title: Assessment of chloroquine treatment for modulating autophagy flux in brain of WT and HD mice.
abstract: BACKGROUND
  Increasing mutant huntingtin (mHTT) clearance through the autophagy pathway may be a way to treat Huntington's disease (HD). Tools to manipulate and measure autophagy flux in brain in vivo are not well established.
  
  
  OBJECTIVE
  To examine the in vivo pharmacokinetics and pharmacodynamics of the lysosomal inhibitor chloroquine (CQ) and the levels of selected autophagy markers to determine usefulness of CQ as a tool to study autophagy flux in brain.
  
  
  METHODS
  Intraperitoneal injections of CQ were administered to WT and HD(Q175/Q175) mice. CQ levels were measured by LC-MS/MS in WT brain, muscle and blood at 4 to 24 hours after the last dose. Two methods of tissue preparation were used to detect by Western blot levels of the macroautophagy markers LC3 II and p62, the chaperone mediated autophagy receptor LAMP-2A and the late endosome/lysosomal marker RAB7.
  
  
  RESULTS
  Following peripheral administration, CQ levels were highest in muscle and declined rapidly between 4 and 24 hours. In the brain, CQ levels were greater in the cortex than striatum, and levels persisted up to 24 hours post-injection. CQ treatment induced changes in LC3 II and p62 that were variable across regions and tissue preparations. HD(Q175/Q175) mice exposed to CQ had variable but diminished levels of LC3 II, p62 and LAMP-2A, and increased levels of RAB7. Higher levels of mHTT were found in the membrane compartment of CQ treated HD mice.
  
  
  CONCLUSIONS
  Our findings suggest that the response of brain to CQ treatment, a blocker of autophagy flux, is variable and not as robust as it has been demonstrated in vitro, suggesting that CQ treatment has limitations for modulating autophagy flux in vivo. Alternative methods, compounds, and technologies need to be developed to further investigate autophagy flux in vivo, especially in the brain.
authors: Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA, USA.
authors: Petr
authors: P
authors: Vodicka
authors: Department of Neurology, Department of Neuroscience, Friedman Brain Institute, Icahn School of Medicine at Mount Sinai, Leon and Norma Hess Center for Science and Medicine, New York, NY, USA.
authors: Junghyun
authors: J
authors: Lim
authors: Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA, USA.
authors: Dana T
authors: DT
authors: Williams
authors: Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA, USA.
authors: Kimberly B
authors: KB
authors: Kegel
authors: Departments of Medicine and Cell Biology, University of Massachusetts, Worcester, MA, USA.
authors: Kathryn
authors: K
authors: Chase
authors: CHDI Management/CHDI Foundation, Los Angeles, CA, USA.
authors: Hyunsun
authors: H
authors: Park
authors: CHDI Management/CHDI Foundation, Los Angeles, CA, USA.
authors: Deanna
authors: D
authors: Marchionini
authors: BioFocus DPI Limited, Chesterford Research Park, UK.
authors: Stephen
authors: S
authors: Wilkinson
authors: BioFocus DPI Limited, Chesterford Research Park, UK.
authors: Tania
authors: T
authors: Mead
authors: BioFocus DPI Limited, Chesterford Research Park, UK.
authors: Helen
authors: H
authors: Birch
authors: BioFocus DPI Limited, Chesterford Research Park, UK.
authors: Dawn
authors: D
authors: Yates
authors: Independent consultant.
authors: Kathy
authors: K
authors: Lyons
authors: CHDI Management/CHDI Foundation, Los Angeles, CA, USA.
authors: Celia
authors: C
authors: Dominguez
authors: CHDI Management/CHDI Foundation, Los Angeles, CA, USA.
authors: Maria
authors: M
authors: Beconi
authors: Department of Neurology, Department of Neuroscience, Friedman Brain Institute, Icahn School of Medicine at Mount Sinai, Leon and Norma Hess Center for Science and Medicine, New York, NY, USA.
authors: Zhenyu
authors: Z
authors: Yue
authors: Departments of Medicine and Cell Biology, University of Massachusetts, Worcester, MA, USA.
authors: Neil
authors: N
authors: Aronin
authors: Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA, USA.
authors: Marian
authors: M
authors: DiFiglia
mesh_terms: D000818:Animals; D000962:Antimalarials / Q000493:pharmacokinetics / Q000494:pharmacology; D001343:Autophagy / Q000187:drug effects*; D001921:Brain / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D002738:Chloroquine / Q000493:pharmacokinetics / Q000494:pharmacology*; D004195:Disease Models, Animal; D055879:Gene Knock-In Techniques; D000071058:Huntingtin Protein; D006816:Huntington Disease / Q000188:drug therapy* / Q000473:pathology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D051758:Transcription Factor TFIIH; D014157:Transcription Factors / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000378:metabolism; D000090802:rab7 GTP-Binding Proteins
chemical_list: D000962:Antimalarials; C490981:Gtf2h1 protein, mouse; C506812:Htt protein, mouse; D000071058:Huntingtin Protein; C480860:Map1lc3b protein, mouse; D008869:Microtubule-Associated Proteins; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; D014157:Transcription Factors; D000090802:rab7 GTP-Binding Proteins; C000716465:rab7 GTP-binding proteins, mouse; D051758:Transcription Factor TFIIH; D002738:Chloroquine; D020691:rab GTP-Binding Proteins