominal time of biopsy had any significant impact on any of the four markers expression levels. Inter- and intrasubject coefficients of variation (CVs) that could be used to size future intervention studies for pRb, phospho-pRb, total p27 and phospho-p27 were 14, 19, 18 and 16%; and 18, 29, 25 and 19%, respectively. In conclusion, quantitation of such markers in 3 mm buccal punch biopsies would be suitable to explore as PD end points within intervention studies of drugs acting on these pathways

Pleiotropic effects of statins in distal human pulmonary artery smooth muscle cells

<p>Abstract</p> <p>Background</p> <p>Recent clinical data suggest statins have transient but significant effects in patients with pulmonary arterial hypertension. In this study we explored the molecular effects of statins on distal human pulmonary artery smooth muscle cells (PASMCs) and their relevance to proliferation and apoptosis in pulmonary arterial hypertension.</p> <p>Methods</p> <p>Primary distal human PASMCs from patients and controls were treated with lipophilic (simvastatin, atorvastatin, mevastatin and fluvastatin), lipophobic (pravastatin) and nitric-oxide releasing statins and studied in terms of their DNA synthesis, proliferation, apoptosis, matrix metalloproteinase-9 and endothelin-1 release.</p> <p>Results</p> <p>Treatment of human PASMCs with selected statins inhibited DNA synthesis, proliferation and matrix metalloproteinase-9 production in a concentration-dependent manner. Statins differed in their effectiveness, the rank order of anti-mitogenic potency being simvastatin > atorvastatin > > pravastatin. Nevertheless, a novel nitric oxide-releasing derivative of pravastatin (NCX 6550) was effective. Lipophilic statins, such as simvastatin, also enhanced the anti-proliferative effects of iloprost and sildenafil, promoted apoptosis and inhibited the release of the mitogen and survival factor endothelin-1. These effects were reversed by mevalonate and the isoprenoid intermediate geranylgeranylpyrophosphate and were mimicked by inhibitors of the Rho and Rho-kinase.</p> <p>Conclusions</p> <p>Lipophilic statins exert direct effects on distal human PASMCs and are likely to involve inhibition of Rho GTPase signalling. These findings compliment some of the recently documented effects in patients with pulmonary arterial hypertension.</p

Outcomes of obstructed abdominal wall hernia: results from the UK national small bowel obstruction audit

Background: Abdominal wall hernia is a common surgical condition. Patients may present in an emergency with bowel obstruction, incarceration or strangulation. Small bowel obstruction (SBO) is a serious surgical condition associated with significant morbidity. The aim of this study was to describe current management and outcomes of patients with obstructed hernia in the UK as identified in the National Audit of Small Bowel Obstruction (NASBO). Methods: NASBO collated data on adults treated for SBO at 131 UK hospitals between January and March 2017. Those with obstruction due to abdominal wall hernia were included in this study. Demographics, co-morbidity, imaging, operative treatment, and in-hospital outcomes were recorded. Modelling for factors associated with mortality and complications was undertaken using Cox proportional hazards and multivariable regression modelling. Results: NASBO included 2341 patients, of whom 415 (17·7 per cent) had SBO due to hernia. Surgery was performed in 312 (75·2 per cent) of the 415 patients; small bowel resection was required in 198 (63·5 per cent) of these operations. Non-operative management was reported in 35 (54 per cent) of 65 patients with a parastomal hernia and in 34 (32·1 per cent) of 106 patients with an incisional hernia. The in-hospital mortality rate was 9·4 per cent (39 of 415), and was highest in patients with a groin hernia (11·1 per cent, 17 of 153). Complications were common, including lower respiratory tract infection in 16·3 per cent of patients with a groin hernia. Increased age was associated with an increased risk of death (hazard ratio 1·05, 95 per cent c.i. 1·01 to 1·10; P = 0·009) and complications (odds ratio 1·05, 95 per cent c.i. 1·02 to 1·09; P = 0·001). Conclusion: NASBO has highlighted poor outcomes for patients with SBO due to hernia, highlighting the need for quality improvement initiatives in this group

A Lung Segmental Model of Chronic <em>Pseudomonas</em> Infection in Sheep

Chronic lung infection with Pseudomonas aeruginosa is a major contributor to morbidity, mortality and premature death in cystic fibrosis. A new paradigm for managing such infections is needed, as are relevant and translatable animal models to identify and test concepts. We sought to improve on limitations associated with existing models of infection in small animals through developing a lung segmental model of chronic Pseudomonas infection in sheep.Using local lung instillation of P. aeruginosa suspended in agar beads we were able to demonstrate that such infection led to the development of a suppurative, necrotising and pyogranulomatous pneumonia centred on the instilled beads. No overt evidence of organ or systemic compromise was apparent in any animal during the course of infection. Infection persisted in the lungs of individual animals for as long as 66 days after initial instillation. Quantitative microbiology applied to bronchoalveolar lavage fluid derived from infected segments proved an insensitive index of the presence of significant infection in lung tissue (>10(4) cfu/g).The agar bead model of chronic P. aeruginosa lung infection in sheep is a relevant platform to investigate both the pathobiology of such infections as well as novel approaches to their diagnosis and therapy. Particular ethical benefits relate to the model in terms of refining existing approaches by compromising a smaller proportion of the lung with infection and facilitating longitudinal assessment by bronchoscopy, and also potentially reducing animal numbers through facilitating within-animal comparisons of differential therapeutic approaches

Outcomes of obstructed abdominal wall hernia: results from the UK national small bowel obstruction audit

