Using inositol as a biocompatible ligand for efficient transgene expression

Abstract

Transgene transfection techniques using cationic polymers such as polyethylenimines (PEIs) and PEI derivatives as gene vectors have shown efficacy, although they also have shortcomings. PEIs have decent DNA-binding capability and good cell internalization performance, but they cannot deliver gene payloads very efficiently to cell nuclei. In this study, three hyperbranched polyglycerol-polyethylenimine (PG6-PEI) polymers conjugated with myo-inositol (INO) molecules were developed. The three resulting PG6-PEI-INO polymers have an increased number of INO ligands per molecule. PG6-PEI-INO 1 had only 14 carboxymethyl INO (CMINO) units per molecule. PG6-PEI-INO 2 had approximately 130 CMINO units per molecule. PG6-PEI-INO 3 had as high as 415 CMINO units approximately. Mixing PG6-PEI-INO polymers with DNA produced compact nanocomposites. We then performed localization studies using fluorescent microscopy. As the number of conjugated inositol ligands increased in PG6-PEI-INO polymers, there was a corresponding increase in accumulation of the polymers within 293T cell nuclei. Transfection performed with spherical 293T cells yielded 82% of EGFP-positive cells when using PG6-PEI-INO 3 as the vehicle. Studies further revealed that extracellular adenosine triphosphate (eATP) can inhibit the transgene efficiency of PG6-PEI-INO polymers, as compared with PEI and PG6-PEI that were not conjugated with inositol. Our work unveiled the possibility of using inositol as an effective ligand for transgene expression.

Keywords:

• myo-inositol
• nuclear localization
• biocompatibility
• polyglycerol-polyethylenimine
• hyperbranched polymers
• extracellular ATP

Acknowledgments

We thank the National Nature Science Foundation of China (31300650 and 31270790), the Key Project of Fujian Province (2013N0039, 2012Y0070), the Nature Science Foundation of Fujian Province (2013J01151), the Science Foundation for Post Doctorate Research of Wuhan University of China (203-180462), the National Thousand Talents Program of China, and the Scientific Research Starting Foundation for Returned Overseas Chinese Scholars of State Human Resource Ministry of China for support on this work.

Disclosure

The authors report no conflicts of interest in this work.