Bactericidal antisense effects of peptide–PNA conjugates

Abstract

Antisense peptide nucleic acids (PNAs) can specifically inhibit Escherichia coli gene expression and growth and hold promise as anti-infective agents and as tools for microbial functional genomics. Here we demonstrate that chemical modification improves the potency of standard PNAs. We show that 9- to 12-mer PNAs, especially when attached to the cell wall/membrane-active peptide KFFKFFKFFK, provide improvements in antisense potency in E. coli amounting to two orders of magnitude while retaining target specificity. Peptide–PNA conjugates targeted to ribosomal RNA (rRNA) and to messenger RNA (mRNA) encoding the essential fatty acid biosynthesis protein Acp prevented cell growth. The anti-acpP PNA at 2 μM concentration cured HeLa cell cultures noninvasively infected with E. coli K12 without any apparent toxicity to the human cells. These results indicate that peptides can be used to carry antisense PNA agents into bacteria. Such peptide–PNA conjugates open exciting possibilities for anti-infective drug development and provide new tools for microbial genetics.

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Figure 1: (A) Chemical structure of DNA and PNA oligomers, with the bases indicated as b.
Figure 2: LacZ expression and inhibition in E. coli with a range of different-sized PNAs.
Figure 3: LacZ expression and inhibition in E. coli with PNA, free peptide, and a peptide–PNA conjugate.
Figure 4: E. coli K12 growth and inhibition in MH broth by an anti-acpP peptide–PNA conjugate.
Figure 5: Bactericidal antisense effects of an anti-acpP PNA against E. coli.
Figure 6: HeLa cell culture, noninvasive E. coli infection, and antimicrobial PNA treatment.

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Acknowledgements

This research was supported by the Danish Biotechnology Program, Pharmacia Corporation and the Swedish Foundation for Strategic Research

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Correspondence to Liam Good or Peter E. Nielsen.

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Good, L., Awasthi, S., Dryselius, R. et al. Bactericidal antisense effects of peptide–PNA conjugates. Nat Biotechnol 19, 360–364 (2001). https://doi.org/10.1038/86753

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