biostudies-literatureUnknownMcDiarmid TANICHD NIH HHSNIA NIH HHSNIMH NIH HHSNHGRI NIH HHSNINDS NIH HHSCRISPR-based gene activation (CRISPRa) has emerged as a promising therapeutic approach for neurodevelopmental disorders (NDD) caused by haploinsufficiency. However, scaling this <i>cis</i>-regulatory therapy (CRT) paradigm requires pinpointing which candidate <i>cis</i>-regulatory elements (cCREs) are active in human neurons, and which can be targeted with CRISPRa to yield specific and therapeutic levels of target gene upregulation. Here, we combine Massively Parallel Reporter Assays (MPRAs) and a multiplex single cell CRISPRa screen to discover functional human neural enhancers whose CRISPRa targeting yields specific upregulation of NDD risk genes. First, we tested 5,425 candidate neuronal enhancers with MPRA, identifying 2,422 that are active in human neurons. Selected cCREs also displaybioRxiv : the preprint server for biology2025.11.04.686611https://www.ebi.ac.uk/biostudies/studies/S-EPMC12637570biostudies-literatureLarge-scale discovery of neural enhancers for cis-regulation therapies.PMC12637570F32 MH124337R01 MH116999R01 NS114086F32 HG011817R01 MH123828R01 MH115957R01 NS129914U01 MH122681R01 AG050986R01 HG003988RM1 MH132651U01 HG011755UM1 HG011966R01 NS131615R01 HD105266U01 MH116489R01 MH125246R01 MH129722Chen GTFink KPhilpot BDDaza RMShendure JJames LMVij PPennacchio LAChardon FMLee ASCalderon DLaboy-Cintron DMcDiarmid TANourie HCMartin BKLalanne JBMuotri ARSanders SJKosicki MTalkowski MEPage NFAhituv NGeschwind DHfalseLarge-scale discovery of neural enhancers for cis-regulation therapies.CRISPR-based gene activation (CRISPRa) has emerged as a promising therapeutic approach for neurodevelopmental disorders (NDD) caused by haploinsufficiency. However, scaling this <i>cis</i>-regulatory therapy (CRT) paradigm requires pinpointing which candidate <i>cis</i>-regulatory elements (cCREs) are active in human neurons, and which can be targeted with CRISPRa to yield specific and therapeutic levels of target gene upregulation. Here, we combine Massively Parallel Reporter Assays (MPRAs) and a multiplex single cell CRISPRa screen to discover functional human neural enhancers whose CRISPRa targeting yields specific upregulation of NDD risk genes. First, we tested 5,425 candidate neuronal enhancers with MPRA, identifying 2,422 that are active in human neurons. Selected cCREs also display2025-01-01T00:00:00Z2025 Nov2026-06-10T03:14:13.498Z2026-06-10T03:07:53.522ZS-EPMC126375704127895310.1101/2025.11.04.686611