A genome-wide cytotoxicity screen of cluster F1 mycobacteriophage Girr reveals novel inhibitors of Mycobacterium smegmatis growth.

Over the past decade, thousands of bacteriophage genomes have been sequenced and annotated. A striking observation from this work is that known structural features and functions cannot be assigned for >65% of the encoded proteins. One approach to begin experimentally elucidating the function of these uncharacterized gene products is genome-wide screening to identify phage genes that confer phenotypes of interest like inhibition of host growth. This study describes the results of a screen evaluating the effects of overexpressing each gene encoded by the temperate Cluster F1 mycobacteriophage Girr on the growth of the host bacterium Mycobacterium smegmatis. Overexpression of 29 of the 102 Girr genes (~28% of the genome) resulted in mild to severe cytotoxicity. Of the 29 toxic genes described, 12 have no known function and are predominately small proteins of <125 amino acids. Overexpression of the majority of these 12 cytotoxic no known functions proteins resulted in moderate to severe growth reduction and represent novel antimicrobial products. The remaining 17 toxic genes have predicted functions, encoding products involved in phage structure, DNA replication/modification, DNA binding/gene regulation, or other enzymatic activity. Comparison of this dataset with prior genome-wide cytotoxicity screens of mycobacteriophages Waterfoul and Hammy reveals some common functional themes, though several of the predicted Girr functions associated with cytotoxicity in our report, including genes involved in lysogeny, have not been described previously. This study, completed as part of the HHMI-supported SEA-GENES project, highlights the power of parallel, genome-wide overexpression screens to identify novel interactions between phages and their hosts.

Genome Sequence of VanLee, a Singleton Actinobacteriophage That Infects Multiple Gordonia Strains.

VanLee is a singleton phage that was isolated from soil in Florida using Gordonia rubripertincta NRRL B-16540 as the host. The genome is 84,560 bp and has a GC content of 67.8%. VanLee has 164 predicted protein-coding genes and one tRNA. VanLee can infect Gordonia terrae with the same efficiency as G. rubripertincta.

Fostering Engagement With Health and Housing Innovation: Development of Participant Personas in a Social Housing Cohort.

BACKGROUND: Personas, based on customer or population data, are widely used to inform design decisions in the commercial sector. The variety of methods available means that personas can be produced from projects of different types and scale. OBJECTIVE: This study aims to experiment with the use of personas that bring together data from a survey, household air measurements and electricity usage sensors, and an interview within a research and innovation project, with the aim of supporting eHealth and eWell-being product, process, and service development through broadening the engagement with and understanding of the data about the local community. METHODS: The project participants were social housing residents (adults only) living in central Cornwall, a rural unitary authority in the United Kingdom. A total of 329 households were recruited between September 2017 and November 2018, with 235 (71.4%) providing complete baseline survey data on demographics, socioeconomic position, household composition, home environment, technology ownership, pet ownership, smoking, social cohesion, volunteering, caring, mental well-being, physical and mental health-related quality of life, and activity. K-prototype cluster analysis was used to identify 8 clusters among the baseline survey responses. The sensor and interview data were subsequently analyzed by cluster and the insights from all 3 data sources were brought together to produce the personas, known as the Smartline Archetypes. RESULTS: The Smartline Archetypes proved to be an engaging way of presenting data, accessible to a broader group of stakeholders than those who accessed the raw anonymized data, thereby providing a vehicle for greater research engagement, innovation, and impact. CONCLUSIONS: Through the adoption of a tool widely used in practice, research projects could generate greater policy and practical impact, while also becoming more transparent and open to the public.

