Fig. 3: Chemical modification pattern affects allele specificity and efficacy. | Nature Communications

Fig. 3: Chemical modification pattern affects allele specificity and efficacy.

From: Chemical engineering of therapeutic siRNAs for allele-specific gene silencing in Huntington’s disease models

Fig. 3

a Combinations of 2′-F and 2′-OMe enrichment around critical positions 6 and 11 affect siRNA activity. Compounds were tested using a dual-luciferase reporter assay system in HeLa cells. The (psiCheck) reporter plasmids contain a 40 mer region of huntingtin, including the target (A) SNP isoform, and non-target (G) SNP isoform. Cells were treated for 72 h with 1.5 µM siRNA. b–e 7-point dose-responses of alternative modification patterns show that backbone modification pattern impacts efficacy and discrimination. Compared to the original alternating modification pattern (b), a heavily-fluorinated SNP region shows an increase in target efficacy, but a complete loss of discrimination between alleles (c). A heavily methylated mismatch region increases discrimination but decreases target efficacy (d). Combining a fluorinated SNP region with a methylated mismatch region improves efficacy but does not increase discrimination above the original alternating modification pattern (e); n = 3 biological replicates for all experiments. All data are presented as mean ± standard deviation. Source data are provided as a source data file.

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