Peptide-oligonucleotide phosphorothioate conjugates with membrane translocation and nuclear localization properties

Area of Science:

• Medicinal Chemistry
• Molecular Biology
• Bioconjugation Chemistry

Background:

• Antisense oligonucleotides (ASOs) are promising therapeutic agents.
• Delivery of ASOs into target cells remains a significant challenge.
• Peptide-oligonucleotide conjugates offer a potential strategy for enhanced cellular uptake.

Purpose of the Study:

• To synthesize and characterize peptide-oligonucleotide phosphorothioate conjugates.
• To evaluate the cellular uptake, intracellular distribution, and antisense activity of these conjugates.
• To investigate the potential of peptide-mediated delivery for ASO therapeutics.

Main Methods:

• Synthesis of 18 peptide-oligonucleotide phosphorothioate conjugates.
• Characterization using electrospray ionization mass spectrometry.
• Cellular uptake and localization studies using fluorescein-labeled conjugates.
• Assessment of antisense effect in living cells.

Main Results:

• Conjugates were prepared in good yield and characterized.
• Conjugates demonstrated membrane translocation and nuclear localization.
• Efficient intracellular penetration was observed.
• No significant antisense effect was detected.
• Fluorescein-labeled conjugates were found to be trapped in endosomes.

Conclusions:

• Peptide-oligonucleotide conjugates can facilitate cellular entry but do not inherently confer antisense activity.
• Endosomal entrapment is a major barrier to the efficacy of these conjugates.
• Strategies to escape endosomes are crucial for developing effective peptide-ASO therapeutics.