ConspectusThe field of gene therapy, which aims to treat patients by modulating gene expression, has come to fruition and has landed several landmark FDA approvals. Most gene therapies currently rely on viral vectors to deliver nucleic acid cargo into cells, but there is significant interest in moving toward chemical-based methods, such as polymer-based vectors, due to their low cost, immunocompatibility, and tunability. The full potential of polymer-based delivery systems has yet to be realized, however, because most polymeric transfection reagents are either too inefficient or too toxic for use in the clinic. In this Account, we describe developments in carbohydrate-based cationic polymers, termed glycopolymers, for enhanced nonviral gene delivery. As ubiquitous components of biological systems, carbohydrates are a rich class of compounds that can be harnessed to improve the biocompatibility of non-native polymers, such as linear polyamines used for promoting transfection. Reineke et al. developed a new class of carbohydrate-based polymers called poly(glycoamidoamine)s (PGAAs) by step-growth polymerization of linear monosaccharides with linear ethyleneamines. These glycopolymers were shown to be both efficient and biocompatible transfection reagents. Systematic modifications of the structural components of the PGAA system revealed structure-activity relationships important to its function, including its ability to degrade in situ.Expanding upon the development of step-growth glycopolymers, monosaccharides, such as glucose, were functionalized as vinyl-based monomers for the formation of diblock copolymers via radical addition-fragmentation chain-transfer (RAFT) polymerization. Upon complexation with plasmid DNA, the glucose-containing block creates a hydrophilic shell that promotes colloidal stability as effectively as PEG functionalization. An N-acetyl-d-galactosamine variant of this diblock polymer yields colloidally stable particles that show increased receptor-mediated uptake by liver hepatocytes in vitro and promotes liver targeting in mice. Finally, the disaccharide trehalose was incorporated into polycationic structures using both step-growth and RAFT techniques. It was shown that these trehalose-based copolymers imparted increased colloidal stability and yielded plasmid and siRNA polyplexes that resist aggregation upon lyophilization and reconstitution in water. The aforementioned series of glycopolymers use carbohydrates to promote effective and safe delivery of nucleic acid cargo into a variety of human cells types by promoting vehicle degradation, tissue-targeting, colloidal stabilization, and stability toward lyophilization to extend shelf life. Work is currently underway to translate the use of glycopolymers for safe and efficient delivery of nucleic acid cargo for gene therapy and gene editing applications.