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

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Gallagher, Ferdia A 
Sladen, Helen 
Kettunen, Mikko I 
Serrao, Eva M 
Rodrigues, Tiago B 


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.



Animals, Antigens, Neoplasm, Bicarbonates, Carbon Dioxide, Carbon Isotopes, Carbonic Anhydrase IX, Carbonic Anhydrases, Cell Line, Tumor, Colorectal Neoplasms, Heterografts, Humans, Hydrogen-Ion Concentration, Magnetic Resonance Spectroscopy, Male, Mice, Mice, Inbred NOD, Mice, SCID, Neoplasm Proteins, Recombinant Fusion Proteins, Tumor Microenvironment

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Cancer Res

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American Association for Cancer Research (AACR)
Cancer Research Uk (None)
Cancer Research Uk (None)
Medical Research Council (G1000265)
Cancer Research Uk (None)
Cancer Research UK (CB4100)
Cancer Research UK (C14303/A17197)
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.