In vitro resistance selection and in vivo efficacy of morpholino oligomers against west nile virus

Tia S. Deas, Corey J. Bennett, Susan A. Jones, Mark Tilgner, Ping Ren, Melissa J. Behr, David A. Stein, Patrick L. Iversen, Laura D. Kramer, Kristen A. Bernard, Pei-Yong Shi

Research output: Contribution to journalArticle

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Abstract

We characterize in vitro resistance to and demonstrate the in vivo efficacy of two antisense phosphorodiamidate morpholino oligomers (PMOs) against West Nile virus (WNV). Both PMOs were conjugated with an Arg-rich peptide. One peptide-conjugated PMO (PPMO) binds to the 5′ terminus of the viral genome (5′-end PPMO); the other targets an essential 3′ RNA element required for genome cyclization (3′ conserved sequence I [3′ CSI] PPMO). The 3′ CSI PPMO displayed a broad spectrum of antiflavivirus activity, suppressing WNV, Japanese encephalitis virus, and St. Louis encephalitis virus, as demonstrated by reductions in viral titers of 3 to 5 logs in cell cultures, likely due to the absolute conservation of the 3′ CSI PPMO-targeted sequences among these viruses. The selection and sequencing of PPMO-resistant WNV showed that the 5′-end-PPMO-resistant viruses contained two to three mismatches within the PPMO-binding site whereas the 3′ CSI PPMO-resistant viruses accumulated mutations outside the PPMO-targeted region. The mutagenesis of a WNV infectious clone demonstrated that the mismatches within the PPMO-binding site were responsible for the 5′-end PPMO resistance. In contrast, a U insertion or a G deletion located within the 3′-terminal stem-loop of the viral genome was the determinant of the 3′ CSI PPMO resistance. In a mouse model, both the 5′-end and 3′ CSI PPMOs (administered at 100 or 200 μg/day) partially protected mice from WNV disease, with minimal to no PPMO-mediated toxicity. A higher treatment dose (300 μg/day) caused toxicity. Unconjugated PMOs (3 mg/day) showed neither efficacy nor toxicity, suggesting the importance of the peptide conjugate for efficacy. The results suggest that a modification of the peptide conjugate composition to reduce its toxicity yet maintain its ability to effectively deliver PMO into cells may improve PMO-mediated therapy.

Original languageEnglish (US)
Pages (from-to)2470-2482
Number of pages13
JournalAntimicrobial Agents and Chemotherapy
Volume51
Issue number7
DOIs
StatePublished - Jul 2007
Externally publishedYes

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Morpholinos
West Nile virus
Peptides
Conserved Sequence
In Vitro Techniques
Viral Genome
Viruses
Japanese Encephalitis Virus
St. Louis Encephalitis Viruses
Binding Sites

ASJC Scopus subject areas

  • Pharmacology (medical)

Cite this

In vitro resistance selection and in vivo efficacy of morpholino oligomers against west nile virus. / Deas, Tia S.; Bennett, Corey J.; Jones, Susan A.; Tilgner, Mark; Ren, Ping; Behr, Melissa J.; Stein, David A.; Iversen, Patrick L.; Kramer, Laura D.; Bernard, Kristen A.; Shi, Pei-Yong.

In: Antimicrobial Agents and Chemotherapy, Vol. 51, No. 7, 07.2007, p. 2470-2482.

Research output: Contribution to journalArticle

Deas, TS, Bennett, CJ, Jones, SA, Tilgner, M, Ren, P, Behr, MJ, Stein, DA, Iversen, PL, Kramer, LD, Bernard, KA & Shi, P-Y 2007, 'In vitro resistance selection and in vivo efficacy of morpholino oligomers against west nile virus', Antimicrobial Agents and Chemotherapy, vol. 51, no. 7, pp. 2470-2482. https://doi.org/10.1128/AAC.00069-07
Deas, Tia S. ; Bennett, Corey J. ; Jones, Susan A. ; Tilgner, Mark ; Ren, Ping ; Behr, Melissa J. ; Stein, David A. ; Iversen, Patrick L. ; Kramer, Laura D. ; Bernard, Kristen A. ; Shi, Pei-Yong. / In vitro resistance selection and in vivo efficacy of morpholino oligomers against west nile virus. In: Antimicrobial Agents and Chemotherapy. 2007 ; Vol. 51, No. 7. pp. 2470-2482.
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AU - Stein, David A.

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