Analysis of cancer metabolism with high-throughput technologies

<p>Abstract</p> <p>Background</p> <p>Recent advances in genomics and proteomics have allowed us to study the nuances of the Warburg effect – a long-standing puzzle in cancer energy metabolism – at an unprecedented level of detail. While modern next-generation sequencing technologies are extremely powerful, the lack of appropriate data analysis tools makes this study difficult. To meet this challenge, we developed a novel application for comparative analysis of gene expression and visualization of RNA-Seq data.</p> <p>Results</p> <p>We analyzed two biological samples (normal human brain tissue and human cancer cell lines) with high-energy, metabolic requirements. We calculated digital topology and the copy number of every expressed transcript. We observed subtle but remarkable qualitative and quantitative differences between the citric acid (TCA) cycle and glycolysis pathways. We found that in the first three steps of the TCA cycle, digital expression of aconitase 2 (<it>ACO2</it>) in the brain exceeded both citrate synthase (<it>CS</it>) and isocitrate dehydrogenase 2 (<it>IDH2</it>), while in cancer cells this trend was quite the opposite. In the glycolysis pathway, all genes showed higher expression levels in cancer cell lines; and most notably, digital gene expression of glyceraldehyde-3-phosphate dehydrogenase (<it>GAPDH</it>) and enolase (<it>ENO</it>) were considerably increased when compared to the brain sample.</p> <p>Conclusions</p> <p>The variations we observed should affect the rates and quantities of ATP production. We expect that the developed tool will provide insights into the subtleties related to the causality between the Warburg effect and neoplastic transformation. Even though we focused on well-known and extensively studied metabolic pathways, the data analysis and visualization pipeline that we developed is particularly valuable as it is global and pathway-independent.</p

Comparison of quality of life after stereotactic body radiotherapy and surgery for early-stage prostate cancer

Background: As the long-term efficacy of stereotactic body radiation therapy (SBRT) becomes established and other prostate cancer treatment approaches are refined and improved, examination of quality of life (QOL) following prostate cancer treatment is critical in driving both patient and clinical treatment decisions. We present the first study to compare QOL after SBRT and radical prostatectomy, with QOL assessed at approximately the same times pre- and post-treatment and using the same validated QOL instrument. Methods: Patients with clinically localized prostate cancer were treated with either radical prostatectomy (n = 123 Spanish patients) or SBRT (n = 216 American patients). QOL was assessed using the Expanded Prostate Cancer Index Composite (EPIC) grouped into urinary, sexual, and bowel domains. For comparison purposes, SBRT EPIC data at baseline, 3 weeks, 5, 11, 24, and 36 months were compared to surgery data at baseline, 1, 6, 12, 24,and 36 months. Differences in patient characteristics between the two groups were assessed using Chi-squared tests for categorical variables and t-tests for continuous variables. Generalized estimating equation (GEE) models were constructed for each EPIC scale to account for correlation among repeated measures and used to assess the effect of treatment on QOL. Results: The largest differences in QOL occurred in the first 1-6 months after treatment, with larger declines following surgery in urinary and sexual QOL as compared to SBRT, and a larger decline in bowel QOL following SBRT as compared to surgery. Long-term urinary and sexual QOL declines remained clinically significantly lower for surgery patients but not for SBRT patients. Conclusions: Overall, these results may have implications for patient and physician clinical decision making which are often influenced by QOL. These differences in sexual, urinary and bowel QOL should be closely considered in selecting the right treatment, especially in evaluating the value of non-invasive treatments, such as SBRT

Evidence for Loss of a Partial Flagellar Glycolytic Pathway during Trypanosomatid Evolution

