Direct enhancement of nuclear singlet order by dynamic nuclear polarization

Hyperpolarized singlet order is available immediately after dissolution DNP, avoiding need for additional preparation steps. We demonstrate this procedure on a sample of [1,2–13C2]pyruvic aci

Effects of fasting on serial measurements of hyperpolarized [1-(13) C]pyruvate metabolism in tumors.

Imaging of the metabolism of hyperpolarized [1-(13) C]pyruvate has shown considerable promise in preclinical studies in oncology, particularly for the assessment of early treatment response. The repeatability of measurements of (13) C label exchange between pyruvate and lactate was determined in a murine lymphoma model in fasted and non-fasted animals. The fasted state showed lower intra-individual variability, although the [1-(13) C]lactate/[1-(13) C]pyruvate signal ratio was significantly greater in fasted than in non-fasted mice, which may be explained by the higher tumor lactate concentrations in fasted animals. These results indicate that the fasted state may be preferable for the measurement of (13) C label exchange between pyruvate and lactate, as it reduces the variability and therefore should make it easier to detect the effects of therapy. © 2016 The Authors. NMR in Biomedicine published by John Wiley & Sons Ltd.Cancer Research UK (Grant ID: 17242)This is the final version of the article. It first appeared from Wiley via https://doi.org/10.1002/nbm.356

Hyperpolarized [1-13C]-ascorbic and dehydroascorbic acid: vitamin C as a probe for imaging redox status in vivo.

Dynamic nuclear polarization (DNP) of (13)C-labeled metabolic substrates in vitro and their subsequent intravenous administration allow both the location of the hyperpolarized substrate and the dynamics of its subsequent conversion into other metabolic products to be detected in vivo. We report here the hyperpolarization of [1-(13)C]-ascorbic acid (AA) and [1-(13)C]-dehydroascorbic acid (DHA), the reduced and oxidized forms of vitamin C, respectively, and evaluate their performance as probes of tumor redox state. Solution-state polarization of 10.5 ± 1.3% was achieved for both forms at pH 3.2, whereas at pH 7.0, [1-(13)C]-AA retained polarization of 5.1 ± 0.6% and [1-(13)C]-DHA retained 8.2 ± 1.1%. The spin-lattice relaxation times (T(1)'s) for these labeled nuclei are long at 9.4 T: 15.9 ± 0.7 s for AA and 20.5 ± 0.9 s for DHA. Extracellular oxidation of [1-(13)C]-AA and intracellular reduction of [1-(13)C]-DHA were observed in suspensions of murine lymphoma cells. The spontaneous reaction of DHA with the cellular antioxidant glutathione was monitored in vitro and was approximately 100-fold lower than the rate observed in cell suspensions, indicating enzymatic involvement in the intracellular reduction. [1-(13)C]-DHA reduction was also detected in lymphoma tumors in vivo. In contrast, no detectable oxidation of [1-(13)C]-AA was measured in the same tumors, consistent with the notion that tumors maintain a reduced microenvironment. This study demonstrates that hyperpolarized (13)C-labeled vitamin C could be used as a noninvasive biomarker of redox status in vivo, which has the potential to translate to the clinic

Alcohol Co-Administration Changes Mephedrone-Induced Alterations of Neuronal Activity

Mephedrone (4-MMC), despite its illegal status, is still a widely used psychoactive substance. Its effects closely mimic those of the classical stimulant drug methamphetamine (METH). Recent research suggests that unlike METH, 4-MMC is not neurotoxic on its own. However, the neurotoxic effects of 4-MMC may be precipitated under certain circumstances, such as administration at high ambient temperatures. Common use of 4-MMC in conjunction with alcohol raises the question whether this co-consumption could also precipitate neurotoxicity. A total of six groups of adolescent rats were treated twice daily for four consecutive days with vehicle, METH (5 mg/kg) or 4-MMC (30 mg/kg), with or without ethanol (1.5 g/kg). To investigate persistent delayed effects of the administrations at two weeks after the final treatments, manganese-enhanced magnetic resonance imaging brain scans were performed. Following the scans, brains were collected for Golgi staining and spine analysis. 4-MMC alone had only subtle effects on neuronal activity. When administered with ethanol, it produced a widespread pattern of deactivation, similar to what was seen with METH-treated rats. These effects were most profound in brain regions which are known to have high dopamine and serotonin activities including hippocampus, nucleus accumbens and caudate-putamen. In the regions showing the strongest activation changes, no morphological changes were observed in spine analysis. By itself 4-MMC showed few long-term effects. However, when co-administered with ethanol, the apparent functional adaptations were profound and comparable to those of neurotoxic METH.Peer reviewe

(13) C magnetic resonance spectroscopy measurements with hyperpolarized [1-(13) C] pyruvate can be used to detect the expression of transgenic pyruvate decarboxylase activity in vivo.

