The 5'untranslated region of the mRNA for the Parkinson's a-synuclein (ASYN 5'UTR) is a key translational regulatory element that contributes to setting the amount of alpha-synuclein production in neural cells, particularly in response to Fe influx (iron accumulation is one of the key hallmarks of PD in the Substantia nigra). Our preliminary data showed ASYN gene expression is controlled by iron at the level of message translation without change to transcription or steady state levels of ASYN mRNA in neuroblastoma (SH-SY5Y cells). This regulation is similar to another metalloprotein, the Amyloid Precursor Protein of Alzheimer's disease, implying that neurodegenerative disease genes may be linked to the mis-metabolism of iron and/or associated oxidative events. These findings, and our experience with the regulation of APP mRNA translation, suggested the ASYN 5'UTR would be an excellent drug target. We already generated both ASYN and APP 5'UTR constructs and stable cell lines available for screening small molecules from the MLSCN molecular library of 500,000 compounds. Our screening strategy to identify APP 5'UTR drug hits was previously conducted with FDA and LDDN molecular libraries, and has now been adapted to screen the alpha-synuclein 5'UTR for this NIMH solicitation. As prior validation of our approach to screen the 5'UTRs of neurodegenerative disease transcripts, our APP 5'UTR screens generated drug selectivity since paroxetine was an APP 5'UTR directed FDA inhibitor that did not change APLP-1 and APLP-2 expression in SH-SY5Y cells (Payton et al., 2003). Paroxetine subsequently reduced the amyloid burden in a transgenic mouse model for AD (TgCRND8 mice) (Tucker et al., (2005/2006).
The aim of this grant application is to collaborate with the MLSCN to screen for 5'UTR directed small molecule inhibitors of translation of alpha synuclein mRNA for PD therapeutics. Our lab has stable cell lines that express luciferase reporter genes translationally driven by the 5'UTRs of target neurodegenerative proteins including ASYN (PD), APP (AD) and Prion protein (PrP, Creutzfeldt-Jacob disease). Dr. Ippolita Cantuti-Castelvetri (coPI, MIND, MGH) will provide the correct reagents, cell lines and experience to successfully ensure ASYN 5'UTR specific leads maintain neuronal viability and are sufficiently specific to inhibit toxic alpha-synuclein expression but leave compensatory beta-synuclein and gamma- synuclein expression unchanged. The data to be accumulated through this R21 molecular libraries screening cooperative mechanism will permit a high throughput transfection based screen of an important RNA target, the 5'untranslated region of the Parkinson's alpha synuclein (ASYN) transcript. This 5'UTR is an attractive therapeutic target for Parkinson's disease (PD), and newly identified ASYN 5'UTR specific leads may be developed by medicinal chemistry potentially to limit neurotoxic ASYN production in dopaminergic neurons. Use of 5'UTR specific MLSCN hits will probe the mechanism of translation of ASYN mRNA relevant to PD. Compounds directed to 5'UTRs of other mRNAs will probe mechanism of translation of the Abeta-amyloid precursor protein mRNA in Alzheimer's disease (SOD-1 mRNA in ALS, and PrP mRNA in Cruetzsfeld- Jacob Syndrome). ?
| Gao, Jin; Cahill, Catherine M; Huang, Xudong et al. (2018) S-Adenosyl Methionine and Transmethylation Pathways in Neuropsychiatric Diseases Throughout Life. Neurotherapeutics 15:156-175 |
| Bandyopadhyay, Sanghamitra; Rogers, Jack T (2014) Alzheimer's disease therapeutics targeted to the control of amyloid precursor protein translation: maintenance of brain iron homeostasis. Biochem Pharmacol 88:486-94 |
| Cahill, Catherine M; Lahiri, Debomoy K; Huang, Xudong et al. (2009) Amyloid precursor protein and alpha synuclein translation, implications for iron and inflammation in neurodegenerative diseases. Biochim Biophys Acta 1790:615-28 |
