Modification of SMN2 exon 7 (E7) splicing to increase SMN protein production is a validated therapeutic strategy against Spinal Muscular Atrophy (SMA). Based on this, we have performed the first small molecule screening described for SMA, choosing a stem-loop RNA structure TSL2 that partially overlaps with the E7 5’ splicing site (5’ ss) of SMN2 as the biological target. TSL2-binding hit PK4C9 was found to also increase E7 splicing and rescued downstream molecular alterations in transfected HeLa cells, transgenic Drosophila, and SMA patient cells. High-resolution NMR combined with in silico modeling revealed that PK4C9 binding to TSL2 promotes a conformational shift towards a triloop conformation, which we also demonstrate that is associated with an enhanced E7 splicing efficiency. This work not only provides one of the few examples of small molecules with direct SMN2-spllicing modifier activity, but also opens new avenues for rational drug discovery in SMA and other splicing-mediated diseases where similar RNA structures are involved.