Single-base 2'OMe-modified LNA and MOE gapmers selectively silence ACVR1 R206H in fibrodysplasia ossificans progressiva.

Fibrodysplasia ossificans progressiva (FOP) is an ultra-rare disorder caused by gain-of-function mutations in ACVR1, most commonly c.617G>A (R206H), leading to progressive heterotopic ossification. In this study, we developed novel antisense gapmers selectively targeting the mutant ACVR1 R206H transcript while sparing the wild-type allele. We engineered locked nucleic acid (LNA) and 2'-O-methoxyethyl (MOE) gapmers incorporating a single 2'-O-methyl (2'OMe) modification at gap position 2. This is hypothesized to synergize with the wild-type sequence mismatch to restrict RNase H1 cleavage, limiting wild-type degradation while preserving mutant target engagement. In FOP patient-derived fibroblasts carrying the endogenous ACVR1 R206H mutation and in murine-derived C2C12 cells ectopically expressing ACVR1 R206H constructs, 2'OMe-modified gapmers demonstrated robust and preferential suppression of ACVR1 R206H at both RNA and protein levels. Gapmer treatment also reduced osteogenic differentiation, as shown by decreased alkaline phosphatase and Alizarin Red S staining, and lower expression of osteogenic markers. In wild-type mice, 2'OMe modification was associated with higher apparent gapmer levels in skeletal muscle and tendon and lower hepatic and renal stress marker readouts. These findings provide preliminary proof-of-concept that a single-base chemical modification can modulate allele selectivity and biodistribution of gapmers targeting ACVR1 R206H . Further studies in disease-relevant FOP models will be needed to establish therapeutic efficacy and long-term safety.