Scaling haplospecific antisense oligonucleotides from N-of-1 to broad use in genetic disease populations by diplotyping

Antisense oligonucleotides (ASO) are versatile disease modifying therapies for genetic diseases. An accelerated FD) pathway enables ASO treatment trial initiation in single patients within a year. However, this rapid N-of-1 pathway lacks extensibility to broad use necessary for sustainability. Individualized ASOs bind pre-mRNAs encompassing an entire locus. Thus, ASOs targeting common heterozygous polymorphisms (SNPs) are potentially haplospecific in many patients with dominant disorders. We developed haplospecific ASOs for two patients with SCN2A-Complex Neurodevelopmental Disorder (CND) and gain-of-function (GOF) or mixed gain-loss dysfunction (GLD) variants. The ASOs targeted reference SCN2A intronic sequences containing SNPs. The patients each had SCN2A haplotypes with reference SNP alleles in cis with causal variants and alternate SNP alleles in cis with normal mRNA. Following N-of-1 demonstration of safety and efficacy, we evaluated their applicability to 21 SCN2A-CND patients using whole genome sequencing (WGS) with haplotyping by read proximity. Ten (48%) patients had ASO-eligible diplotypes. Haplotype analysis of 1000 Genomes Project (1kGP) participants revealed 16 additional SCN2A haplotypes present in >20% of subjects, tagged by 156 SNPs. In silico assessment of specificity and potency identified additional haplospecific ASOs for validation in reprogrammed 1kGP cells heterozygous for the tagging SNPs. A combination of 4 haplospecific ASOs provided coverage for 76% of 1kGP subjects, potentially scaling N-of-1 FDA applications in future SCN2A-CND patients with GOF/GLD variants by ASO selection by diagnosis by WGS with haplotyping. Thus, population resources have potential to prepare haplospecific ASO therapies a priori for many patients and genetic diseases, with individual selection by WGS haplotyping. One Sentence SummaryPopulation genome sequencing with haplotyping identifies haplospecific antisense oligonucleotides for disease modifying therapy of genetic disorders.