Predicting lung cancer recurrence from circulating tumour DNA. Commentary on 'Phylogenetic ctDNA analysis depicts early-stage lung cancer evolution'

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The Rewiring of Ubiquitination Targets in a Pathogenic Yeast Promotes Metabolic Flexibility, Host Colonization and Virulence

Funding: This work was funded by the European Research Council [http://erc.europa.eu/], AJPB (STRIFE Advanced Grant; C-2009-AdG-249793). The work was also supported by: the Wellcome Trust [www.wellcome.ac.uk], AJPB (080088, 097377); the UK Biotechnology and Biological Research Council [www.bbsrc.ac.uk], AJPB (BB/F00513X/1, BB/K017365/1); the CNPq-Brazil [http://cnpq.br], GMA (Science without Borders fellowship 202976/2014-9); and the National Centre for the Replacement, Refinement and Reduction of Animals in Research [www.nc3rs.org.uk], DMM (NC/K000306/1). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Acknowledgments We thank Dr. Elizabeth Johnson (Mycology Reference Laboratory, Bristol) for providing strains, and the Aberdeen Proteomics facility for the biotyping of S. cerevisiae clinical isolates, and to Euroscarf for providing S. cerevisiae strains and plasmids. We are grateful to our Microscopy Facility in the Institute of Medical Sciences for their expert help with the electron microscopy, and to our friends in the Aberdeen Fungal Group for insightful discussions.Peer reviewedPublisher PD

Metabolic determinants of cancer cell sensitivity to glucose limitation and biguanides

As the concentrations of highly consumed nutrients, particularly glucose, are generally lower in tumours than in normal tissues1,2, cancer cells must adapt their metabolism to the tumour microenvironment. A better understanding of these adaptations might reveal cancer cell liabilities that can be exploited for therapeutic benefit. Here, we developed a continuous flow culture apparatus (Nutrostat) for maintaining proliferating cells in low nutrient media for long periods of time and used it to undertake competitive proliferation assays on a pooled collection of barcoded cancer cell lines cultured in low glucose conditions. Sensitivity to low glucose varies amongst cell lines, and an RNAi screen pinpointed mitochondrial oxidative phosphorylation (OXPHOS) as the major pathway required for optimal proliferation in low glucose. We found that cell lines most sensitive to low glucose are defective in the upregulation of OXPHOS normally caused by glucose limitation as a result of either mtDNA mutations in Complex I genes or impaired glucose utilization. These defects predict sensitivity to biguanides, anti-diabetic drugs that inhibit OXPHOS3,4, when cancer cells are grown in low glucose or as tumour xenografts. Remarkably, the biguanide sensitivity of cancer cells with mtDNA mutations was reversed by ectopic expression of yeast NDI1, a ubiquinone oxidoreductase that allows bypass of Complex I function5. Thus, we conclude that mtDNA mutations and impaired glucose utilization are potential biomarkers for identifying tumours with increased sensitivity to OXPHOS inhibitors

Prehospital Electronic Patient Care Report Systems: Early Experiences from Emergency Medical Services Agency Leaders

Background: As the United States embraces electronic health records (EHRs), improved emergency medical services (EMS) information systems are also a priority; however, little is known about the experiences of EMS agencies as they adopt and implement electronic patient care report (e-PCR) systems. We sought to characterize motivations for adoption of e-PCR systems, challenges associated with adoption and implementation, and emerging implementation strategies. Methods: We conducted a qualitative study using semi-structured in-depth interviews with EMS agency leaders. Participants were recruited through a web-based survey of National Association of EMS Physicians (NAEMSP) members, a didactic session at the 2010 NAEMSP Annual Meeting, and snowball sampling. Interviews lasted approximately 30 minutes, were recorded and professionally transcribed. Analysis was conducted by a five-person team, employing the constant comparative method to identify recurrent themes. Results: Twenty-three interviewees represented 20 EMS agencies from the United States and Canada; 14 EMS agencies were currently using e-PCR systems. The primary reason for adoption was the potential for e-PCR systems to support quality assurance efforts. Challenges to e-PCR system adoption included those common to any health information technology project, as well as challenges unique to the prehospital setting, including: fear of increased ambulance run times leading to decreased ambulance availability, difficulty integrating with existing hospital information systems, and unfunded mandates requiring adoption of e-PCR systems. Three recurring strategies emerged to improve e-PCR system adoption and implementation: 1) identify creative funding sources; 2) leverage regional health information organizations; and 3) build internal information technology capacity. Conclusion: EMS agencies are highly motivated to adopt e-PCR systems to support quality assurance efforts; however, adoption and implementation of e-PCR systems has been challenging for many. Emerging strategies from EMS agencies and others that have successfully implemented EHRs may be useful in expanding e-PCR system use and facilitating this transition for other EMS agencies

Code Status Discussions Between Attending Hospitalist Physicians and Medical Patients at Hospital Admission

BackgroundBioethicists and professional associations give specific recommendations for discussing cardiopulmonary resuscitation (CPR).ObjectiveTo determine whether attending hospitalist physicians' discussions meet these recommendations.DesignCross-sectional observational study on the medical services at two hospitals within a university system between August 2008 and March 2009.ParticipantsAttending hospitalist physicians and patients who were able to communicate verbally about their medical care.Main measuresWe identified code status discussions in audio-recorded admission encounters via physician survey and review of encounter transcripts. A quantitative content analysis was performed to determine whether discussions included elements recommended by bioethicists and professional associations. Two coders independently coded all discussions; Cohen's kappa was 0.64-1 for all reported elements.Key resultsAudio-recordings of 80 patients' admission encounters with 27 physicians were obtained. Eleven physicians discussed code status in 19 encounters. Discussions were more frequent in seriously ill patients (OR 4, 95% CI 1.2-14.6), yet 66% of seriously ill patients had no discussion. The median length of the code status discussions was 1 min (range 0.2-8.2). Prognosis was discussed with code status in only one of the encounters. Discussions of patients' preferences focused on the use of life-sustaining interventions as opposed to larger life goals. Descriptions of CPR as an intervention used medical jargon, and the indication for CPR was framed in general, as opposed to patient-specific scenarios. No physician quantitatively estimated the outcome of or provided a recommendation about the use of CPR.ConclusionsCode status was not discussed with many seriously ill patients. Discussions were brief, and did not include elements that bioethicists and professional associations recommend to promote patient autonomy. Local and national guidelines, research, and clinical practice changes are needed to clarify and systematize with whom and how CPR is discussed at hospital admission

