{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,5]],"date-time":"2026-08-05T11:19:01Z","timestamp":1785928741469,"version":"3.56.0"},"reference-count":40,"publisher":"MDPI AG","issue":"12","license":[{"start":{"date-parts":[[2021,12,7]],"date-time":"2021-12-07T00:00:00Z","timestamp":1638835200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Annovis Bio","award":["None"],"award-info":[{"award-number":["None"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Pharmaceutics"],"abstract":"<jats:p>Posiphen tartrate (Posiphen) is an orally available small molecule that targets a conserved regulatory element in the mRNAs of amyloid precursor protein (APP) and \u03b1-synuclein (\u03b1SYN) and inhibits their translation. APP and \u03b1SYN can cause neurodegeneration when their aggregates induce neurotoxicity. Therefore, Posiphen is a promising drug candidate for neurodegenerative diseases, including Alzheimer\u2019s disease and Parkinson\u2019s disease. Posiphen\u2019s safety has been demonstrated in three independent phase I clinical trials. Moreover, in a proof of concept study, Posiphen lowered neurotoxic proteins and inflammatory markers in cerebrospinal fluid of mild cognitive impaired patients. Herein we investigated whether Posiphen reduced the expression of other proteins, as assessed by stable isotope labeling with amino acids in cell culture (SILAC) followed by mass spectrometry (MS)-based proteomics. Neuroblastoma SH-SY5Y cells, an in vitro model of neuronal function, were used for the SILAC protein profiling response. Proteins whose expression was altered by Posiphen treatment were characterized for biological functions, pathways and networks analysis. The most significantly affected pathway was the Huntington\u2019s disease signaling pathway, which, along with huntingtin (HTT) protein, was down-regulated by Posiphen in the SH-SY5Y cells. The downregulation of HTT protein by Posiphen was confirmed by quantitative Western blotting and immunofluorescence. Unchanged mRNA levels of HTT and a comparable decay rate of HTT proteins after Posiphen treatment supported the coclusion that Posiphen reduced HTT via downregulation of the translation of HTT mRNA. Meanwhile, the downregulation of APP and \u03b1SYN proteins by Posiphen was also confirmed. The mRNAs encoding HTT, APP and \u03b1SYN contain an atypical iron response element (IRE) in their 5\u2032-untranslated regions (5\u2032-UTRs) that bind iron regulatory protein 1 (IRP1), and Posiphen specifically bound this complex. Conversely, Posiphen did not bind the IRP1\/IRE complex of mRNAs with canonical IREs, and the translation of these mRNAs was not affected by Posiphen. Taken together, Posiphen shows high affinity binding to the IRE\/IRP1 complex of mRNAs with an atypical IRE stem loop, inducing their translation suppression, including the mRNAs of neurotoxic proteins APP, \u03b1SYN and HTT.<\/jats:p>","DOI":"10.3390\/pharmaceutics13122109","type":"journal-article","created":{"date-parts":[[2021,12,7]],"date-time":"2021-12-07T11:00:23Z","timestamp":1638874823000},"page":"2109","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":18,"title":["Posiphen Reduces the Levels of Huntingtin Protein through Translation Suppression"],"prefix":"10.3390","volume":"13","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-9799-7246","authenticated-orcid":false,"given":"Xu-Qiao","family":"Chen","sequence":"first","affiliation":[{"name":"Department of Neurosciences, University of California San Diego, La Jolla, CA 92093, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5779-6642","authenticated-orcid":false,"given":"Carlos A.","family":"Barrero","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Science, Moulder Center for Drug Discovery, School of Pharmacy, Temple University, Philadelphia, PA 19140, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rodrigo","family":"Vasquez-Del Carpio","sequence":"additional","affiliation":[{"name":"Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA"},{"name":"Sandoz, Novartis Division, Princeton, NJ 08540, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"E. Premkumar","family":"Reddy","sequence":"additional","affiliation":[{"name":"Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chiara","family":"Fecchio","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Science, Moulder Center for Drug Discovery, School of Pharmacy, Temple University, Philadelphia, PA 19140, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Salim","family":"Merali","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Science, Moulder Center for Drug