A versatile vector for gene and oligonucleotide transfer into cells in culture and in vivo: polyethylenimine.

Abstract

Several polycations possessing substantial buffering capacity below physiological pH, such as lipopolyamines and polyamidoamine polymers, are efficient transfection agents per se-i.e., without the addition of cell targeting or membrane-disruption agents. This observation led us to test the cationic polymer polyethylenimine (PEI) for its genedelivery potential. Indeed, every third atom of PEI is a protonable amino nitrogen atom, which makes the polymeric network an effective "proton sponge" at virtually any pH. Luciferase reporter gene transfer with this polycation into a variety of cell lines and primary cells gave results comparable to, or even better than, lipopolyamines. Cytotoxicity was low and seen only at concentrations well above those required for optimal transfection. Delivery of oligonucleotides into embryonic neurons was followed by using a fluorescent probe.Virtually all neurons showed nuclear labeling, with no toxic effects. The optimal PEI cation/anion balance for in vitro transfection is only slightly on the cationic side, which is advantageous for in vivo delivery. Indeed, intracerebral luciferase gene transfer into newborn mice gave results comparable (for a given amount of DNA) to the in vitro transfection of primary rat brain endothelial cells or chicken embryonic neurons. Together, these properties make PEI a promising vector for gene therapy and an outstanding core for the design of more sophisticated devices. Our hypothesis is that its efficiency relies on extensive lysosome buffering that protects DNA from nuclease degradation, and consequent lysosomal swelling and rupture that provide an escape mechanism for the PEI/DNA particles. Nonviral gene-delivery techniques remain several orders of magnitude behind viral vectors when compared on the basis of the mean number of gene copies needed to transfect a cell. Despite this limitation, plasmid-mediated transfection has the major advantage that it raises none of the concerns of biological vectors for human therapy. Thus, much effort is presently devoted to improving nonviral techniques (1). Indeed, the advent of gene therapy has provided the impetus for improving, by appropriate chemical design, the efficiency of classical transfection agents such as cationic polymers (DEAE-dextran, Polybrene, polylysine) or inorganic aggregates (e.g., calcium phosphate). Various other polycationic cores have been developed, whether macromolecules (for review, see refs. 2-4), amphiphilic aggregates (for review, see ref. 5), or mixtures of both (6, 7), all of which ionically condense plasmid DNA and bind to the cell surface. Additional viral-like molecular properties have been added to these particles (2-9) to promote receptor-mediated endocytosis, fusogenicity, and karyophily. Among the cationic cores described so far, two are constitutively efficient gene-delivery agents without any extra virusderived function or lysosomotropic additive. Polyamidoamine cascade polymers (4) and lipopolyamines (5, 10-12), although quite different in chemical structures, ...

A STING-activating nanovaccine for cancer immunotherapy

“…Once taken up by cells, these nanoparticles can buffer the luminal pH of endocytic organelles at specific pH values 7 . Inspired by “proton sponge” polymers for cytosolic delivery of biologics 8 and small nanoparticle size for lymph node targeting, we performed an in vivo screening of UPS nanoparticles to evaluate their abilities in generating cytotoxic T lymphocyte (CTL) response. The UPS library consists of copolymers containing tertiary amines with linear or cyclic side chains (Fig.…”

Glycosylation-mediated targeting of carriers

“…Of these, the release of pDNA from endosomes/lysosomes into the cytoplasm after internalization is a major obstacle for efficient gene expression in hepatocytes. Therefore, manipulation of this process by functional peptides or the use of PEI with pH buffering functions in lysosomes [53] by galactosylated macromolecules might allow high transfection rates in vivo [40,49,[54][55][56]. Our group developed a mannosylated macromolecular carrier system for cell-selective targeting based on dextran, which has high solubility, an abundance of hydroxyl groups for chemical modification, low immunogenicity, and is used in the clinic as a plasma expander [2].…”

Nanomedicine in the diagnosis and therapy of neurodegenerative disorders

“…Examples include polymeric micelles [118,[123][124][125][126], DNA/polycation complexes ("polyplexes") [127][128][129][130][131][132][133][134][135], block ionomer complexes [116,[136][137][138][139], nanogels [140][141][142][143][144] and others. Of these materials, polymeric micelles have been evaluated in human trials for the delivery of anti-cancer agents [145][146][147][148][149].…”