Mitochondrial Dysfunction Increases Oxidative Stress and Decreases Chronological Life Span in Fission Yeast

Background: Oxidative stress is a probable cause of aging and associated diseases. Reactive oxygen species (ROS) originate mainly from endogenous sources, namely the mitochondria. Methodology/Principal Findings: We analyzed the effect of aerobic metabolism on oxidative damage in Schizosaccharomyces pombe by global mapping of those genes that are required for growth on both respiratory-proficient media and hydrogen-peroxide-containing fermentable media. Out of a collection of approximately 2700 haploid yeast deletion mutants, 51 were sensitive to both conditions and 19 of these were related to mitochondrial function. Twelve deletion mutants lacked components of the electron transport chain. The growth defects of these mutants can be alleviated by the addition of antioxidants, which points to intrinsic oxidative stress as the origin of the phenotypes observed. These respiration-deficient mutants display elevated steady-state levels of ROS, probably due to enhanced electron leakage from their defective transport chains, which compromises the viability of chronologically-aged cells. Conclusion/Significance: Individual mitochondrial dysfunctions have often been described as the cause of diseases or aging, and our global characterization emphasizes the primacy of oxidative stress in the etiology of such processes.This work was supported by Dirección General de Investigación of Spain Grant BFU2006-02610, and by the Spanish program Consolider-Ingenio 2010 Grant CSD 2007-0020 to E.H

Robust Metabolic Responses to Varied Carbon Sources in Natural and Laboratory Strains of Saccharomyces cerevisiae

Understanding factors that regulate the metabolism and growth of an organism is of fundamental biologic interest. This study compared the influence of two different carbon substrates, dextrose and galactose, on the metabolic and growth rates of the yeast Saccharomyces cerevisiae. Yeast metabolic and growth rates varied widely depending on the metabolic substrate supplied. The metabolic and growth rates of a yeast strain maintained under long-term laboratory conditions was compared to strain isolated from natural condition when grown on different substrates. Previous studies had determined that there are numerous genetic differences between these two strains. However, the overall metabolic and growth rates of a wild isolate of yeast was very similar to that of a strain that had been maintained under laboratory conditions for many decades. This indicates that, at in least this case, metabolism and growth appear to be well buffered against genetic differences. Metabolic rate and cell number did not co-vary in a simple linear manner. When grown in either dextrose or galactose, both strains showed a growth pattern in which the number of cells continued to increase well after the metabolic rate began a sharp decline. Previous studied have reported that O2 consumption in S. cerevisiae grown in reduced dextrose levels were elevated compared to higher levels. Low dextrose levels have been proposed to induce caloric restriction and increase life span in yeast. However, there was no evidence that reduced levels of dextrose increased metabolic rates, measured by either O2 consumption or CO2 production, in the strains used in this study

Energy Metabolism in H460 Lung Cancer Cells: Effects of Histone Deacetylase Inhibitors

BACKGROUND: Tumor cells are characterized by accelerated growth usually accompanied by up-regulated pathways that ultimately increase the rate of ATP production. These cells can suffer metabolic reprogramming, resulting in distinct bioenergetic phenotypes, generally enhancing glycolysis channeled to lactate production. In the present work we showed metabolic reprogramming by means of inhibitors of histone deacetylase (HDACis), sodium butyrate and trichostatin. This treatment was able to shift energy metabolism by activating mitochondrial systems such as the respiratory chain and oxidative phosphorylation that were largely repressed in the untreated controls. METHODOLOGY/PRINCIPAL FINDINGS: Various cellular and biochemical parameters were evaluated in lung cancer H460 cells treated with the histone deacetylase inhibitors (HDACis), sodium butyrate (NaB) and trichostatin A (TSA). NaB and TSA reduced glycolytic flux, assayed by lactate release by H460 cells in a concentration dependent manner. NaB inhibited the expression of glucose transporter type 1 (GLUT 1), but substantially increased mitochondria bound hexokinase (HK) activity. NaB induced increase in HK activity was associated to isoform HK I and was accompanied by 1.5 fold increase in HK I mRNA expression and cognate protein biosynthesis. Lactate dehydrogenase (LDH) and pyruvate kinase (PYK) activities were unchanged by HDACis suggesting that the increase in the HK activity was not coupled to glycolytic flux. High resolution respirometry of H460 cells revealed NaB-dependent increased rates of oxygen consumption coupled to ATP synthesis. Metabolomic analysis showed that NaB altered the glycolytic metabolite profile of intact H460 cells. Concomitantly we detected an activation of the pentose phosphate pathway (PPP). The high O(2) consumption in NaB-treated cells was shown to be unrelated to mitochondrial biogenesis since citrate synthase (CS) activity and the amount of mitochondrial DNA remained unchanged. CONCLUSION: NaB and TSA induced an increase in mitochondrial function and oxidative metabolism in H460 lung tumor cells concomitant with a less proliferative cellular phenotype