Discovery, School of Pharmacy, Temple University, Philadelphia, PA 19140, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alessia","family":"Deglincerti","sequence":"additional","affiliation":[{"name":"Laboratory of Stem Cell Biology and Molecular Embryology, The Rockefeller University, New York, NY 10065, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Cheng","family":"Fang","sequence":"additional","affiliation":[{"name":"Annovis Bio, Berwyn, PA 19312, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1262-8578","authenticated-orcid":false,"given":"Jack","family":"Rogers","sequence":"additional","affiliation":[{"name":"Neurochemistry Laboratory, Psychiatry-Neuroscience, Massachusetts General Hospital, Charlestown, MA 02129, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maria L.","family":"Maccecchini","sequence":"additional","affiliation":[{"name":"Annovis Bio, Berwyn, PA 19312, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2021,12,7]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"281","DOI":"10.2174\/1567205054367829","article-title":"An overview of phenserine tartrate, a novel acetylcholinesterase inhibitor for the treatment of Alzheimer\u2019s disease","volume":"2","author":"Greig","year":"2005","journal-title":"Curr. Alzheimer Res."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"386","DOI":"10.1124\/jpet.106.112102","article-title":"The experimental Alzheimer\u2019s disease drug posiphen [(+)-phenserine] lowers amyloid-beta peptide levels in cell culture and mice","volume":"320","author":"Lahiri","year":"2007","journal-title":"J. Pharmacol. Exp. Ther."},{"key":"ref_3","first-page":"142372","article-title":"The anticholinesterase phenserine and its enantiomer posiphen as 5\u2032untranslated-region-directed translation blockers of the Parkinson\u2019s alpha synuclein expression","volume":"2012","author":"Mikkilineni","year":"2012","journal-title":"Parkinsons Dis."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"615","DOI":"10.1016\/j.bbagen.2008.12.001","article-title":"Amyloid precursor protein and alpha synuclein translation, implications for iron and inflammation in neurodegenerative diseases","volume":"1790","author":"Cahill","year":"2009","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"493","DOI":"10.1007\/s00702-010-0513-5","article-title":"The alpha-synuclein 5\u2032untranslated region targeted translation blockers: Anti-alpha synuclein efficacy of cardiac glycosides and Posiphen","volume":"118","author":"Rogers","year":"2011","journal-title":"J. Neural Transm."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"117","DOI":"10.2174\/1871523011312020003","article-title":"Synthesis of the Alzheimer drug Posiphen into its primary metabolic products (+)-N1-norPosiphen, (+)-N8-norPosiphen and (+)-N1, N8-bisnorPosiphen, their inhibition of amyloid precursor protein, alpha-Synuclein synthesis, interleukin-1beta release, and cholinergic action","volume":"12","author":"Yu","year":"2013","journal-title":"Antiinflamm Antiallergy Agents Med. Chem."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"12506","DOI":"10.1073\/pnas.0705346104","article-title":"Modulation of human neural stem cell differentiation in Alzheimer (APP23) transgenic mice by phenserine","volume":"104","author":"Marutle","year":"2007","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"271","DOI":"10.1002\/alz.12185","article-title":"Targeting increased levels of APP in Down syndrome: Posiphen-mediated reductions in APP and its products reverse endosomal phenotypes in the Ts65Dn mouse model","volume":"17","author":"Chen","year":"2021","journal-title":"Alzheimers Dement."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"2420","DOI":"10.4103\/1673-5374.313044","article-title":"A pilot exploration with Posiphen to normalize amyloid precursor protein in Down syndrome","volume":"16","author":"Chen","year":"2021","journal-title":"Neural Regen. Res."},{"key":"ref_10","first-page":"37","article-title":"Translational inhibition of APP by Posiphen: Efficacy, pharmacodynamics, and pharmacokinetics in the APP\/PS1 mouse","volume":"4","author":"Teich","year":"2018","journal-title":"Alzheimers Dement."},{"key":"ref_11","first-page":"1","article-title":"Translational inhibition of alpha-synuclein by Posiphen normalizes distal colon motility in transgenic Parkinson mice","volume":"8","author":"Kuo","year":"2019","journal-title":"Am. J. Neurodegener. Dis."