Differences in resistance to 5-fluorouracil as a function of cell cycle delay and not apoptosis.

A series of human embryo fibroblasts has previously been shown to display increasing resistance to the antimetabolites methotrexate (MTX) and N-phosphonacetyl-L-aspartate (PALA) with increasing tumorigenicity. This increased resistance was found to be further increased as a result of salvage pathway activity for purine and pyrimidine biosynthesis. A similar pattern of increasing resistance paralleling increasing tumorigenicity has now been shown to occur with 5-fluorouracil (5-FU), which is independent of salvage pathway activity. The KMS normal cell line was found to be more sensitive to 5-FU than either the immortalised KMST or tumorigenic KN-NM cell lines. Immunohistochemical analysis of the three cell lines demonstrated high levels of p53 protein in the KMST and KN-NM cell lines, but undetectable p53 levels in the KMS cell line. From these data it was hypothesised that a difference in p53 function may be causing the difference in the patterns of sensitivity observed in the three cell lines. P53 is now believed to function as a regulator of a G1 to S cell cycle checkpoint and as an inducer of apoptosis following DNA damage to the cell. The differences in sensitivity of the cell lines could not be explained by differences in the levels of apoptosis but could be attributed to differences in cell cycle response. Our evidence suggests that loss of cell cycle control, possibly through loss of p53 function, is an important factor in increasing the drug resistance of fibroblast cell lines

A caspase-3 'death-switch' in colorectal cancer cells for induced and synchronous tumor apoptosis in vitro and in vivo facilitates the development of minimally invasive cell death biomarkers

Novel anticancer drugs targeting key apoptosis regulators have been developed and are undergoing clinical trials. Pharmacodynamic biomarkers to define the optimum dose of drug that provokes tumor apoptosis are in demand; acquisition of longitudinal tumor biopsies is a significant challenge and minimally invasive biomarkers are required. Considering this, we have developed and validated a preclinical 'death-switch' model for the discovery of secreted biomarkers of tumour apoptosis using in vitro proteomics and in vivo evaluation of the novel imaging probe [ 18 F]ML-10 for non-invasive detection of apoptosis using positron emission tomography (PET). The 'death-switch' is a constitutively active mutant caspase-3 that is robustly induced by doxycycline to drive synchronous apoptosis in human colorectal cancer cells in vitro or grown as tumor xenografts. Deathswitch induction caused caspase-dependent apoptosis between 3 and 24 hours in vitro and regression of 'death-switched' xenografts occurred within 24 h correlating with the percentage of apoptotic cells in tumor and levels of an established cell death biomarker (cleaved cytokeratin-18) in the blood. We sought to define secreted biomarkers of tumor apoptosis from cultured cells using Discovery Isobaric Tag proteomics, which may provide candidates to validate in blood. Early after caspase-3 activation, levels of normally secreted proteins were decreased (e.g. Gelsolin and Midkine) and proteins including CD44 and High Mobility Group protein B1 (HMGB1) that were released into cell culture media in vitro were also identified in the bloodstream of mice bearing death-switched tumors. We also exemplify the utility of the death-switch model for the validation of apoptotic imaging probes using [ 18 F]ML-10, a PET tracer currently in clinical trials. Results showed increased tracer uptake of [ 18 F]ML-10 in tumours undergoing apoptosis, compared with matched tumour controls imaged in the same animal. Overall, the death-switch model represents a robust and versatile tool for the discovery and validation of apoptosis biomarkers. © 2013 Macmillan Publishers Limited. All rights reserved

Beyond platitudes: a qualitative study of Australian Aboriginal people's perspectives on biobanking.

BACKGROUND: Biobanks are vital resources for genetics and genomics, and it is broadly recognised that for maximal benefit it is essential that they include samples and data from diverse ancestral groups. The inclusion of First Nations people, in particular, is important to prevent biobanking research from exacerbating existing health inequities, and to ensure that these communities share in the benefits arising from research. AIMS: To explore the perspectives of Australian Aboriginal people whose tissue - or that of their family members - has been stored in the biobank of the National Centre for Indigenous Genomics (NCIG). METHODS: Semi-structured interviews with 42 Aboriginal people from the Titjikala, Galiwinku, Tiwi Islands, Yarrabah, Fitzroy Crossing, Derby, One Arm Point and Mulan communities, as well as a formal discussion with A. Hermes, an Indigenous Community Engagement Coordinator at the NCIG who had conducted the interviews. The interviews and the structured discussion were double coded using a procedure informed by Charmaz's outline of grounded theory analysis and Morse's outline of the cognitive basis of qualitative research. RESULTS: In this article, we report on A. Hermes' interviews with members from the above Aboriginal communities, as well as on her personal views, experiences, and interpretations of the interviews she conducted with other community members. We found that participation in the NCIG biobank raised issues around broken trust, grief and loss, but also - somewhat unexpectedly - was perceived as a source of empowerment, hope and reconnection. CONCLUSIONS: This research reminds us (again) of the need to engage deeply with communities in order to respond appropriately with respect for their cultural values and norms, and to develop culturally relevant policies and processes that enhance the benefits of biobank participation and minimise potential harms

Balancing selection of a frame-shift mutation in the MRC2 gene accounts for the outbreak of the crooked tail syndrome in Belgian blue cattle

Abstract We herein describe the positional identification of a 2-bp deletion in the open reading frame of the MRC2 receptor causing the recessive Crooked Tail Syndrome in cattle. The resulting frame-shift reveals a premature stop codon that causes nonsense-mediated decay of the mutant messenger RNA, and the virtual absence of functional Endo180 protein in affected animals. Cases exhibit skeletal anomalies thought to result from impaired extracellular matrix remodeling during ossification, and as of yet unexplained muscular symptoms. We demonstrate that carrier status is very significantly associated with desired characteristics in the general population, including enhanced muscular development, and that the resulting heterozygote advantage caused a selective sweep which explains the unexpectedly high frequency (25%) of carriers in the Belgian Blue Cattle Breed

