Description

Autism Spectrum Disorder (ASD) patients are affected with various symptoms of severity in their daily lives, including communication difficulties, problems with social interaction, and repetitive behaviors together with sensory problems. The advanced studies of ASD show that de-novo single-nucleotide mutations cause developmental disability. Our study will look to demonstrate that antisense oligonucleotide (ASO) therapy could reduce the effects of monogenetic ASD mutations. Materials and methods: To evaluate the efficacy of the ASO approach, we have collected patient-induced pluripotent stem cells (iPSCs) for four monogenetic forms of ASD genes: ASXL3, PACS1, PPP2R5D, and SYNGAP1. We generate induced neurons (iNs) from the patient iPSCs to use as vectors of study. Lentivirus is used to induce Neurogenin2 (NGN2) gene expression in the patient cells with doxycycline (DOX). To determine the effects of ASOs on the transcriptome, control iPSC lines were differentiated and then collected for RNAseq for three weeks at weekly intervals. Results: From the collected data, we demonstrated that iPSC cell markers expression decreased over the timeframe, and neural markers expression increased over the same span of time. Together with these findings, ASO dosing was optimized using qRT-PCR. Our study uses 5-10-5 2’-O-methoxyethyl (MOE) gapmer ASOs targeting alpha-actinin as a control. The findings suggested that using ASOs could achieve 80-85% knockdown of targeted genes in patient iNs. Conclusion and future steps: After assessing control cell lines, we would like to follow the same differentiation protocol for patient cell lines and knockdown gene expression with ASOs. Our approach will contribute to the development of RNA therapeutics for ASD patients. The study will aid our understanding of ASD and underline the cellular mechanisms of neurodevelopmental disease.