CRISPR-based therapeutics have advanced rapidly, but clinical translation remains bottlenecked by delivery. Standard carriers like viral vectors and lipid nanoparticles (LNPs) face challenges regarding cargo capacity, immunogenicity, and restricted tissue tropism. Peptide-mediated delivery platforms offer a highly tunable, non-viral alternative. These chemically defined sequences are uniquely modular: they are small enough to penetrate dense tissues, inexpensive to synthesize, and open to redesign residue by residue. In cell culture, peptide platforms compete effectively against standard electroporation, delivering high editing efficiency in primary human cells while significantly improving cell viability. Despite these in vitro successes, translating peptide-mediated delivery in vivo introduces physiological barriers. When introduced by tissue injection in vivo, peptide delivered CRISPR editors must tackle more barriers than mere cell entry and endosomal escape they had to face in the dish. They typically reach modest editing efficiencies, which, however, could still yield a therapeutic benefit. For systemic delivery, more fragile but transient ribonucleoprotein (RNP) complexes face hepatic clearance, phagocytic capture, and protein corona interference-obstacles that are driving a shift toward engineered nanoassemblies and mRNA/DNA cargo alternatives. This review outlines the evolution of peptide-based CRISPR delivery from cell culture to local and systemic in vivo applications, evaluating physiological boundaries and highlighting engineering avenues required to overcome them. Looking forward, the next decade of peptide-mediated delivery must reconcile the conservative demands of clinical translation with radical chemical innovation. To advance, the field must transition past qualitative readouts and integrate isolated advancements into a single cohesive vehicle engineered for robust in vivo efficacy. By systematically combining these chemical innovations, engineered peptides might overcome the restrictions of viruses and LNPs, providing a truly versatile platform for systemic CRISPR delivery.