Classically viewed as a cytosolic pathway, glycolysis is increasingly recognized as a metabolic pathway exhibiting surprisingly wide-ranging variations in compartmentalization within eukaryotic cells. Trypanosomatid parasites provide an extreme view of glycolytic enzyme compartmentalization as several glycolytic enzymes are found exclusively in peroxisomes. Here, we characterize Trypanosoma brucei flagellar proteins resembling glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and phosphoglycerate kinase (PGK): we show the latter associates with the axoneme and the former is a novel paraflagellar rod component. The paraflagellar rod is an essential extra-axonemal structure in trypanosomes and related protists, providing a platform into which metabolic activities can be built. Yet, bioinformatics interrogation and structural modelling indicate neither the trypanosome PGK-like nor the GAPDH-like protein is catalytically active. Orthologs are present in a free-living ancestor of the trypanosomatids, Bodo saltans: the PGK-like protein from B. saltans also lacks key catalytic residues, but its GAPDH-like protein is predicted to be catalytically competent. We discuss the likelihood that the trypanosome GAPDH-like and PGK-like proteins constitute molecular evidence for evolutionary loss of a flagellar glycolytic pathway, either as a consequence of niche adaptation or the re-localization of glycolytic enzymes to peroxisomes and the extensive changes to glycolytic flux regulation that accompanied this re-localization. Evidence indicating loss of localized ATP provision via glycolytic enzymes therefore provides a novel contribution to an emerging theme of hidden diversity with respect to compartmentalization of the ubiquitous glycolytic pathway in eukaryotes. A possibility that trypanosome GAPDH-like protein additionally represents a degenerate example of a moonlighting protein is also discussed

Identification and studying of the role of new cytosolic factors implicated in tRNA import in yeast mitochondria

Chez S. cerevisiae un ARNtLys (appelé tRK1) est partiellement importé dans la mitochondrie. Une pré-protéine, préMsk1p, a été identifiée comme nécessaire mais non suffisante pour diriger cet adressage. L'objectif de la thèse était d'identifier de nouveauIn yeast S. cerevisiae, one ARNtLys (referred to as tRK1) is partially imported into mitochondria. One pre-protein, preMsk1p, was previously identified as necessary but not sufficient to direct the import. The aim of the thesis work was to identify new p

Identification and studying of the role of new cytosolic factors implicated in tRNA import in yeast mitochondria

Chez S. cerevisiae un ARNtLys (appelé tRK1) est partiellement importé dans la mitochondrie. Une pré-protéine, préMsk1p, a été identifiée comme nécessaire mais non suffisante pour diriger cet adressage. L'objectif de la thèse était d'identifier de nouveaux facteurs participant à l'importation d'ARNt dans les mitochondries de levure et d'étudier leur rôle dans ce processus. Un des facteurs d'import a été identifié comme un enzyme glycolytique, enolase. Cette découverte a permis de reconstruire un système minimal d'importation de tRK1 in vitro à partir de deux protéines recombinantes, préMsk1p et enolase, tRK1 et les mitochondries isolées.Trois protéines du système ubiquitine/protéasome (UPS) ont été identifiées par des cribles 2- et 3-hybride comme des facteurs potentiels d'import de tRK1. Une corrélation a été observée entre l'efficacité d'importation de tRK1 et l'activité protéolytique du protéasome, suggérant que l'UPS pourrait être impliqué dans la régulation de l'importation de tRK1.In yeast S. cerevisiae, one ARNtLys (referred to as tRK1) is partially imported into mitochondria. One pre-protein, preMsk1p, was previously identified as necessary but not sufficient to direct the import. The aim of the thesis work was to identify new protein factors of tRK1 mitochondrial import and characterisation of their precise role in this pathway.One import factor of tRK1 mitochondrial targeting was identified as a glycolytic enzyme, enolase. This finding permitted to reconstruct in vitro the import mechanism by using individual recombinant proteins (preMsk1p and enolase), tRK1 and mitochondria.Three proteins of ubiquitin/protéasome system (UPS) were identified by two and three-hybrid screenings for interactions with tRK1 or preMsk1 and are proposed as tRNA import factors. Direct correlation between efficiency of tRNA import and proteolytic activity of proteasome was observed. We therefore suggest that UPS is implicated in tRNA import regulation in yeast

