Protection from experimental endotoxemia by a recombinant adeno-associated virus encoding interleukin 10.
BACKGROUND
Interleukin 10 (IL-10) is a homodimeric cytokine that shows considerable clinical promise. Adeno-associated virus (AAV) vectors appear increasingly useful for in vivo gene-transfer applications.


METHODS
A recombinant AAV type 2 vector encoding human IL-10 (rAAVhIL10) was constructed by using an adenoviral-free, three-plasmid co-transfection. Cytokine production was measured by using an enzyme-linked immunosorbent assay. Endotoxic shock was induced by lipopolysaccharide (LPS) injection.


RESULTS
As media from rAAVhIL10-infected COS cells caused a dose-dependent blockade of IL-12 secretion from spleen cells of IL-10 knockout (KO) mice challenged with Brucella abortus, it was clear that vector-derived hIL-10 was biologically active in vitro. Intravenous or intramuscular administration of relatively modest levels of rAAVhIL10 (10(10) genomes) to IL-10 KO mice resulted in hIL-10 secretion into the bloodstream, which, at 8 weeks, gave median serum levels of 0.9 and 0.45 pg/ml, respectively. Acute endotoxic shock led to a 33% mortality rate, and severe morbidity, in control IL-10 KO mice, whereas no mortality and little morbidity were seen in IL-10 KO mice given rAAVhIL10 7 weeks earlier.


CONCLUSIONS
The findings demonstrate that a modest dose of rAAVhIL10 administered in vivo provides long-term protection against LPS-induced endotoxic shock in a murine model. Thus, this vector may be useful for clinical applications requiring sustained IL-10 expression, for example in the treatment of several autoimmune diseases.
Yamano
S
S
Gene Therapy and Therapeutics Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD 20892, USA. syamano@dir.nidcr.nih.gov
Scott
D E
DE
Huang
L Y
LY
Mikolajczyk
M
M
Pillemer
S R
SR
Chiorini
J A
JA
Golding
B
B
Baum
B J
BJ
D000818:Animals; D019556:COS Cells / Q000378:metabolism; D000229:Dependovirus / Q000235:genetics*; D019446:Endotoxemia / Q000139:chemically induced / Q000235:genetics / Q000517:prevention & control*; D015870:Gene Expression; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D005822:Genetic Vectors; D007167:Immunotherapy; D016753:Interleukin-10 / Q000235:genetics* / Q000378:metabolism; D018664:Interleukin-12 / Q000037:antagonists & inhibitors; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D014409:Tumor Necrosis Factor-alpha; D016753:Interleukin-10; D018664:Interleukin-12