Enhanced Gene Delivery and CRISPR/Cas9 Homology-Directed Repair in Serum by Minimally Succinylated Polyethylenimine

Nasir Uddin, Logan W. Warriner, Daniel W. Pack, Jason E. Derouchey

Research output: Contribution to journalArticlepeer-review

1 Scopus citations

Abstract

Gene therapy aims to treat patients by altering or controlling gene expression. The field of gene therapy has had increasing success in recent years primarily using viral-based approaches; however, there is still significant interest toward the use of polymeric materials due to their potential as flexible, low-cost scaffolds for gene delivery that do not suffer the mutagenesis and immunogenicity concerns of viral vectors. To address the challenges of efficiency and biocompatibility, a series of zwitterion-like polyethylenimine derivatives (zPEIs) were produced via the succinylation of 2-11.5% of polyethylenimine (PEI) amines. With increasing modification, zPEI polyplexes exhibited decreased serum-protein aggregation and dissociated more easily in the presence of a competitor polyanion when compared to unmodified PEI. Surprisingly, the gene delivery mediated in the presence of serum showed that succinylation of as few as 2% of PEI amines resulted in transgene expression 260- to 480-fold higher than that of unmodified PEI and 50- to 65-fold higher than that of commercial PEI-PEG2k in HEK293 and HeLa cells, respectively. Remarkably, the same zPEIs also produced 16-fold greater efficiency of CRISPR/Cas9 gene knock-in compared to unmodified PEI in the presence of serum. In addition, we show that 2% succinylation does not significantly decrease polymer/DNA binding ability or serum protein interaction to a significant extent, yet this small modification is still sufficient to provide a remarkable increase in transgene expression and gene knock-in in the presence of serum.

Original languageEnglish
Pages (from-to)3452-3463
Number of pages12
JournalMolecular Pharmaceutics
Volume18
Issue number9
DOIs
StatePublished - Sep 6 2021

Bibliographical note

Funding Information:
J.E.D. acknowledges financial support from the National Science Foundation (MCB-1453168).

Publisher Copyright:
© 2021 American Chemical Society.

Keywords

  • gene delivery
  • in-serum stability
  • nonviral vectors
  • polyampholytes
  • polymer chemistry
  • transfection efficiency

ASJC Scopus subject areas

  • Molecular Medicine
  • Pharmaceutical Science
  • Drug Discovery

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