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A plasmid-based self-amplifying Sindbis virus vector

H Herweijer1, J S Latendresse, P Williams

  • 1Department of Pediatrics and Medical Genetics, University of Wisconsin, Madison 53705, USA.

Human Gene Therapy
|September 1, 1995
PubMed
Summary
This summary is machine-generated.

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This study introduces a novel Sindbis virus-based expression vector for high-level gene expression. This self-amplifying vector shows superior performance in vitro and transient, safe gene delivery in vivo for gene therapy applications.

Area of Science:

  • Molecular Biology
  • Virology
  • Biotechnology

Background:

  • Traditional expression vectors often yield suboptimal transgene expression levels.
  • Developing efficient and safe gene delivery systems is crucial for research and therapeutic applications.
  • Sindbis virus, a (+)-strand RNA virus, possesses a unique replication mechanism that can be harnessed for enhanced gene expression.

Purpose of the Study:

  • To engineer a self-amplifying eukaryotic expression vector based on Sindbis virus.
  • To evaluate the expression efficiency of this novel vector in mammalian cell lines and primary cells.
  • To assess the in vitro and in vivo performance of the Sindbis virus-based vector for transgene expression and its potential in gene therapy.

Main Methods:

  • A recombinant Sindbis virus cDNA genome was constructed under the control of a Rous sarcoma virus long terminal repeat (RSV LTR) promoter.

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  • The plasmid construct was transfected into various mammalian cell lines (3T3, HepG2, 293) and primary rat myoblasts.
  • Luciferase reporter gene expression was measured to quantify vector performance.
  • In vivo expression was assessed following injection into mouse quadriceps.

    • Main Results:

    • The Sindbis virus vector achieved significantly higher expression levels compared to a conventional RSV expression vector in differentiated myotubes (up to 200-fold increase).
    • High expression was observed in various cell lines and primary myoblasts.
    • Transient in vivo expression was detected after plasmid DNA injection in mice, becoming undetectable by day 16.

    Conclusions:

    • The Sindbis virus-based self-amplifying vector enables high-level transgene expression both in vitro and in vivo.
    • The transient nature of in vivo expression suggests potential for safe, short-term gene product delivery in gene therapy.
    • This vector system offers a valuable tool for studying Sindbis virus and other RNA viruses, and for advancing gene delivery technologies.