Mol Ther Nucleic Acids. 2023 Oct 17
Shukkwan K. Lau, Zachary C.E. Hawley, Maria I. Zavodszky, Sam Hana, Daniel Ferretti, Branka Grubor, Michael Hawes, Shanqin Xu, Stefan Hamann, Galina Marsh, Patrick Cullen, Ravi Challa, Thomas M. Carlile, Hang Zhang, Wan-Hung Lee, Andrea Peralta, Pete Clarner, Cong Wei, Kathryn Koszka, Feng Gao, Shih-Ching Lo
Products used in the paper Details Operation
AAV vector packaging PackGene Biotech (Worcester, MA) produced AAV9-amiR vectors using triple plasmid transfection protocol (Worchester, MA) as described previously.Transmission electron microscopy and image analysis were conducted by Packgene to confirm minimal number of empty AAV capsids for each vector produced (Fig. S10). AAV titers were quantified by ddPCR using a custom mCherry primer/probe Request Quote

Research Field: CNS

AAV Serotype: AAV9

Dose: 8E+10GC or 16E+10GC

Routes of Administration: ICV

Targeted organ: brain

Animal or cell line strain: Mouse


Toxic gain-of-function mutations in superoxide dismutase 1 (SOD1) contribute to ∼2-3% of all amyotrophic lateral sclerosis (ALS) cases. Artificial microRNAs (amiRs) delivered by adeno-associated virus (AAV) have been proposed as a potential treatment option to silence SOD1 expression and mitigate disease progression. Primary microRNA (pri-miRNA) scaffolds are used in amiRs to shuttle a hairpin RNA into the endogenous miRNA pathway, but it is unclear whether different primary miRNA scaffolds impact the potency and safety profile of the expressed amiR in vivo. In our process to develop an AAV amiR targeting SOD1, we performed a preclinical characterization of two primary miRNA scaffolds, miR155 and miR30a, sharing the same guide strand sequence. We report that while the miR155-based vector, compared to the miR30a-based vector, leads to a higher level of the amiR and more robust suppression of SOD1 in vitro and in vivo, it also presents significantly greater risks for CNS-related toxicities in vivo. Despite miR30a-based vector showing relatively lower potency, it can significantly delay the development of ALS-like phenotypes in SOD1-G93A mice and increase survival in a dose-dependent manner. This data highlights the importance of scaffold selection in the pursuit of highly efficacious and safe amiRs for RNAi gene therapy.

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