Systemic Injection of Peptide-PMOs into Humanized DMD Mice and Evaluation by RT-PCR and ELISA

Abstract

Duchenne muscular dystrophy (DMD) is an X-linked recessive disorder due to the lack of dystrophin production. The disease is characterized by muscle wasting, with the most common causes of death being respiratory failure or heart failure. Recently, exon skipping using a phosphorodiamidate morpholino oligomer (PMO) is used as an FDA approved treatment for DMD. Peptide-conjugated PMOs (PPMOs) are used to increase exon skipping efficacy in the heart and are a promising therapy for DMD. Researchers have previously relied on high-performance liquid chromatography (HPLC) or liquid chromatography-mass spectrometry (LC/MS) methods for detecting PPMO uptake, but an enzyme-linked immunosorbent assay (ELISA) has been shown to have greater sensitivity. Here, we present methodologies to determine the uptake efficiency of a PPMO into the heart and efficacy of exon 51 skipping by a PPMO injected retro-orbitally into a humanized DMD mouse model via ELISA and RT-PCR, respectively.