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"894","DOI":"10.1136\/jnnp-2012-302589","article-title":"Posiphen as a candidate drug to lower CSF amyloid precursor protein, amyloid-beta peptide and tau levels: Target engagement, tolerability and pharmacokinetics in humans","volume":"83","author":"Maccecchini","year":"2012","journal-title":"J. Neurol. Neurosurg. Psychiatry"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1226","DOI":"10.3390\/ijms10031226","article-title":"Physiological and pathological role of alpha-synuclein in Parkinson\u2019s disease through iron mediated oxidative stress; the role of a putative iron-responsive element","volume":"10","author":"Olivares","year":"2009","journal-title":"Int. J. Mol. Sci."},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Rogers, J.T., Xia, N., Wong, A., Bakshi, R., and Cahill, C.M. (2019). Targeting the Iron-Response Elements of the mRNAs for the Alzheimer\u2019s Amyloid Precursor Protein and Ferritin to Treat Acute Lead and Manganese Neurotoxicity. Int. J. Mol. Sci., 20.","DOI":"10.3390\/ijms20040994"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"75","DOI":"10.1186\/s13024-017-0218-4","article-title":"Iron regulatory protein (IRP)-iron responsive element (IRE) signaling pathway in human neurodegenerative diseases","volume":"12","author":"Zhou","year":"2017","journal-title":"Mol. Neurodegener."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"31217","DOI":"10.1074\/jbc.M110.149161","article-title":"Selective translational control of the Alzheimer amyloid precursor protein transcript by iron regulatory protein-1","volume":"285","author":"Cho","year":"2010","journal-title":"J. Biol. Chem."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"7605","DOI":"10.1073\/pnas.131152998","article-title":"Phenserine regulates translation of beta -amyloid precursor protein mRNA by a putative interleukin-1 responsive element, a target for drug development","volume":"98","author":"Shaw","year":"2001","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"34","DOI":"10.1196\/annals.1332.003","article-title":"The integrated role of desferrioxamine and phenserine targeted to an iron-responsive element in the APP-mRNA 5\u2032-untranslated region","volume":"1035","author":"Venti","year":"2004","journal-title":"Ann. N. Y. Acad. Sci."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"9866","DOI":"10.1073\/pnas.94.18.9866","article-title":"Iron accumulation in Alzheimer disease is a source of redox-generated free radicals","volume":"94","author":"Smith","year":"1997","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"77","DOI":"10.1023\/A:1020731021276","article-title":"Adventiously-bound redox active iron and copper are at the center of oxidative damage in Alzheimer disease","volume":"16","author":"Perry","year":"2003","journal-title":"Biometals"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"5","DOI":"10.1097\/01.rmr.0000245461.90406.ad","article-title":"Role of iron in neurodegenerative disorders","volume":"17","author":"Berg","year":"2006","journal-title":"Top. Magn. Reson. Imaging"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"371","DOI":"10.1001\/archneurol.2008.586","article-title":"Iron accumulation in the substantia nigra of patients with Alzheimer disease and parkinsonism","volume":"66","author":"Brar","year":"2009","journal-title":"Arch. Neurol."},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Bandyopadhyay, S., Cahill, C., Balleidier, A., Huang, C., Lahiri, D.K., Huang, X., and Rogers, J.T. (2013). Novel 5\u2032 untranslated region directed blockers of iron-regulatory protein-1 dependent amyloid precursor protein translation: Implications for down syndrome and Alzheimer\u2019s disease. PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0065978"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"90","DOI":"10.1016\/j.ydbio.2006.03.026","article-title":"Contribution of human embryonic stem cells to mouse blastocysts","volume":"295","author":"James","year":"2006","journal-title":"Dev. Biol."},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Barrero, C.A., Datta, P.K., Sen, S., Deshmane, S., Amini, S., Khalili, K., and Merali, S. (2013). HIV-1 Vpr modulates macrophage metabolic pathways: A SILAC-based quantitative analysis. PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0068376"},{"key":"ref_26","doi-asserted-by":"crossref","unstructured":"Obakan, P., Barrero, C., Coker-Gurkan, A., Arisan, E.D., Merali, S., and Palavan-Unsal, N. (2015). SILAC-Based Mass Spectrometry Analysis Reveals That Epibrassinolide Induces Apoptosis via Activating Endoplasmic Reticulum Stress in Prostate Cancer Cells. PLoS ONE, 10.","DOI":"10.1371\/journal.pone.0135788"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"3","DOI":"10.1016\/B978-0-12-391940-3.00001-9","article-title":"Examining histone posttranslational modification patterns by high-resolution mass spectrometry","volume":"512","author":"Lin","year":"2012","journal-title":"Methods Enzymol."