AZD2014, an Inhibitor of mTORC1 and mTORC2, Is Highly Effective in ER+ Breast Cancer When Administered Using Intermittent or Continuous Schedules.

mTOR is an atypical serine threonine kinase involved in regulating major cellular functions, such as nutrients sensing, growth, and proliferation. mTOR is part of the multiprotein complexes mTORC1 and mTORC2, which have been shown to play critical yet functionally distinct roles in the regulation of cellular processes. Current clinical mTOR inhibitors only inhibit the mTORC1 complex and are derivatives of the macrolide rapamycin (rapalogs). Encouraging effects have been observed with rapalogs in estrogen receptor-positive (ER(+)) breast cancer patients in combination with endocrine therapy, such as aromatase inhibitors. AZD2014 is a small-molecule ATP competitive inhibitor of mTOR that inhibits both mTORC1 and mTORC2 complexes and has a greater inhibitory function against mTORC1 than the clinically approved rapalogs. Here, we demonstrate that AZD2014 has broad antiproliferative effects across multiple cell lines, including ER(+) breast models with acquired resistance to hormonal therapy and cell lines with acquired resistance to rapalogs. In vivo, AZD2014 induces dose-dependent tumor growth inhibition in several xenograft and primary explant models. The antitumor activity of AZD2014 is associated with modulation of both mTORC1 and mTORC2 substrates, consistent with its mechanism of action. In combination with fulvestrant, AZD2014 induces tumor regressions when dosed continuously or using intermittent dosing schedules. The ability to dose AZD2014 intermittently, together with its ability to block signaling from both mTORC1 and mTORC2 complexes, makes this compound an ideal candidate for combining with endocrine therapies in the clinic. AZD2014 is currently in phase II clinical trials.

Bedside ultrasound maximizes patient satisfaction.

BACKGROUND: Bedside ultrasound (US) is associated with improved patient satisfaction, perhaps as a consequence of improved time to diagnosis and decreased length of stay (LOS). OBJECTIVES: Our study aimed to quantify the association between beside US and patient satisfaction and to assess patient attitudes toward US and perception of their interaction with the clinician performing the examination. METHODS: We enrolled a convenience sample of adult patients who received a bedside US. The control group had similar LOS and presenting complaints but did not have a bedside US. Both groups answered survey questions during their emergency department (ED) visit and again by telephone 1 week later. The questionnaire assessed patient perceptions and satisfaction on a 5-point Likert scale. RESULTS: Seventy patients were enrolled over 10 months. The intervention group had significantly higher scores on overall ED satisfaction (4.69 vs. 4.23; mean difference 0.46; 95% confidence interval [CI] 0.17-0.75), diagnostic testing (4.54 vs. 4.09; mean difference 0.46; 95% CI 0.16-0.76), and skills/abilities of the emergency physician (4.77 vs. 4.14; mean difference 0.63; 95% CI 0.29-0.96). A trend to higher scores for the intervention group persisted on follow-up survey. CONCLUSIONS: Patients who had a bedside US had statistically significant higher satisfaction scores with overall ED care, diagnostic testing, and with their perception of the emergency physician. Bedside US has the potential not only to expedite care and diagnosis, but also to maximize satisfaction scores and improve the patient-physician relationship, which has increasing relevance to health care organizations and hospitals that rely on satisfaction surveys.

Helicopter emergency medical services crew administration of antibiotics for open fractures.

This study had 3 major aims: (1) to ascertain the degree to which helicopter emergency medical services (HEMS) administration of antibiotics (Abx) can streamline the time to Abx in open fracture patients, (2) to determine whether any clinical outcome improvements were associated with HEMS Abx therapy, and (3) to calculate the cost-effectiveness of prehospital HEMS Abx. The design of the study was a prospective, nonrandomized, nonintervention, natural study of timing and clinical outcomes for patients with suspected open extremity fracture. There were 138 scene trauma cases transported by 8 participating HEMS programs from July 2009 to June 2010. The participating HEMS programs were both urban and rural. The diagnosis of an open fracture by the HEMS crews had an accuracy rate of 97.8% (95% confidence interval, 90.8%-98.4%). The time from the incident to Abx was 30 minutes shorter (P = .0001) when Abx were administered by HEMS crews. There was no statistical significance (P = 1.0) regarding the endpoint of infection or nonunion development in HEMS- versus hospital-administered Abx. In conclusion, the administration of Abx by HEMS crews to patients diagnosed with open extremity fractures is feasible, it may decrease the time to Abx by 30 minutes, and the effect magnitude (40.3% relative risk reduction) was promising.