PURPOSE: Dissolution dynamic nuclear polarization can increase the sensitivity of the (13) C magnetic resonance spectroscopy experiment by at least four orders of magnitude and offers a novel approach to the development of MRI gene reporters based on enzymes that metabolize (13) C-labeled tracers. We describe here a gene reporter based on the enzyme pyruvate decarboxylase (EC 4.1.1.1), which catalyzes the decarboxylation of pyruvate to produce acetaldehyde and carbon dioxide. METHODS: Pyruvate decarboxylase from Zymomonas mobilis (zmPDC) and a mutant that lacked enzyme activity were expressed using an inducible promoter in human embryonic kidney (HEK293T) cells. Enzyme activity was measured in the cells and in xenografts derived from the cells using (13) C MRS measurements of the conversion of hyperpolarized [1-(13) C] pyruvate to H(13) CO3-. RESULTS: Induction of zmPDC expression in the cells and in the xenografts derived from them resulted in an approximately two-fold increase in the H(13) CO3-/[1-(13) C] pyruvate signal ratio following intravenous injection of hyperpolarized [1-(13) C] pyruvate. CONCLUSION: We have demonstrated the feasibility of using zmPDC as an in vivo reporter gene for use with hyperpolarized (13) C MRS. Magn Reson Med 76:391-401, 2016. © 2015 The Authors. Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance in Medicine. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.PD was in receipt of a studentship funded by CRUK and S.-S.T. a Yousef Jameel studentship. TBR was in receipt of an Intra-European Marie Curie (FP7-PEOPLE- 2009-IEF, Imaging Lymphoma) and Long-term EMBO (EMBO-ALT-1145-2009) fellowships and E.M.S. and I.M.R were recipients of fellowships from the European Union Seventh Framework Programme (FP7/2007-2013) under the Marie Curie Initial Training Network METAFLUX (project number 264780). E.M.S. also acknowledges the educational support of Programme for Advanced Medical Education from Calouste Gulbenkian Foundation, Champalimaud Foundation, Ministerio de Saude and Fundacao para a Ciencia e Tecnologia, Portugal. The work was supported by a CRUK Programme Grant (17242) to KMB. The polarizer and related materials were provided by GE-Healthcare.This is the final version of the article. It first appeared from Wiley via http://dx.doi.org/10.1002/mrm.2587

Carbonic Anhydrase Activity Monitored In Vivo by Hyperpolarized 13C-Magnetic Resonance Spectroscopy Demonstrates Its Importance for pH Regulation in Tumors.

Carbonic anhydrase buffers tissue pH by catalyzing the rapid interconversion of carbon dioxide (CO2) and bicarbonate (HCO3 (-)). We assessed the functional activity of CAIX in two colorectal tumor models, expressing different levels of the enzyme, by measuring the rate of exchange of hyperpolarized (13)C label between bicarbonate (H(13)CO3(-)) and carbon dioxide ((13)CO2), following injection of hyperpolarized H(13)CO3(-), using (13)C-magnetic resonance spectroscopy ((13)C-MRS) magnetization transfer measurements. (31)P-MRS measurements of the chemical shift of the pH probe, 3-aminopropylphosphonate, and (13)C-MRS measurements of the H(13)CO3(-)/(13)CO2 peak intensity ratio showed that CAIX overexpression lowered extracellular pH in these tumors. However, the (13)C measurements overestimated pH due to incomplete equilibration of the hyperpolarized (13)C label between the H(13)CO3(-) and (13)CO2 pools. Paradoxically, tumors overexpressing CAIX showed lower enzyme activity using magnetization transfer measurements, which can be explained by the more acidic extracellular pH in these tumors and the decreased activity of the enzyme at low pH. This explanation was confirmed by administration of bicarbonate in the drinking water, which elevated tumor extracellular pH and restored enzyme activity to control levels. These results suggest that CAIX expression is increased in hypoxia to compensate for the decrease in its activity produced by a low extracellular pH and supports the hypothesis that a major function of CAIX is to lower the extracellular pH.The authors acknowledge funding support from Cancer Research UK (CRUK; C19212/A16628; C19212/A911376), the National Institute for Health Research Cambridge Biomedical Research Centre and the School of Clinical Medicine at the University of Cambridge, the CRUK and Engineering and Physical Sciences Research Council (EPSRC) Cancer Imaging Centre in Cambridge and Manchester. E.M.S. is a recipient of funding from the European Union Seventh Framework Programme (FP7/2007-2013) under the Marie Curie Initial Training Network METAFLUX and has support from the Calouste Gulbenkian Foundation, Champalimaud Foundation, Ministerio de Saude and Fundacao para a Ciencia e Tecnologia, Portugal.This is the author accepted manuscript. The final version is available from American Association for Cancer Research via http://dx.doi.org/10.1158/0008-5472.CAN-15-085

Magnetic resonance imaging with hyperpolarized [1-13C]pyruvate detects advanced pancreatic preneoplasia prior to invasive disease in a mouse model

