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Efficient Cytoplasmic Delivery of Antisense Probes Assisted by Cyclized-Peptide-Mediated Photoinduced Endosomal

Xiaohong Tan1, Marcel P Bruchez1,2, Bruce A Armitage1

  • 1Departments of Chemistry, Center for Nucleic Acids Science and Technology, Carnegie Mellon University, 4400 Fifth Avenue, Pittsburgh, PA, 15213, USA.

Chembiochem : a European Journal of Chemical Biology
|November 20, 2018
PubMed

Insights

Cyclized TAT peptides (Cyc-TAT) enhance cellular uptake and endosomal escape of antisense oligonucleotides. Photoactivation significantly shortens treatment time for gene-targeted therapeutics while maintaining high efficacy.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Gene Therapy

Background:

  • Antisense oligonucleotides face challenges in intracellular delivery and endosomal release.
  • Cyclized TAT peptide (Cyc-TAT) shows improved cellular entry compared to linear analogues.
  • Cyc-TAT can be modified to co-deliver nucleic acid probes for targeted gene inhibition.

Purpose of the Study:

  • To investigate the enhanced endosomal release of cyclized TAT peptide (Cyc-TAT) using photoactivation.
  • To evaluate the efficiency of Cyc-TAT in delivering anti-telomerase gamma-peptide nucleic acid (γPNA) for gene therapy.
  • To reduce the required cell treatment time for effective gene silencing.

Main Methods:

  • Utilized a noncovalently cyclized TAT peptide (Cyc-TAT) with an overhanging γPNA sequence for co-delivery.
  • Employed fluorescence-labeled Cyc-TAT (Cyc-TAT(FAM)) to track endosomal escape after LED illumination (490 nm).
  • Assessed the inhibition of telomerase activity in A549 cells using the Cyc-TAT(FAM)/anti-telomerase γPNA complex.

Main Results:

  • Cyc-TAT demonstrated superior endosomal escape compared to linear TAT after light exposure.
  • The Cyc-TAT(FAM)/anti-telomerase γPNA complex effectively reduced telomerase activity by over 97% in A549 cells.
  • Photoactivation significantly enhanced endosomal release, reducing cell treatment time from 60 hours to 3 hours.

Conclusions:

  • Photoactivated endosomal release significantly improves the efficiency of Cyc-TAT-mediated gene delivery.
  • Cyc-TAT represents a promising platform for developing effective gene-targeted therapeutics with reduced treatment times.
  • This approach offers a strategy to overcome intracellular delivery barriers for antisense oligonucleotides.
Keywords:
anti-telomerase gammaPNAcyclized TATcytoplasmic deliveryphotoactivation

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