Chemical probes to potently and selectively inhibit endocannabinoid cellular reuptake

Proc Natl Acad Sci U S A. 2017 Jun 20;114(25):E5006-E5015. doi: 10.1073/pnas.1704065114. Epub 2017 Jun 5.

Abstract

The extracellular effects of the endocannabinoids anandamide and 2-arachidonoyl glycerol are terminated by enzymatic hydrolysis after crossing cellular membranes by facilitated diffusion. The lack of potent and selective inhibitors for endocannabinoid transport has prevented the molecular characterization of this process, thus hindering its biochemical investigation and pharmacological exploitation. Here, we report the design, chemical synthesis, and biological profiling of natural product-derived N-substituted 2,4-dodecadienamides as a selective endocannabinoid uptake inhibitor. The highly potent (IC50 = 10 nM) inhibitor N-(3,4-dimethoxyphenyl)ethyl amide (WOBE437) exerted pronounced cannabinoid receptor-dependent anxiolytic, antiinflammatory, and analgesic effects in mice by increasing endocannabinoid levels. A tailored WOBE437-derived diazirine-containing photoaffinity probe (RX-055) irreversibly blocked membrane transport of both endocannabinoids, providing mechanistic insights into this complex process. Moreover, RX-055 exerted site-specific anxiolytic effects on in situ photoactivation in the brain. This study describes suitable inhibitors to target endocannabinoid membrane trafficking and uncovers an alternative endocannabinoid pharmacology.

Keywords: 2-AG; endocannabinoid reuptake; endocannabinoid system; inhibitor; lipid transport.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Anti-Anxiety Agents / pharmacology
  • Anti-Inflammatory Agents / pharmacology
  • Arachidonic Acids / metabolism
  • Biological Transport / drug effects*
  • Brain / drug effects
  • Brain / metabolism
  • Cell Line, Tumor
  • Cell Membrane / drug effects
  • Cell Membrane / metabolism
  • Endocannabinoids / metabolism*
  • Glycerides / metabolism
  • Humans
  • Hydrolysis / drug effects
  • Mice
  • Mice, Inbred BALB C
  • Mice, Inbred C57BL
  • Polyunsaturated Alkamides / metabolism
  • Receptors, Cannabinoid / metabolism
  • U937 Cells

Substances

  • Anti-Anxiety Agents
  • Anti-Inflammatory Agents
  • Arachidonic Acids
  • Endocannabinoids
  • Glycerides
  • Polyunsaturated Alkamides
  • Receptors, Cannabinoid
  • glyceryl 2-arachidonate
  • anandamide