Alternative splicing enhances proteomic diversity, yet its dysregulation drives cancer, neurodegeneration, and inherited disease. Small-molecule splicing modulators, while clinically validated, like risdiplam, often lack locus specificity, producing off-target effects. CRISPR/Cas13 enables programmable transcript-level targeting, but dCas13 fusion effectors are bulky and can hamper delivery and RNA homeostasis. Building on our previous Chem-CRISPR/dCas9 FCPF system for epigenome editing, we now introduce Chem-CRISPR/dCas13 FCPF , a modular platform that covalently tethers a perfluorobiphenyl-tagged small molecule to dCas13 via a four-residue FCPF -clamp tag. Guided by crRNAs, dRfxCas13d FCPF recruits a risdipla... More
Alternative splicing enhances proteomic diversity, yet its dysregulation drives cancer, neurodegeneration, and inherited disease. Small-molecule splicing modulators, while clinically validated, like risdiplam, often lack locus specificity, producing off-target effects. CRISPR/Cas13 enables programmable transcript-level targeting, but dCas13 fusion effectors are bulky and can hamper delivery and RNA homeostasis. Building on our previous Chem-CRISPR/dCas9 FCPF system for epigenome editing, we now introduce Chem-CRISPR/dCas13 FCPF , a modular platform that covalently tethers a perfluorobiphenyl-tagged small molecule to dCas13 via a four-residue FCPF -clamp tag. Guided by crRNAs, dRfxCas13d FCPF recruits a risdiplam-derived conjugate to the SMN2 exon 7 splice region, inducing exon inclusion at ligand doses 500-fold lower than those of free risdiplam and with no detectable effects at known risdiplam-sensitive transcripts in our assays. The approach generalizes to additional transcripts by crRNA redesign. By coupling CRISPR addressability with dose-sparing chemical action, Chem-CRISPR/dCas13 FCPF establishes a proximity-induced, chemically controllable route to precise RNA modulation suitable for therapeutic exploration.
