Construction and characterization of encapsidated poliovirus replicons that express biologically active murine interleukin-2.
Poliovirus genomes have been constructed in which the capsid genes have been substituted with the murine gene encoding interleukin-2 (IL-2) (referred to as replicons). One replicon contained the gene for IL-2 in place of the poliovirus capsid VP2 and VP3 genes, and a second replicon was constructed that contained the murine IL-2 substituted for the poliovirus VP3 and VP1 genes. The IL-2 genes were cloned into the replicon so as to maintain the translational reading frame with the remaining poliovirus proteins. Transfection of either replicon into cells resulted in the expression of replicon-encoded proteins and replication of replicon RNA. Using a procedure developed in this laboratory, we have encapsidated these replicons into authentic polio virions by passaging the replicons in the presence of a recombinant vaccinia virus, VVP1, which expresses the capsid precursor, P1, protein. Using a quantitative immunoassay, we determined that the majority of the IL-2 produced remained intracellular, with approximately 1%-2% released from the infected cells, and that the IL-2 was biologically active. The results of these studies demonstrate the utility of poliovirus replicons for expression of small bioactive molecules and are discussed with respect to future applications as immune adjuvants as well as potential new tumor therapies.
Basak
S
S
Department of Microbiology, University of Alabama at Birmingham, 35294, USA.
McPherson
S
S
Kang
S
S
Collawn
J F
JF
Morrow
C D
CD
D000818:Animals; D002213:Capsid / Q000235:genetics*; D004274:DNA, Recombinant / Q000235:genetics; D015967:Gene Expression Regulation, Viral / Q000502:physiology*; D016679:Genome, Viral; D006023:Glycoproteins / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D007376:Interleukin-2 / Q000235:genetics*; D051379:Mice; D017955:Poliovirus / Q000235:genetics*; D021382:Protein Sorting Signals / Q000235:genetics; D012093:Replicon / Q000235:genetics*
D004274:DNA, Recombinant; D006023:Glycoproteins; D007376:Interleukin-2; D021382:Protein Sorting Signals