Abstract
Genetic diseases can be diagnosed early during pregnancy, but many monogenic disorders continue to cause considerable neonatal and pediatric morbidity and mortality. Early intervention through intrauterine gene editing, however, could correct the genetic defect, potentially allowing for normal organ development, functional disease improvement, or cure. Here we demonstrate safe intravenous and intra-amniotic administration of polymeric nanoparticles to fetal mouse tissues at selected gestational ages with no effect on survival or postnatal growth. In utero introduction of nanoparticles containing peptide nucleic acids (PNAs) and donor DNAs corrects a disease-causing mutation in the β-globin gene in a mouse model of human β-thalassemia, yielding sustained postnatal elevation of blood hemoglobin levels into the normal range, reduced reticulocyte counts, reversal of splenomegaly, and improved survival, with no detected off-target mutations in partially homologous loci. This work may provide the basis for a safe and versatile method of fetal gene editing for human monogenic disorders.
Highlights
Genetic diseases can be diagnosed early during pregnancy, but many monogenic disorders continue to cause considerable neonatal and pediatric morbidity and mortality
We previously demonstrated that peptide nucleic acids (PNAs) and donor DNA could be efficiently encapsulated in NPs fabricated from poly(lactic-co-glycolic acid) (PLGA), a polymer that has been approved by the FDA for numerous drug delivery applications[17]
We showed that genomic correction achieved by PNA/DNA NPs leads to significant gene editing and phenotypic disease improvement in mouse models of β-thalassemia and cystic fibrosis[5,6,7]
Summary
Genetic diseases can be diagnosed early during pregnancy, but many monogenic disorders continue to cause considerable neonatal and pediatric morbidity and mortality. The PNAs contain nucleobases supported by a modified polyamide backbone[8] and bind to their specific genomic target site via both Watson−Crick and Hoogsteen base-pairing[9], yielding PNA/DNA/PNA triplex structures that induce endogenous DNA repair to mediate the recombination of the donor DNA molecule containing the correct sequence and produce specific, in situ gene correction[10,11,12]. This process is, in part, dependent on the nucleotide excision repair and homology dependent repair pathways10,13 [reviewed in ref. Unlike gene editing technologies that rely on the activity of exogenously delivered nucleases18,19—such as zinc finger nucleases, TAL effector nucleases, and CRISPR/Cas9— PNA/DNA NPs can be readily administered in vivo and have been shown to have extremely low to undetectable off-target effects in the genome because the PNA editing molecules lack inherent nuclease activity[5,6,7]
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