Background Abdominal wall hernia is a common surgical condition. Patients may present in an emergency with bowel obstruction, incarceration or strangulation. Small bowel obstruction (SBO) is a serious surgical condition associated with significant morbidity. The aim of this study was to describe current management and outcomes of patients with obstructed hernia in the UK as identified in the National Audit of Small Bowel Obstruction (NASBO). Methods NASBO collated data on adults treated for SBO at 131 UK hospitals between January and March 2017. Those with obstruction due to abdominal wall hernia were included in this study. Demographics, co‐morbidity, imaging, operative treatment, and in‐hospital outcomes were recorded. Modelling for factors associated with mortality and complications was undertaken using Cox proportional hazards and multivariable regression modelling. Results NASBO included 2341 patients, of whom 415 (17·7 per cent) had SBO due to hernia. Surgery was performed in 312 (75·2 per cent) of the 415 patients; small bowel resection was required in 198 (63·5 per cent) of these operations. Non‐operative management was reported in 35 (54 per cent) of 65 patients with a parastomal hernia and in 34 (32·1 per cent) of 106 patients with an incisional hernia. The in‐hospital mortality rate was 9·4 per cent (39 of 415), and was highest in patients with a groin hernia (11·1 per cent, 17 of 153). Complications were common, including lower respiratory tract infection in 16·3 per cent of patients with a groin hernia. Increased age was associated with an increased risk of death (hazard ratio 1·05, 95 per cent c.i. 1·01 to 1·10; P = 0·009) and complications (odds ratio 1·05, 95 per cent c.i. 1·02 to 1·09; P = 0·001). Conclusion NASBO has highlighted poor outcomes for patients with SBO due to hernia, highlighting the need for quality improvement initiatives in this group

Outcomes following small bowel obstruction due to malignancy in the national audit of small bowel obstruction

Introduction Patients with cancer who develop small bowel obstruction are at high risk of malnutrition and morbidity following compromise of gastrointestinal tract continuity. This study aimed to characterise current management and outcomes following malignant small bowel obstruction. Methods A prospective, multicentre cohort study of patients with small bowel obstruction who presented to UK hospitals between 16th January and 13th March 2017. Patients who presented with small bowel obstruction due to primary tumours of the intestine (excluding left-sided colonic tumours) or disseminated intra-abdominal malignancy were included. Outcomes included 30-day mortality and in-hospital complications. Cox-proportional hazards models were used to generate adjusted effects estimates, which are presented as hazard ratios (HR) alongside the corresponding 95% confidence interval (95% CI). The threshold for statistical significance was set at the level of P ≤ 0.05 a-priori. Results 205 patients with malignant small bowel obstruction presented to emergency surgery services during the study period. Of these patients, 50 had obstruction due to right sided colon cancer, 143 due to disseminated intraabdominal malignancy, 10 had primary tumours of the small bowel and 2 patients had gastrointestinal stromal tumours. In total 100 out of 205 patients underwent a surgical intervention for obstruction. 30-day in-hospital mortality rate was 11.3% for those with primary tumours and 19.6% for those with disseminated malignancy. Severe risk of malnutrition was an independent predictor for poor mortality in this cohort (adjusted HR 16.18, 95% CI 1.86 to 140.84, p = 0.012). Patients with right-sided colon cancer had high rates of morbidity. Conclusions Mortality rates were high in patients with disseminated malignancy and in those with right sided colon cancer. Further research should identify optimal management strategy to reduce morbidity for these patient groups

National prospective cohort study of the burden of acute small bowel obstruction

Background Small bowel obstruction is a common surgical emergency, and is associated with high levels of morbidity and mortality across the world. The literature provides little information on the conservatively managed group. The aim of this study was to describe the burden of small bowel obstruction in the UK. Methods This prospective cohort study was conducted in 131 acute hospitals in the UK between January and April 2017, delivered by trainee research collaboratives. Adult patients with a diagnosis of mechanical small bowel obstruction were included. The primary outcome was in‐hospital mortality. Secondary outcomes included complications, unplanned intensive care admission and readmission within 30 days of discharge. Practice measures, including use of radiological investigations, water soluble contrast, operative and nutritional interventions, were collected. Results Of 2341 patients identified, 693 (29·6 per cent) underwent immediate surgery (within 24 h of admission), 500 (21·4 per cent) had delayed surgery after initial conservative management, and 1148 (49·0 per cent) were managed non‐operatively. The mortality rate was 6·6 per cent (6·4 per cent for non‐operative management, 6·8 per cent for immediate surgery, 6·8 per cent for delayed surgery; P = 0·911). The major complication rate was 14·4 per cent overall, affecting 19·0 per cent in the immediate surgery, 23·6 per cent in the delayed surgery and 7·7 per cent in the non‐operative management groups (P < 0·001). Cox regression found hernia or malignant aetiology and malnutrition to be associated with higher rates of death. Malignant aetiology, operative intervention, acute kidney injury and malnutrition were associated with increased risk of major complication. Conclusion Small bowel obstruction represents a significant healthcare burden. Patient‐level factors such as timing of surgery, acute kidney injury and nutritional status are factors that might be modified to improve outcomes

In vivo efficacy of novel monobactam LYS228 in murine models of carbapenemase-producing Klebsiella pneumoniae infection

LYS228 has potent antibacterial activity against carbapenem-resistant strains of Enterobacteriaceae. LYS228 was efficacious in neutropenic thigh models established with K. pneumoniae producing either KPC-2 or NDM-1; pre-treatment with uranyl nitrate considerably shifted calculated static doses of LYS228. In murine ascending pyelonephritis, LYS228 reduced bacterial burden in the kidney, urine and bladder. The successful treatment of murine infection models established with carbapenem resistant K. pneumoniae further supports the clinical development of LYS228