OBJECTIVES: Pancreatic cancer (PCa) is treatable by surgery when detected at an early stage. Non-invasive imaging methods able to detect both established tumours and their precursor lesions are needed to select patients for surgery. We investigated here whether pancreatic preneoplasia could be detected prior to the development of invasive cancers in genetically engineered mouse models of PCa using metabolic imaging. DESIGN: The concentrations of alanine and lactate and the activities of lactate dehydrogenase (LDH) and alanine aminotransferase (ALT) were measured in extracts prepared from the pancreas of animals at different stages of disease progression; from pancreatitis, through tissue with predominantly low-grade and then high-grade pancreatic intraepithelial neoplasia and then tumour. (13)C magnetic resonance spectroscopic imaging ((13)C-MRSI) was used to measure non-invasively changes in (13)C labelling of alanine and lactate with disease progression, following injection of hyperpolarised [1-(13)C]pyruvate. RESULTS: Progressive decreases in the alanine/lactate concentration ratio and ALT/LDH activity ratio with disease progression were accompanied by a corresponding decrease in the [1-(13)C]alanine/[1-(13)C]lactate signal ratio observed in (13)C-MRSI images of the pancreas. CONCLUSIONS: Metabolic imaging with hyperpolarised [1-(13)C]pyruvate enables detection and monitoring of the progression of PCa precursor lesions. Translation of this MRI technique to the clinic has the potential to improve the management of patients at high risk of developing PCa.The work was supported by a Cancer Research UK Programme grant (17242) to K.M.B.. E.M.S. is a recipient of a fellowship from the European Union Seventh Framework Programme (FP7/2007-2013) under the Marie Curie Initial Training Network METAFLUX (project number 264780). T.B.R. is a recipient of an Intra- European Marie Curie (FP7-PEOPLE-2009-IEF, Imaging Lymphoma) fellowship and a Long-term European Molecular Biology Organization (EMBO-ALT-1145-2009) fellowship. E.M.S. and J.A. acknowledge the educational support of Programme for Advanced Medical Education from Calouste Gulbenkian Foundation, Champalimaud Foundation, Ministerio de Saude and Fundacao para a Ciencia e Tecnologia, Portugal. The polarizer and related materials were provided by GE Healthcare. The polarimeter was provided by NIHR Cambridge Biomedical Centre. The laboratory is a member of and receives support from the Cancer Research UK & Engineering and Physical Science Research Council Cancer Imaging Center in Cambridge and Manchester. The authors would also like to acknowledge Dr. Judit Espana, Dr. Athena Matakidou, Dr. Madhu Basetti, Dr. Jose Sandoval and Sarah McGuire for their help with experiments as well as the Tumor Models Core of Cancer Research UK-Cambridge Institute.This is the final version of the article. It first appeared from BMJ Group via http://dx.doi.org/10.1136/gutjnl-2015-31011

A comparison between radiolabeled fluorodeoxyglucose uptake and hyperpolarized (13)C-labeled pyruvate utilization as methods for detecting tumor response to treatment.

Published versio

CuII(atsm) Attenuates Neuroinflammation

Background: Neuroinflammation and biometal dyshomeostasis are key pathological features of several neurodegenerative diseases, including Alzheimer’s disease (AD). Inflammation and biometals are linked at the molecular level through regulation of metal buffering proteins such as the metallothioneins. Even though the molecular connections between metals and inflammation have been demonstrated, little information exists on the effect of copper modulation on brain inflammation.Methods: We demonstrate the immunomodulatory potential of the copper bis(thiosemicarbazone) complex CuII(atsm) in an neuroinflammatory model in vivo and describe its anti-inflammatory effects on microglia and astrocytes in vitro.Results: By using a sophisticated in vivo magnetic resonance imaging (MRI) approach, we report the efficacy of CuII(atsm) in reducing acute cerebrovascular inflammation caused by peripheral administration of bacterial lipopolysaccharide (LPS). CuII(atsm) also induced anti-inflammatory outcomes in primary microglia [significant reductions in nitric oxide (NO), monocyte chemoattractant protein 1 (MCP-1), and tumor necrosis factor (TNF)] and astrocytes [significantly reduced NO, MCP-1, and interleukin 6 (IL-6)] in vitro. These anti-inflammatory actions were associated with increased cellular copper levels and increased the neuroprotective protein metallothionein-1 (MT1) in microglia and astrocytes.Conclusion: The beneficial effects of CuII(atsm) on the neuroimmune system suggest copper complexes are potential therapeutics for the treatment of neuroinflammatory conditions

Direct Enhancement of Nuclear Singlet Order by Dynamic Nuclear Polarization

Hyperpolarized singlet order is available immediately after dissolution DNP, avoiding need for additional preparation steps. We demonstrate this procedure on a sample of [1,2–13C2]pyruvic aci