Fig. 1

Pipeline for high-throughput screening for sarcospan modulators. a Large chemical libraries were screened using the hSSPN-EGFP C2C12 reporter cell line (n = 1). The top 1000 hits were rescreened in both hSSPN-EGFP and hSSPN-luciferase C2C12 reporter cell lines (n = 3 each). Sixty-three of 1000 compounds increased reporter expression in both cell lines and were therefore considered confirmed hits. The confirmed hits were sorted into 3 groups based on common structural features: pharmacophore 1 (PC1), pharmacophore 2 (PC2), and other, which had no unifying structural theme. Dystrophin-deficient H2K mdx cells were treated with 5.5 μM of b pharmacophore 1 (PC1) or c other compounds for 48 h. All cells were treated at day 2 of differentiation and harvested 48 h posttreatment. Gene expression was normalized to housekeeping gene β-actin and vehicle-treated cells (0.5% DMSO). Data represents individual replicates and mean value. n = 3–8. SSPN, sarcospan; R.U., relative units. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001