A Small Molecule Exploits Hidden Structural Features Within the RNA Repeat Expansion That Causes C9ALS/FTD and Rescues Pathological Hallmarks

Overview

Journal ACS Chem Neurosci

Publisher American Chemical Society

Specialty Neurology

Date 2021 Oct 22

PMID 34677935

Citations 7

Authors

Andrei Ursu

Jared T Baisden

Jessica A Bush

Amirhossein Taghavi

Shruti Choudhary

Yong-Jie Zhang

Tania F Gendron

Leonard Petrucelli

Ilyas Yildirim

Matthew D Disney

Affiliations

Soon will be listed here.

Abstract

The hexanucleotide repeat expansion GGGGCC [r(GC)] within intron 1 of causes genetically defined amyotrophic lateral sclerosis and frontotemporal dementia, collectively named c9ALS/FTD. , the repeat expansion causes neurodegeneration via deleterious phenotypes stemming from r(GC) RNA gain- and loss-of-function mechanisms. The r(GC) RNA folds into both a hairpin structure with repeating 1 × 1 nucleotide GG internal loops and a G-quadruplex structure. Here, we report the identification of a small molecule (CB253) that selectively binds the hairpin form of r(GC). Interestingly, the small molecule binds to a previously unobserved conformation in which the RNA forms 2 × 2 nucleotide GG internal loops, as revealed by a series of binding and structural studies. NMR and molecular dynamics simulations suggest that the r(GC) hairpin interconverts between 1 × 1 and 2 × 2 internal loops through the process of strand slippage. We provide experimental evidence that CB253 binding indeed shifts the equilibrium toward the 2 × 2 GG internal loop conformation, inhibiting mechanisms that drive c9ALS/FTD pathobiology, such as repeat-associated non-ATG translation formation of stress granules and defective nucleocytoplasmic transport in various cellular models of c9ALS/FTD.

Citing Articles

The evolution and application of RNA-focused small molecule libraries.

Taghavi A, Springer N, Zanon P, Li Y, Li C, Childs-Disney J RSC Chem Biol. 2025; .

PMID: 39957993 PMC: 11824871. DOI: 10.1039/d4cb00272e.

Therapeutic targeting of RNA for neurological and neuromuscular disease.

Bubenik J, Scotti M, Swanson M Genes Dev. 2024; 38(15-16):698-717.

PMID: 39142832 PMC: 11444190. DOI: 10.1101/gad.351612.124.

Antisense RNA C9orf72 hexanucleotide repeat associated with amyotrophic lateral sclerosis and frontotemporal dementia forms a triplex-like structure and binds small synthetic ligand.

Blaszczyk L, Ryczek M, Das B, Mateja-Pluta M, Bejger M, Sliwiak J Nucleic Acids Res. 2024; 52(11):6707-6717.

PMID: 38738637 PMC: 11194091. DOI: 10.1093/nar/gkae376.

Mapping of repeat-associated non-AUG (RAN) translation knowledge: A bibliometric analysis.

Zhao T, Duan S, Li J, Zheng H, Liu C, Zhang H Heliyon. 2024; 10(8):e29141.

PMID: 38628764 PMC: 11019168. DOI: 10.1016/j.heliyon.2024.e29141.

Molecular basis of RNA-binding and autoregulation by the cancer-associated splicing factor RBM39.

Campagne S, Jutzi D, Malard F, Matoga M, Romane K, Feldmuller M Nat Commun. 2023; 14(1):5366.

PMID: 37666821 PMC: 10477243. DOI: 10.1038/s41467-023-40254-5.

References

Furtig B, Wenter P, Reymond L, Richter C, Pitsch S, Schwalbe H . Conformational dynamics of bistable RNAs studied by time-resolved NMR spectroscopy. J Am Chem Soc. 2007; 129(51):16222-9. DOI: 10.1021/ja076739r. View

Simone R, Balendra R, Moens T, Preza E, Wilson K, Heslegrave A . G-quadruplex-binding small molecules ameliorate FTD/ALS pathology and . EMBO Mol Med. 2017; 10(1):22-31. PMC: 5760849. DOI: 10.15252/emmm.201707850. View

Mukherjee S, Blaszczyk L, Rypniewski W, Falschlunger C, Micura R, Murata A . Structural insights into synthetic ligands targeting A-A pairs in disease-related CAG RNA repeats. Nucleic Acids Res. 2019; 47(20):10906-10913. PMC: 6847237. DOI: 10.1093/nar/gkz832. View

Connelly C, Moon M, Schneekloth Jr J . The Emerging Role of RNA as a Therapeutic Target for Small Molecules. Cell Chem Biol. 2016; 23(9):1077-1090. PMC: 5064864. DOI: 10.1016/j.chembiol.2016.05.021. View

