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Peptide nanoparticle delivery of charge-neutral splice-switching morpholino oligonucleotides.

Järver P, Zaghloul EM, Arzumanov AA, Saleh AF, McClorey G, Hammond SM, Hällbrink M, Langel Ü, Smith CI, Wood MJ, Gait MJ, El Andaloussi S - Nucleic Acid Ther (2015)

Bottom Line: Negatively charged oligonucleotides, such as 2'-O-Methyl RNA and locked nucleic acids have in recent years been delivered successfully into cells through complex formation with cationic polymers, peptides, liposomes, or similar nanoparticle delivery systems.However, due to the lack of electrostatic interactions, this promising delivery method has been unsuccessful to date using charge-neutral oligonucleotide analogs.To our knowledge, this is the first study to show delivery through complex formation of biologically active charge-neutral oligonucleotides by cationic peptides.

View Article: PubMed Central - PubMed

Affiliation: 1 Medical Research Council , Laboratory of Molecular Biology, Cambridge Biomedical Campus, Cambridge, United Kingdom .

ABSTRACT
Oligonucleotide analogs have provided novel therapeutics targeting various disorders. However, their poor cellular uptake remains a major obstacle for their clinical development. Negatively charged oligonucleotides, such as 2'-O-Methyl RNA and locked nucleic acids have in recent years been delivered successfully into cells through complex formation with cationic polymers, peptides, liposomes, or similar nanoparticle delivery systems. However, due to the lack of electrostatic interactions, this promising delivery method has been unsuccessful to date using charge-neutral oligonucleotide analogs. We show here that lipid-functionalized cell-penetrating peptides can be efficiently exploited for cellular transfection of the charge-neutral oligonucleotide analog phosphorodiamidate morpholino. The lipopeptides form complexes with splice-switching phosphorodiamidate morpholino oligonucleotide and can be delivered into clinically relevant cell lines that are otherwise difficult to transfect while retaining biological activity. To our knowledge, this is the first study to show delivery through complex formation of biologically active charge-neutral oligonucleotides by cationic peptides.

Show MeSH
Exon skipping activity in H2K mdx myotubes by 1 μM PMO formulated with selected lipopeptides at different molar ratios (2:1 to 7:1). Results are from at least three experiments performed in duplicate.
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f6: Exon skipping activity in H2K mdx myotubes by 1 μM PMO formulated with selected lipopeptides at different molar ratios (2:1 to 7:1). Results are from at least three experiments performed in duplicate.

Mentions: To see if the relative amounts of lipopeptides affect the SSO activity, they were incubated with PMO at ratios between 2 and 7 prior to cell treatments. For most lipopeptides, an increased ratio over PMO was found to improve splice-switching in the DMD model muscle cells (Fig. 6). The increase seems to be more pronounced in the peptides containing less positively charged amino acids such as the secondary amphipathic peptide St-KL4 (four lysines). In contrast for some peptides such as the more cationic St-RXR3 (six Arginines), the increased peptide ratio over PMO did not improve, but instead decreased, splice-switching activity (Fig. 6).


Peptide nanoparticle delivery of charge-neutral splice-switching morpholino oligonucleotides.

Järver P, Zaghloul EM, Arzumanov AA, Saleh AF, McClorey G, Hammond SM, Hällbrink M, Langel Ü, Smith CI, Wood MJ, Gait MJ, El Andaloussi S - Nucleic Acid Ther (2015)

Exon skipping activity in H2K mdx myotubes by 1 μM PMO formulated with selected lipopeptides at different molar ratios (2:1 to 7:1). Results are from at least three experiments performed in duplicate.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4376484&req=5

f6: Exon skipping activity in H2K mdx myotubes by 1 μM PMO formulated with selected lipopeptides at different molar ratios (2:1 to 7:1). Results are from at least three experiments performed in duplicate.
Mentions: To see if the relative amounts of lipopeptides affect the SSO activity, they were incubated with PMO at ratios between 2 and 7 prior to cell treatments. For most lipopeptides, an increased ratio over PMO was found to improve splice-switching in the DMD model muscle cells (Fig. 6). The increase seems to be more pronounced in the peptides containing less positively charged amino acids such as the secondary amphipathic peptide St-KL4 (four lysines). In contrast for some peptides such as the more cationic St-RXR3 (six Arginines), the increased peptide ratio over PMO did not improve, but instead decreased, splice-switching activity (Fig. 6).

Bottom Line: Negatively charged oligonucleotides, such as 2'-O-Methyl RNA and locked nucleic acids have in recent years been delivered successfully into cells through complex formation with cationic polymers, peptides, liposomes, or similar nanoparticle delivery systems.However, due to the lack of electrostatic interactions, this promising delivery method has been unsuccessful to date using charge-neutral oligonucleotide analogs.To our knowledge, this is the first study to show delivery through complex formation of biologically active charge-neutral oligonucleotides by cationic peptides.

View Article: PubMed Central - PubMed

Affiliation: 1 Medical Research Council , Laboratory of Molecular Biology, Cambridge Biomedical Campus, Cambridge, United Kingdom .

ABSTRACT
Oligonucleotide analogs have provided novel therapeutics targeting various disorders. However, their poor cellular uptake remains a major obstacle for their clinical development. Negatively charged oligonucleotides, such as 2'-O-Methyl RNA and locked nucleic acids have in recent years been delivered successfully into cells through complex formation with cationic polymers, peptides, liposomes, or similar nanoparticle delivery systems. However, due to the lack of electrostatic interactions, this promising delivery method has been unsuccessful to date using charge-neutral oligonucleotide analogs. We show here that lipid-functionalized cell-penetrating peptides can be efficiently exploited for cellular transfection of the charge-neutral oligonucleotide analog phosphorodiamidate morpholino. The lipopeptides form complexes with splice-switching phosphorodiamidate morpholino oligonucleotide and can be delivered into clinically relevant cell lines that are otherwise difficult to transfect while retaining biological activity. To our knowledge, this is the first study to show delivery through complex formation of biologically active charge-neutral oligonucleotides by cationic peptides.

Show MeSH