title: Hydroxyurea enhances SMN2 gene expression through nitric oxide release.
abstract: Small molecules that increase full-length survivor motor neuron (SMN) gene transcript are promising therapeutic candidates for spinal muscular atrophy (SMA). Hydroxyurea (HU) has recently been shown to increase full-length SMN transcript in cultured lymphocytes from patients with SMA. We investigate the mechanism by which HU enhances full-length SMN2 gene expression in SMA lymphocytes. Nitric oxide (NO) is a major intracellular metabolite of HU. We test whether NO donors can themselves enhance full-length SMN2 expression. Eighteen cell lines (five type I, five type II, six type III SMA, and two non-SMA controls) were treated with or without NO donors for 48 h. SMA cells treated with HU and three NO donors: two long-acting donors, Deta-NONOate and S-nitrosoglutathione, and one short-acting donor, 3-ethyl-3-(ethylaminoethyl)-1-hydroxy-2-oxo-1-triazene, resulted in significant increase in full-length SMN2 mRNA. These effects were abolished by co-treatment with an NO scavenger 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide. One short-acting NO donor, S-nitroso-N-acetyl-DL-penicillamine, failed to show significant effect on full-length SMN2 expression, possibly due to high degree of cytotoxicity. These results were observed using both densitometry and quantitative PCR methods. We conclude that HU enhances SMN2 expression through the release of NO. NO donors may themselves be considered as new therapeutic candidates for SMA.
authors.affiliation: Department of Neurology and Neurological Sciences, Stanford University Medical Center, 300 Pasteur Drive, Stanford, CA 94305-5235, USA.
authors.forename: Cheng
authors.initials: C
authors.lastname: Xu
authors.forename: Xin
authors.initials: X
authors.lastname: Chen
authors.forename: Susanna M
authors.initials: SM
authors.lastname: Grzeschik
authors.forename: Madhuri
authors.initials: M
authors.lastname: Ganta
authors.forename: Ching H
authors.initials: CH
authors.lastname: Wang
mesh_terms: D001483:Base Sequence; D002460:Cell Line; D003497:Cyclic N-Oxides / Q000494:pharmacology; D017931:DNA Primers / Q000235:genetics; D016166:Free Radical Scavengers / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D006918:Hydroxyurea / Q000494:pharmacology*; D007093:Imidazoles / Q000494:pharmacology; D008214:Lymphocytes / Q000187:drug effects / Q000378:metabolism; D009134:Muscular Atrophy, Spinal / Q000188:drug therapy* / Q000235:genetics* / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D009603:Nitroso Compounds / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055540:Survival of Motor Neuron 2 Protein / Q000235:genetics
chemical_list: D003497:Cyclic N-Oxides; D017931:DNA Primers; D016166:Free Radical Scavengers; D007093:Imidazoles; D020030:Nitric Oxide Donors; D009603:Nitroso Compounds; D012333:RNA, Messenger; C526477:SMN2 protein, human; D055540:Survival of Motor Neuron 2 Protein; C094210:2,2'-(hydroxynitrosohydrazono)bis-ethanamine; C079391:2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide; D009569:Nitric Oxide; D006918:Hydroxyurea