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"D792","DOI":"10.1093\/nar\/gkp1005","article-title":"PrimerBank: A resource of human and mouse PCR primer pairs for gene expression detection and quantification","volume":"38","author":"Spandidos","year":"2010","journal-title":"Nucleic Acids Res."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"262","DOI":"10.1016\/0076-6879(89)80106-5","article-title":"Computer prediction of RNA structure","volume":"180","author":"Zuker","year":"1989","journal-title":"Methods Enzymol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"183","DOI":"10.1006\/nbdi.1996.0018","article-title":"Differential expression of normal and mutant Huntington\u2019s disease gene alleles","volume":"3","author":"Persichetti","year":"1996","journal-title":"Neurobiol. Dis."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"1457","DOI":"10.1073\/pnas.1905057117","article-title":"Translation of the intrinsically disordered protein alpha-synuclein is inhibited by a small molecule targeting its structured mRNA","volume":"117","author":"Zhang","year":"2020","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"E5655","DOI":"10.1073\/pnas.1603020113","article-title":"TRiC subunits enhance BDNF axonal transport and rescue striatal atrophy in Huntington\u2019s disease","volume":"113","author":"Zhao","year":"2016","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"1051","DOI":"10.1083\/jcb.200108057","article-title":"Tau blocks traffic of organelles, neurofilaments, and APP vesicles in neurons and enhances oxidative stress","volume":"156","author":"Stamer","year":"2002","journal-title":"J. Cell Biol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"5428","DOI":"10.1073\/pnas.1303279110","article-title":"Molecular chaperone Hsp110 rescues a vesicle transport defect produced by an ALS-associated mutant SOD1 protein in squid axoplasm","volume":"110","author":"Song","year":"2013","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"536","DOI":"10.1016\/j.neuron.2013.12.018","article-title":"Axonal transport of TDP-43 mRNA granules is impaired by ALS-causing mutations","volume":"81","author":"Alami","year":"2014","journal-title":"Neuron"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"3365","DOI":"10.1523\/JNEUROSCI.5427-08.2009","article-title":"Dynamic changes in presynaptic and axonal transport proteins combined with striatal neuroinflammation precede dopaminergic neuronal loss in a rat model of AAV alpha-synucleinopathy","volume":"29","author":"Chung","year":"2009","journal-title":"J. Neurosci."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"613","DOI":"10.1038\/nn.3356","article-title":"Physiologic brain activity causes DNA double-strand breaks in neurons, with exacerbation by amyloid-beta","volume":"16","author":"Suberbielle","year":"2013","journal-title":"Nat. Neurosci."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"345","DOI":"10.1038\/s41380-018-0266-3","article-title":"Novel upregulation of amyloid-beta precursor protein (APP) by microRNA-346 via targeting of APP mRNA 5\u2032-untranslated region: Implications in Alzheimer\u2019s disease","volume":"24","author":"Long","year":"2019","journal-title":"Mol. Psychiatry"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"15005","DOI":"10.1038\/nrdp.2015.5","article-title":"Huntington disease","volume":"1","author":"Bates","year":"2015","journal-title":"Nat. Rev. Dis. Primers"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"1031","DOI":"10.1016\/j.neuron.2012.05.009","article-title":"Sustained therapeutic reversal of Huntington\u2019s disease by transient repression of huntingtin synthesis","volume":"74","author":"Kordasiewicz","year":"2012","journal-title":"Neuron"}],"container-title":["Pharmaceutics"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1999-4923\/13\/12\/2109\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T07:42:42Z","timestamp":1760168562000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1999-4923\/13\/12\/2109"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,12,7]]},"references-count":40,"journal-issue":{"issue":"12","published-online":{"date-parts":[[2021,12]]}},"alternative-id":["pharmaceutics13122109"],"URL":"https:\/\/doi.org\/10.3390\/pharmaceutics13122109","relation":{},"ISSN":["1999-4923"],"issn-type":[{"value":"1999-4923","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,12,7]]}}}