Novel Semliki Forest virus vectors with reduced cytotoxicity and temperature sensitivity for long-term enhancement of transgene expression

Kenneth Lundstrom, Alessandra Abenavoli, Antonio Malgaroli, Markus U. Ehrengruber

Research output: Contribution to journalArticle

Abstract

Alphaviral vectors inhibit host cell protein synthesis and are cytotoxic. To overcome these limitations, we modified the nonstructural protein-2 (nsP2) gene in the Semliki Forest virus (SFV) vector, pSFV1. Packaging of SFV replicons with two point mutations in nsP2 resulted in high-titer recombinant SFV(PD) particles. SFV(PD) led to more efficient host cell protein synthesis, exhibited reduced cytotoxicity and improved cell survival, and allowed greater and prolonged transgene expression than the original vector, SFV. In dissociated hippocampal neurons and organotypic rat hippocampal slices, SFV(PD) infection preserved neuronal morphology and synaptic function more efficiently than SFV. Combination of the two point mutations with a replication-persistent mutation in nsP2 resulted in a highly temperature-sensitive vector, SFV(PD713P), which efficiently transduced neurons in hippocampal slice cultures. At 31 °C, SFV(PD713P) allowed continuous transgene expression in BHK cells, at amounts comparable to SFV(PD). These new SFV mutants are expected to substantially broaden the application of alphaviral vectors in neurons and other mammalian cells.

Original languageEnglish
Pages (from-to)202-209
Number of pages8
JournalMolecular Therapy
Volume7
Issue number2
DOIs
Publication statusPublished - Feb 1 2003

Keywords

  • Alphavirus
  • Expression vector
  • Green fluorescent protein (GFP)
  • Hippocampus
  • Metabolic labeling
  • Neuron
  • Slice culture
  • Synapse staining

ASJC Scopus subject areas

  • Molecular Biology

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