A Polyethylenimine-Linoleic Acid Conjugate for Antisense Oligonucleotide Delivery

Jing Xie,Bryant C Yung,Robert J Lee,Chenguang Zhou,Lesheng Teng,Yang Liu,Zhaogang Yang

doi:10.1155/2013/710502

Jing Xie, Bryant C Yung + Show 5 more

Open Access

https://doi.org/10.1155/2013/710502

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Abstract

A novel antisense oligonucleotide (ASO) carrier, polyethylenimine conjugated to linoleic acid (PEI-LA), was synthesized and evaluated for delivery of LOR-2501 to tumor cells. LOR-2501 is an ASO targeting ribonucleotide reductase R1 subunit (RRM1). In this study, PEI-LA was synthesized by reacting PEI (Mw ~ 800) with linoleoyl chloride. Gel retardation assay showed complete complexation between PEI-LA and LOR-2501 at N/P ratio above 8. No significant cytotoxicity was observed with these complexes at the tested dosage levels. Interestingly, at N/P ratio of >6, levels of cellular uptake of PEI-LA/LOR-2501 were double that of PEI/LOR-2501 complexes of the same N/P ratio. PEI-LA/LOR-2501 induced downregulation of 64% and 70% of RRM1 at mRNA and protein levels, respectively. The highest transfection activity was shown by PEI-LA/LOR-2501 complexes at N/P ratio of 10. Finally, using pathway specific inhibitors, clathrin-mediated endocytosis was shown to be the principle mechanism of cellular internalization of these complexes. In conclusion, PEI-LA is a promising agent for the delivery of ASOs and warrants further investigation.

Highlights

Antisense oligonucleotide (ASO) therapy is an emerging therapeutic modality for the treatment of human diseases, including cancer [1,2,3]
LOR-2501 is a 20-mer phosphorothioate ASO targeting the R1 subunit of ribonucleotide reductase [13], an enzyme associated with drug resistance
Degree of complexation between polyethylenimine conjugated to linoleic acid (PEI-LA) and LOR-2501 was measured by an agarose gel retardation assay

Summary

Introduction

Antisense oligonucleotide (ASO) therapy is an emerging therapeutic modality for the treatment of human diseases, including cancer [1,2,3]. LOR-2501 has been studied in a phase I clinical trial in 2006 for the treatment of prostate cancer [13, 14]. High molecular weight (25 kDa) PEI has frequently been used for gene delivery [18,19,20]. It is fairly cytotoxic [21,22,23]. Low molecular weight (∼800 Da) PEI demonstrates much lower cytotoxicity but is much less active in transfection [22, 24, 25]. Previous studies have shown that conjugating PEI to a lipophilic moiety greatly improved its transfection activity [26, 27].

Results

Discussion

Conclusion