Identification et rôles de nouveaux facteurs protéiques cytosoliques impliqués dans l'import d'ARNt dans les mitochondries de levure

Chez S. cerevisiae un ARNtLys (appelé tRK1) est partiellement importé dans la mitochondrie. Une pré-protéine, préMsk1p, a été identifiée comme nécessaire mais non suffisante pour diriger cet adressage. L objectif de la thèse était d identifier de nouveaux facteurs participant à l importation d ARNt dans les mitochondries de levure et d étudier leur rôle dans ce processus. Un des facteurs d'import a été identifié comme un enzyme glycolytique, enolase. Cette découverte a permis de reconstruire un système minimal d importation de tRK1 in vitro à partir de deux protéines recombinantes, préMsk1p et enolase, tRK1 et les mitochondries isolées.Trois protéines du système ubiquitine/protéasome (UPS) ont été identifiées par des cribles 2- et 3-hybride comme des facteurs potentiels d import de tRK1. Une corrélation a été observée entre l efficacité d importation de tRK1 et l activité protéolytique du protéasome, suggérant que l UPS pourrait être impliqué dans la régulation de l importation de tRK1.In yeast S. cerevisiae, one ARNtLys (referred to as tRK1) is partially imported into mitochondria. One pre-protein, preMsk1p, was previously identified as necessary but not sufficient to direct the import. The aim of the thesis work was to identify new protein factors of tRK1 mitochondrial import and characterisation of their precise role in this pathway.One import factor of tRK1 mitochondrial targeting was identified as a glycolytic enzyme, enolase. This finding permitted to reconstruct in vitro the import mechanism by using individual recombinant proteins (preMsk1p and enolase), tRK1 and mitochondria.Three proteins of ubiquitin/protéasome system (UPS) were identified by two and three-hybrid screenings for interactions with tRK1 or preMsk1 and are proposed as tRNA import factors. Direct correlation between efficiency of tRNA import and proteolytic activity of proteasome was observed. We therefore suggest that UPS is implicated in tRNA import regulation in yeast.STRASBOURG-Sc. et Techniques (674822102) / SudocSudocFranceRussian FederationFRR

Challenges and recommendations to improve implementation of phototherapy among neonates in Malawian hospitals

Background Severe neonatal jaundice can result in long term morbidities and mortality when left untreated. Phototherapy is the main-stay intervention for treating moderate jaundice and for prevention of the development of severe jaundice. However, in resource-limited health care settings, phototherapy has been inconsistently used. The objective of this study is to evaluate barriers and facilitators for phototherapy to treat neonatal jaundice at Malawian hospitals. Methods We conducted a convergent mixed-method study comprised of a facility assessment and qualitative interviews with healthcare workers and caregivers in southern Malawi. The facility assessment was conducted at three secondary-level hospitals in rural districts. In-depth interviews following a semi-structured topic guide were conducted at a district hospital and a tertiary-level hospital. Interviews were thematically analysed in NVivo 12 software (QSR International, Melbourne, Australia). Results The facility assessment found critical gaps in initiating and monitoring phototherapy in all facilities. Based on a total of 31 interviews, participants identified key challenges in diagnosing neonatal jaundice, counselling caregivers, and availability of infrastructure. Participants emphasized the need for transcutaneous bilirubinometers to guide treatment decisions. Caregivers were sometimes fearful of potential harmful effects of phototherapy, which required adequate explanation to mothers and family members in non-medical language. Task shifting and engaging peer support for caregivers with concerns about phototherapy was recommended. Conclusion Implementation of a therapeutic intervention is limited if accurate diagnostic tests are unavailable. The scale up of therapeutic interventions, such as phototherapy for neonatal jaundice, requires careful holistic attention to infrastructural needs, supportive services such as laboratory integration as well as trained human resources.Medicine, Faculty ofNon UBCObstetrics and Gynaecology, Department ofPathology and Laboratory Medicine, Department ofReviewedFacultyResearche