Analysis of Tumor Metabolism Reveals Mitochondrial Glucose Oxidation in Genetically Diverse Human Glioblastomas in the Mouse Brain In Vivo

SummaryDysregulated metabolism is a hallmark of cancer cell lines, but little is known about the fate of glucose and other nutrients in tumors growing in their native microenvironment. To study tumor metabolism in vivo, we used an orthotopic mouse model of primary human glioblastoma (GBM). We infused 13C-labeled nutrients into mice bearing three independent GBM lines, each with a distinct set of mutations. All three lines displayed glycolysis, as expected for aggressive tumors. They also displayed unexpected metabolic complexity, oxidizing glucose via pyruvate dehydrogenase and the citric acid cycle, and using glucose to supply anaplerosis and other biosynthetic activities. Comparing the tumors to surrounding brain revealed obvious metabolic differences, notably the accumulation of a large glutamine pool within the tumors. Many of these same activities were conserved in cells cultured ex vivo from the tumors. Thus GBM cells utilize mitochondrial glucose oxidation during aggressive tumor growth in vivo

Taxonomic revaluation of the Ahaetulla prasina (H. Boie in F. Boie, 1827) complex from Northeast India: resurrection and redescription of Ahaetulla flavescens (Wall, 1910) (Reptilia: Serpentes: Colubridae)

The taxonomic status of the nominal taxon Dryophis prasinus flavescens Wall, 1910 is reevaluated herein. Based on molecular data generated from fresh collections of Ahaetulla prasina (H. Boie in F. Boie, 1827) auctorum from Northeast India and, additionally, morphological data from museum specimens originating from the same areas, we resurrect this taxon as Ahaetulla flavescens (Wall, 1910) comb. nov. We clarify the status, identity and locations of its type specimens, rediscover, redescribe and illustrate those specimens and also designate a lectotype in order to effect a proper taxonomic redefinition of this nominal taxon. We provide further details on the morphology and diagnosis of this species and elucidate its phylogenetic position. We also provide a summary of the natural history and distribution of this species. Adding to the known cryptic diversity and genetic divergence within Southeast Asian populations, this work also hints at the need for a taxonomic revision of the A. prasina complex. This work complements a previous study on the A. prasina complex focusing on populations in Indonesia. Taken together, these two studies represent phylogenetic reconstructions from different populations of the A. prasina complex across its distribution range, on the Asian mainland and the surrounding islands

Evaluating the efficacy and safety of human anti-SARS-CoV-2 convalescent plasma in severely ill adults with COVID-19: A structured summary of a study protocol for a randomized controlled trial

