Improved gene delivery to K-562 leukemia cells by lipoic acid modified block copolymer micelles

Friederike Richter,Stephanie Hoeppener,Thomas Ernst,Franziska Hausig,Liam Martin,Prosper Mapfumo,Johannes C. Brendel,Jana I. Solomun,Anja Traeger,Jochen J. Frietsch

doi:10.1186/s12951-021-00801-y

Friederike Richter, Stephanie Hoeppener + Show 8 more

Open Access

https://doi.org/10.1186/s12951-021-00801-y

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Abstract

Although there has been substantial progress in the research field of gene delivery, there are some challenges remaining, e.g. there are still cell types such as primary cells and suspension cells (immune cells) known to be difficult to transfect. Cationic polymers have gained increasing attention due to their ability to bind, condense and mask genetic material, being amenable to scale up and highly variable in their composition. In addition, they can be combined with further monomers exhibiting desired biological and chemical properties, such as antioxidative, pH- and redox-responsive or biocompatible features. By introduction of hydrophobic monomers, in particular as block copolymers, cationic micelles can be formed possessing an improved chance of transfection in otherwise challenging cells. In this study, the antioxidant biomolecule lipoic acid, which can also be used as crosslinker, was incorporated into the hydrophobic block of a diblock copolymer, poly{[2-(dimethylamino)ethyl methacrylate]101-b-[n-(butyl methacrylate)124-co-(lipoic acid methacrylate)22]} (P(DMAEMA101-b-[nBMA124-co-LAMA22])), synthesized by RAFT polymerization and assembled into micelles (LAMA-mic). These micelles were investigated regarding their pDNA binding, cytotoxicity mechanisms and transfection efficiency in K-562 and HEK293T cells, the former representing a difficult to transfect, suspension leukemia cell line. The LAMA-mic exhibited low cytotoxicity at applied concentrations but demonstrated superior transfection efficiency in HEK293T and especially K-562 cells. In-depth studies on the transfection mechanism revealed that transfection efficiency in K-562 cells does not depend on the specific oncogenic fusion gene BCR-ABL alone. It is independent of the cellular uptake of polymer-pDNA complexes but correlates with the endosomal escape of the LAMA-mic. A comparison of the transfection efficiency of the LAMA-mic with structurally comparable micelles without lipoic acid showed that lipoic acid is not solely responsible for the superior transfection efficiency of the LAMA-mic. More likely, a synergistic effect of the antioxidative lipoic acid and the micellar architecture was identified. Therefore, the incorporation of lipoic acid into the core of hydrophobic-cationic micelles represents a promising tailor-made transfer strategy, which can potentially be beneficial for other difficult to transfect cell types.

Highlights

Non-viral gene therapy has become an important research field, with the aim of finding new methods for the treatment of diseases like cancer or genetic disorders or for the development of vaccines [1, 2]
Studies have shown that lipoic acid exhibits redox properties, they can be used as a crosslinking agent when incorporated in micelles or nanoparticles [52]
The polymers were synthesized by reversible addition fragmentation chain transfer (RAFT) polymerization with (4-cyano pentanoic acid)yl ethyl trithiocarbonate (CPAETC) as a chain transfer agent (CTA) and 4,4′-azobis(4-cyanovaleric acid) (ACVA) as the initiator

Summary

Introduction

Non-viral gene therapy has become an important research field, with the aim of finding new methods for the treatment of diseases like cancer or genetic disorders or for the development of vaccines [1, 2]. Polymeric nanocarriers could be a promising strategy to interact more efficiently with immune cells and enhance the intracellular concentration of active agents to counteract mechanisms such as drug resistance or poor response to treatment [14, 15]. Wagner could already show successful transfection of pDNA with poly(ethylene imine) (PEI) only in combination with transferrin [16,17,18] Another promising strategy was demonstrated by H. Uludağ and coworkers who statistically incorporated hydrophobic moieties into polymeric nanocarriers that were beneficial for the transfection of siRNA and subsequent knockdown of the BCR-ABL fusion protein in the CML cell line K-562 [19,20,21]

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