title: Screening with an NMNAT2-MSD platform identifies small molecules that modulate NMNAT2 levels in cortical neurons.
abstract: Nicotinamide mononucleotide adenylyl transferase 2 (NMNAT2) is a key neuronal maintenance factor and provides potent neuroprotection in numerous preclinical models of neurological disorders. NMNAT2 is significantly reduced in Alzheimer's, Huntington's, Parkinson's diseases. Here we developed a Meso Scale Discovery (MSD)-based screening platform to quantify endogenous NMNAT2 in cortical neurons. The high sensitivity and large dynamic range of this NMNAT2-MSD platform allowed us to screen the Sigma LOPAC library consisting of 1280 compounds. This library had a 2.89% hit rate, with 24 NMNAT2 positive and 13 negative modulators identified. Western analysis was conducted to validate and determine the dose-dependency of identified modulators. Caffeine, one identified NMNAT2 positive-modulator, when systemically administered restored NMNAT2 expression in rTg4510 tauopathy mice to normal levels. We confirmed in a cell culture model that four selected positive-modulators exerted NMNAT2-specific neuroprotection against vincristine-induced cell death while four selected NMNAT2 negative modulators reduced neuronal viability in an NMNAT2-dependent manner. Many of the identified NMNAT2 positive modulators are predicted to increase cAMP concentration, suggesting that neuronal NMNAT2 levels are tightly regulated by cAMP signaling. Taken together, our findings indicate that the NMNAT2-MSD platform provides a sensitive phenotypic screen to detect NMNAT2 in neurons.
authors.affiliation: Linda and Jack Gill Center, Department of Psychological and Brain Sciences, Indiana University, Bloomington, Indiana, United States of America.|The Cain Foundation Laboratories, Texas Children's Hospital, Houston, Texas, United States of America.|Jan and Dan Duncan Neurological Research Institute, Texas Children's Hospital, Houston, Texas, United States of America.|Department of Pediatrics, Baylor College of Medicine, Houston, Texas, United States of America.
authors.forename: Yousuf O
authors.initials: YO
authors.lastname: Ali
authors.affiliation: Linda and Jack Gill Center, Department of Psychological and Brain Sciences, Indiana University, Bloomington, Indiana, United States of America.|Developmental Biology Program and Department of Neuroscience, Baylor College of Medicine, Houston, Texas, United States of America.
authors.forename: Gillian
authors.initials: G
authors.lastname: Bradley
authors.affiliation: Linda and Jack Gill Center, Department of Psychological and Brain Sciences, Indiana University, Bloomington, Indiana, United States of America.|The Cain Foundation Laboratories, Texas Children's Hospital, Houston, Texas, United States of America.|Jan and Dan Duncan Neurological Research Institute, Texas Children's Hospital, Houston, Texas, United States of America.|Department of Pediatrics, Baylor College of Medicine, Houston, Texas, United States of America.|Developmental Biology Program and Department of Neuroscience, Baylor College of Medicine, Houston, Texas, United States of America.
authors.forename: Hui-Chen
authors.initials: HC
authors.lastname: Lu
mesh_terms: D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D000697:Central Nervous System Stimulants / Q000494:pharmacology; D002540:Cerebral Cortex / Q000166:cytology* / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D057166:High-Throughput Screening Assays / Q000379:methods*; D018345:Mice, Knockout; D008822:Mice, Transgenic; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D009612:Nicotinamide-Nucleotide Adenylyltransferase / Q000235:genetics / Q000378:metabolism*; D015203:Reproducibility of Results; D054852:Small Molecule Libraries / Q000494:pharmacology*; D024801:Tauopathies / Q000235:genetics / Q000378:metabolism
chemical_list: D000697:Central Nervous System Stimulants; D054852:Small Molecule Libraries; D002110:Caffeine; D009612:Nicotinamide-Nucleotide Adenylyltransferase