Objectives The aim of this study is to evaluate the efficacy and safety of human anti-SARS-CoV-2 convalescent plasma in hospitalized adults with severe SARS-CoV-2 infection. Trial Design This is a prospective, single-center, phase 2, randomized, controlled trial that is blinded to participants and clinical outcome assessor. Participants Eligible participants include adults (≥ 18 years) with evidence of SARS-CoV-2 infection by PCR test of nasopharyngeal or oropharyngeal swab within 14 days of randomization, evidence of infiltrates on chest radiography, peripheral capillary oxygen saturation (SpO2) ≤ 94% on room air, and/or need for supplemental oxygen, non-invasive mechanical ventilation, or invasive mechanical ventilation, who are willing and able to provide written informed consent prior to performing study procedures or who have a legally authorized representative available to do so. Exclusion criteria include participation in another clinical trial of anti-viral agent(s)* for coronavirus disease-2019 (COVID-19), receipt of any anti-viral agent(s)* with possible activity against SARS-CoV-2 <24 hours prior to plasma infusion, mechanical ventilation (including extracorporeal membrane oxygenation [ECMO]) for ≥ 5 days, severe multi-organ failure, history of allergic reactions to transfused blood products per NHSN/CDC criteria, known IgA deficiency, and pregnancy. Included participants will be hospitalized at the time of randomization and plasma infusion. *Use of remdesivir as treatment for COVID-19 is permitted. The study will be undertaken at Columbia University Irving Medical Center in New York, USA. Intervention and comparator The investigational treatment is anti-SARS-CoV-2 human convalescent plasma. To procure the investigational treatment, volunteers who recovered from COVID-19 will undergo testing to confirm the presence of anti-SARS-CoV-2 antibody to the spike trimer at a 1:400 dilution. Donors will also be screened for transfusion-transmitted infections (e.g. HIV, HBV, HCV, WNV, HTLV-I/II, T. cruzi, ZIKV). If donors have experienced COVID-19 symptoms within 28 days, they will be screened with a nasopharyngeal swab to confirm they are SARS-CoV-2 PCR-negative. Plasma will be collected using standard apheresis technology by the New York Blood Center. Study participants will be randomized in a 2:1 ratio to receive one unit (200 – 250 mL) of anti-SARS-CoV-2 plasma versus one unit (200 – 250 mL) of the earliest available control plasma. The control plasma cannot be tested for presence of anti-SARS-CoV-2 antibody prior to the transfusion, but will be tested for anti- SARS-CoV-2 antibody after the transfusion to allow for a retrospective per-protocol analysis. Main outcomes The primary endpoint is time to clinical improvement. This is defined as time from randomization to either discharge from the hospital or improvement by one point on the following seven-point ordinal scale, whichever occurs first. 1. Not hospitalized with resumption of normal activities 2. Not hospitalized, but unable to resume normal activities 3. Hospitalized, not requiring supplemental oxygen 4. Hospitalized, requiring supplemental oxygen 5. Hospitalized, requiring high-flow oxygen therapy or non-invasive mechanical ventilation 6. Hospitalized, requiring ECMO, invasive mechanical ventilation, or both 7. Death This scale, designed to assess clinical status over time, was based on that recommended by the World Health Organization for use in determining efficacy end-points in clinical trials in hospitalized patients with COVID-19. A recent clinical trial evaluating the efficacy and safety of lopinavir- ritonavir for patients hospitalized with severe COVID-19 used a similar ordinal scale, as have recent clinical trials of novel therapeutics for severe influenza, including a post-hoc analysis of a trial evaluating immune plasma. The primary safety endpoints are cumulative incidence of grade 3 and 4 adverse events and cumulative incidence of serious adverse events during the study period. Randomization Study participants will be randomized in a 2:1 ratio to receive anti-SARS-CoV-2 plasma versus control plasma using a web-based randomization platform. Treatment assignments will be generated using randomly permuted blocks of different sizes to minimize imbalance while also minimizing predictability. Blinding (masking) The study participants and the clinicians who will evaluate post-treatment outcomes will be blinded to group assignment. The blood bank and the clinical research team will not be blinded to group assignment. Numbers to be randomized (sample size) We plan to enroll 129 participants, with 86 in the anti-SARS-CoV-2 arm, and 43 in the control arm. Among the participants, we expect ~70% or n = 72 will achieve clinical improvement. This will yield an 80% power for a one-sided Wald test at 0.15 level of significance under the proportional hazards model with a hazard ratio of 1.5. Trial Status Protocol AAAS9924, Version 17APR2020, 4/17/2020 Start of recruitment: April 20, 2020 Recruitment is ongoing. Trial registration ClinicalTrials.gov: NCT04359810 Date of trial registration: April 24, 2020 Retrospectively registered Full protocol The full protocol is attached as an additional file, accessible from the Trials website (Additional file 1). In the interest of expediting dissemination of this material, the familiar formatting has been eliminated; this Letter serves as a summary of the key elements of the full protocol

Haem oxygenase is synthetically lethal with the tumour suppressor fumarate hydratase

Fumarate hydratase (FH) is an enzyme of the tricarboxylic acid cycle (TCA cycle) that catalyses the hydration of fumarate into malate. Germline mutations of FH are responsible for hereditary leiomyomatosis and renal-cell cancer (HLRCC)1. It has previously been demonstrated that the absence of FH leads to the accumulation of fumarate, which activates hypoxia-inducible factors (HIFs) at normal oxygen tensions2, 3, 4. However, so far no mechanism that explains the ability of cells to survive without a functional TCA cycle has been provided. Here we use newly characterized genetically modified kidney mouse cells in which Fh1 has been deleted, and apply a newly developed computer model of the metabolism of these cells to predict and experimentally validate a linear metabolic pathway beginning with glutamine uptake and ending with bilirubin excretion from Fh1-deficient cells. This pathway, which involves the biosynthesis and degradation of haem, enables Fh1-deficient cells to use the accumulated TCA cycle metabolites and permits partial mitochondrial NADH production. We predicted and confirmed that targeting this pathway would render Fh1-deficient cells non-viable, while sparing wild-type Fh1-containing cells. This work goes beyond identifying a metabolic pathway that is induced in Fh1-deficient cells to demonstrate that inhibition of haem oxygenation is synthetically lethal when combined with Fh1 deficiency, providing a new potential target for treating HLRCC patients.demonstrated that the absence of FH leads to the accumulation of fumarate, which activates hypoxia-inducible factors (HIFs) at normal oxygen tensions. However, so far no mechanism that explains the ability of cells to survive without a functional TCA cycle has been provided. Here we use newly characterized genetically modified kidney mouse cells in which Fh1 has been deleted, and apply a newly developed computer model of the metabolism of these cells to predict and experimentally validate a linear metabolic pathway beginning with glutamine uptake and ending with bilirubin excretion from Fh1-deficient cells. This pathway, which involves the biosynthesis and degradation of haem, enables Fh1-deficient cells to use the accumulated TCA cycle metabolites and permits partial mitochondrial NADH production. We predicted and confirmed that targeting this pathway would render Fh1-deficient cells non-viable, while sparing wild-type Fh1-containing cells. This work goes beyond identifying a metabolic pathway that is induced in Fh1-deficient cells to demonstrate that inhibition of haem oxygenation is synthetically lethal when combined with Fh1 deficiency, providing a new potential target for treating HLRCC patients