Vandetanib inhibits both VEGFR-2 and EGFR signalling at clinically relevant drug levels in preclinical models of human cancer.

Vandetanib is a multi-targeted receptor tyrosine kinase inhibitor that is in clinical development for the treatment of solid tumours. This preclinical study examined the inhibition of two key signalling pathways (VEGFR-2, EGFR) at drug concentrations similar to those achieved in the clinic, and their contribution to direct and indirect antitumour effects of vandetanib. For in vitro studies, receptor phosphorylation was assessed by Western blotting and ELISA, cell proliferation was assessed using a cell viability endpoint, and effects on cell cycle determined using flow cytometry. For in vivo studies, Western blotting, ELISA and immunohistochemistry (IHC) were used to assess receptor phosphorylation. Cell culture experiments demonstrated that anti-proliferative effects of vandetanib resulted from inhibition of either EGFR or VEGFR-2 signalling in endothelial cells, but were associated with inhibition of EGFR signalling in tumour cells. Vandetanib inhibited both EGFR and VEGFR-2 signalling in normal lung tissue and in tumour xenografts. In a lung cancer model expressing an activating EGFR mutation, the activity of vandetanib was similar to that of a highly selective EGFR inhibitor (gefitinib), and markedly greater than that of a highly selective VEGFR inhibitor (vatalanib). These data suggest that at the plasma exposures achieved in the clinic, vandetanib will significantly inhibit both VEGFR-2 and EGFR signalling, and that both inhibition of angiogenesis and direct inhibition of tumour cell growth can contribute to treatment response.

Benefits of mTOR kinase targeting in oncology: pre-clinical evidence with AZD8055.

AZD8055 is a small-molecule inhibitor of mTOR (mammalian target of rapamycin) kinase activity. The present review highlights molecular and phenotypic differences between AZD8055 and allosteric inhibitors of mTOR such as rapamycin. Biomarkers, some of which are applicable to clinical studies, as well as biological effects such as autophagy, growth inhibition and cell death are compared between AZD8055 and rapamycin. Potential ways to develop rational combinations with mTOR kinase inhibitors are also discussed. Overall, AZD8055 may provide a better therapeutic strategy than rapamycin and analogues.

AZD8055 is a potent, selective, and orally bioavailable ATP-competitive mammalian target of rapamycin kinase inhibitor with in vitro and in vivo antitumor activity.

The mammalian target of rapamycin (mTOR) kinase forms two multiprotein complexes, mTORC1 and mTORC2, which regulate cell growth, cell survival, and autophagy. Allosteric inhibitors of mTORC1, such as rapamycin, have been extensively used to study tumor cell growth, proliferation, and autophagy but have shown only limited clinical utility. Here, we describe AZD8055, a novel ATP-competitive inhibitor of mTOR kinase activity, with an IC50 of 0.8 nmol/L. AZD8055 showed excellent selectivity (approximately 1,000-fold) against all class I phosphatidylinositol 3-kinase (PI3K) isoforms and other members of the PI3K-like kinase family. Furthermore, there was no significant activity against a panel of 260 kinases at concentrations up to 10 micromol/L. AZD8055 inhibits the phosphorylation of mTORC1 substrates p70S6K and 4E-BP1 as well as phosphorylation of the mTORC2 substrate AKT and downstream proteins. The rapamycin-resistant T37/46 phosphorylation sites on 4E-BP1 were fully inhibited by AZD8055, resulting in significant inhibition of cap-dependent translation. In vitro, AZD8055 potently inhibits proliferation and induces autophagy in H838 and A549 cells. In vivo, AZD8055 induces a dose-dependent pharmacodynamic effect on phosphorylated S6 and phosphorylated AKT at plasma concentrations leading to tumor growth inhibition. Notably, AZD8055 results in significant growth inhibition and/or regression in xenografts, representing a broad range of human tumor types. AZD8055 is currently in phase I clinical trials.