Brain age as a surrogate marker for cognitive performance in multiple sclerosis

Background: Data from neuro-imaging techniques allow us to estimate a brain's age. Brain age is easily interpretable as "how old the brain looks", and could therefore be an attractive communication tool for brain health in clinical practice. This study aimed to investigate its clinical utility by investigating the relationship between brain age and cognitive performance in multiple sclerosis (MS). Methods: A linear regression model was trained to predict age from brain MRI volumetric features and sex in a healthy control dataset (HC_train, n=1673). This model was used to predict brain age in two test sets: HC_test (n=50) and MS_test (n=201). Brain-Predicted Age Difference (BPAD) was calculated as BPAD=brain age minus chronological age. Cognitive performance was assessed by the Symbol Digit Modalities Test (SDMT). Results: Brain age was significantly related to SDMT scores in the MS_test dataset (r=-0.46, p<.001), and contributed uniquely to variance in SDMT beyond chronological age, reflected by a significant correlation between BPAD and SDMT (r=-0.24, p<.001) and a significant weight (-0.25, p=0.002) in a multivariate regression equation with age. Conclusions: Brain age is a candidate biomarker for cognitive dysfunction in MS and an easy to grasp metric for brain health

EPAC-Lung:Pooled analysis of circulating tumor cells in advanced non-small cell lung cancer

Background: We assessed the clinical validity of circulating tumor cell (CTC) quantification for prognostication of patients with advanced non-small cell lung cancer (NSCLC) by undertaking a European pooled analysis of individual patient data. This is the largest study of its kind and the first to examine between-centre heterogeneity of CTC identification in NSCLC.Methods: Nine European NSCLC CTC centers were asked to provide reported/unreported anonymised data for patients with advanced NSCLC who participated in CellSearch CTC studies from January 2003 - March 2017. We used Cox regression models, stratified by centre, to establish the association between CTC count and survival. We assessed the added value of CTCs to prognostic clinico-pathological models using likelihood ratio (LR) statistics and c-indices.Results: Seven out of nine eligible centers provided data for 550 eligible patients, including 209 patients whose prognostic information was previously unpublished. CTC counts of ≥ 2 and ≥5 per 7·5 mL were associated with reduced progression-free survival (≥2 CTCs: HR 1.72, p &lt; 0·001; ≥5 CTCs: HR 2.21, p &lt; 0·001) and overall survival (≥2 CTCs: HR 2·18, p &lt; 0·001; ≥5 CTCs: HR 2·75, p &lt; 0·001), respectively. Survival prediction was significantly improved by addition of baseline CTC count to LR clinico-pathological models (log-transformed CTCs p &lt; 0·0001; ≥2 CTCs p &lt; 0·0001; ≥5 CTCs p &lt; 0·0001), while more moderate improvements were observed with the use of c-index models. There was minor evidence of between-center heterogeneity in the effect on PFS, but not OS.No difference in CTC profile was observed between key NSCLC molecular subsets such as EGFR, ALK, and KRAS.Conclusions: These data confirm CTCs as an independent prognostic indicator of progression-free survival and overall survival in advanced NSCLC. CTC count improves prognostication when added to full clinico-pathological predictive models. ≥2 CTCs is an appropriate cutoff to move towards establishing clinical utility

The efficacy and safety of prokinetic agents in critically ill patients receiving enteral nutrition: a systematic review and meta-analysis of randomized trials.

BACKGROUND: Intolerance to enteral nutrition is common in critically ill adults, and may result in significant morbidity including ileus, abdominal distension, vomiting and potential aspiration events. Prokinetic agents are prescribed to improve gastric emptying. However, the efficacy and safety of these agents in critically ill patients is not well-defined. Therefore, we conducted a systematic review and meta-analysis to determine the efficacy and safety of prokinetic agents in critically ill patients. METHODS: We searched MEDLINE, EMBASE, and Cochrane Library from inception up to January 2016. Eligible studies included randomized controlled trials (RCTs) of critically ill adults assigned to receive a prokinetic agent or placebo, and that reported relevant clinical outcomes. Two independent reviewers screened potentially eligible articles, selected eligible studies, and abstracted pertinent data. We calculated pooled relative risk (RR) for dichotomous outcomes and mean difference for continuous outcomes, with the corresponding 95 % confidence interval (CI). We assessed risk of bias using Cochrane risk of bias tool, and the quality of evidence using grading of recommendations assessment, development, and evaluation (GRADE) methodology. RESULTS: Thirteen RCTs (enrolling 1341 patients) met our inclusion criteria. Prokinetic agents significantly reduced feeding intolerance (RR 0.73, 95 % CI 0.55, 0.97; P = 0.03; moderate certainty), which translated to 17.3 % (95 % CI 5, 26.8 %) absolute reduction in feeding intolerance. Prokinetics also reduced the risk of developing high gastric residual volumes (RR 0.69; 95 % CI 0.52, 0.91; P = 0.009; moderate quality) and increased the success of post-pyloric feeding tube placement (RR 1.60, 95 % CI 1.17, 2.21; P = 0.004; moderate quality). There was no significant improvement in the risk of vomiting, diarrhea, intensive care unit (ICU) length of stay or mortality. Prokinetic agents also did not significantly increase the rate of diarrhea. CONCLUSION: There is moderate-quality evidence that prokinetic agents reduce feeding intolerance in critically ill patients compared to placebo or no intervention. However, the impact on other clinical outcomes such as pneumonia, mortality, and ICU length of stay is unclear