Ganser L, Kelly M, Patwardhan N, Hargrove A, Al-Hashimi H . Demonstration that Small Molecules can Bind and Stabilize Low-abundance Short-lived RNA Excited Conformational States. J Mol Biol. 2019; 432(4):1297-1304. PMC: 7054137. DOI: 10.1016/j.jmb.2019.12.009. View

Rangadurai A, Szymaski E, Kimsey I, Shi H, Al-Hashimi H . Characterizing micro-to-millisecond chemical exchange in nucleic acids using off-resonance R relaxation dispersion. Prog Nucl Magn Reson Spectrosc. 2019; 112-113:55-102. PMC: 6727989. DOI: 10.1016/j.pnmrs.2019.05.002. View

Kiliszek A, Rypniewski W . Structural studies of CNG repeats. Nucleic Acids Res. 2014; 42(13):8189-99. PMC: 4117766. DOI: 10.1093/nar/gku536. View

Hennig M, Scott L, Sperling E, Bermel W, Williamson J . Synthesis of 5-fluoropyrimidine nucleotides as sensitive NMR probes of RNA structure. J Am Chem Soc. 2007; 129(48):14911-21. DOI: 10.1021/ja073825i. View

Zhang Q, An Y, Chen Z, Koon A, Lau K, Ngo J . A Peptidylic Inhibitor for Neutralizing (GGGGCC)-Associated Neurodegeneration in C9ALS-FTD. Mol Ther Nucleic Acids. 2019; 16:172-185. PMC: 6424097. DOI: 10.1016/j.omtn.2019.02.015. View

10.

Reining A, Nozinovic S, Schlepckow K, Buhr F, Furtig B, Schwalbe H . Three-state mechanism couples ligand and temperature sensing in riboswitches. Nature. 2013; 499(7458):355-9. DOI: 10.1038/nature12378. View

11.

Yin W, Rogge M . Targeting RNA: A Transformative Therapeutic Strategy. Clin Transl Sci. 2019; 12(2):98-112. PMC: 6440575. DOI: 10.1111/cts.12624. View

12.

Sholokh M, Sharma R, Grytsyk N, Zaghzi L, Postupalenko V, Dziuba D . Environmentally Sensitive Fluorescent Nucleoside Analogues for Surveying Dynamic Interconversions of Nucleic Acid Structures. Chemistry. 2018; 24(52):13850-13861. DOI: 10.1002/chem.201802297. View

13.

Reddy K, Zamiri B, Stanley S, Macgregor Jr R, Pearson C . The disease-associated r(GGGGCC)n repeat from the C9orf72 gene forms tract length-dependent uni- and multimolecular RNA G-quadruplex structures. J Biol Chem. 2013; 288(14):9860-9866. PMC: 3617286. DOI: 10.1074/jbc.C113.452532. View

14.

Zhao C, Anklin C, Greenbaum N . Use of ¹⁹F NMR methods to probe conformational heterogeneity and dynamics of exchange in functional RNA molecules. Methods Enzymol. 2014; 549:267-85. DOI: 10.1016/B978-0-12-801122-5.00012-X. View

15.

Sztuba-Solinska J, Chavez-Calvillo G, Cline S . Unveiling the druggable RNA targets and small molecule therapeutics. Bioorg Med Chem. 2019; 27(10):2149-2165. PMC: 7126819. DOI: 10.1016/j.bmc.2019.03.057. View

16.

Lee J, Dethoff E, Al-Hashimi H . Invisible RNA state dynamically couples distant motifs. Proc Natl Acad Sci U S A. 2014; 111(26):9485-90. PMC: 4084437. DOI: 10.1073/pnas.1407969111. View

17.

Disney M . Targeting RNA with Small Molecules To Capture Opportunities at the Intersection of Chemistry, Biology, and Medicine. J Am Chem Soc. 2019; 141(17):6776-6790. PMC: 6541398. DOI: 10.1021/jacs.8b13419. View

18.

Cleary J, Ranum L . New developments in RAN translation: insights from multiple diseases. Curr Opin Genet Dev. 2017; 44:125-134. PMC: 5951168. DOI: 10.1016/j.gde.2017.03.006. View

19.

Puffer B, Kreutz C, Rieder U, Ebert M, Konrat R, Micura R . 5-Fluoro pyrimidines: labels to probe DNA and RNA secondary structures by 1D 19F NMR spectroscopy. Nucleic Acids Res. 2009; 37(22):7728-40. PMC: 2794194. DOI: 10.1093/nar/gkp862. View

20.

Su Z, Zhang Y, Gendron T, Bauer P, Chew J, Yang W . Discovery of a biomarker and lead small molecules to target r(GGGGCC)-associated defects in c9FTD/ALS. Neuron. 2014; 83(5):1043-50. PMC: 4232217. DOI: 10.1016/j.neuron.2014.07.041. View