Fig. 5. Assessment of R403Q-allele inactivation and echocardiographic findings in R403Q-129SvEv/S4 mice treated with low, medium and high doses of AAV9-Cas9.

a, Percent of R403Q alleles with indels in three untreated male or AAV9-Cas9 treated (low dose: n = 1 male, 2 females; medium dose, n = 3 females and high dose, n = 4 females) R403Q-129SvEv/S4 mice was assessed by next-generation sequencing of PCR-amplified LV gDNA and analyzed using CRISPResso2. b, Myh6 cDNAs, amplified from RNAs extracted from each cardiac chamber of R403Q-129SvEv/S4 mice, treated with low (1.1 × 10¹² vg kg⁻¹; n = 1 male, 2 females), medium (5.4 × 10¹² vg kg⁻¹; n = 3 females) and high (2.2 × 10¹³ vg kg⁻¹; n = 2 females) doses of AAV9-Cas9 for 18–33 weeks. The percentage of inactivation of the pathogenic variant R403Q was calculated as: (1 – (total number of R403Q reads, divided by total number of WT reads)) multiplied by 100. c, Longitudinal echocardiographic measurements LVPW, LVPW/LVDd and FS (defined in Fig. 2) in WT 129SvEv/S4 (n = 5 males, 5 females, black line) and untreated R403Q-129SvEv/S4 (n = 5 males, 6 females, red line) and treated (AAV9-Cas9 2.2 × 10¹³ vg kg⁻¹) R403Q-129SvEv/S4 (n = 6 females, blue line). Note impaired contractile function (FS <40%) in high-dose-treated mice. d, Echocardiographic measurements of LVPW thickness, LVPW/LVDd ratio and FS (defined in Fig. 2) in R403Q-129SvEv/S4 mice at 20 weeks of age treated with low (1.1 × 10¹² vg kg⁻¹, n = 1 male, 2 females), medium (5.4 × 10¹² vg kg⁻¹, n = 3 females) and high (2.2 × 10¹³ vg kg⁻¹, n = 6 females) doses of AAV9-Cas9, compared to untreated WT 129SvEv/S4 (n = 3 females) and untreated R403Q-129SvEv/S4 (n = 4 females) mice. Note that one mouse treated with the low dose had increased LVPW and increased LVPW/LVDd, indicating a non-therapeutic response. Data are presented as mean values ± s.d., and significance was assessed by two-tailed t-test (Supplementary Information).

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