Immunogenicity of intrathecal plasmid gene delivery: cytokine release and effects on transgene expression.
BACKGROUND
One method for the delivery of therapeutic proteins to the spinal cord is to inject nonviral gene vectors including plasmid DNA into the cerebrospinal fluid (CSF) that surrounds the spinal cord (intrathecal space). This approach has produced therapeutic benefits in animal models of disease and several months of protein expression; however, there is little information available on the immune response to these treatments in the intrathecal space, the relevance of plasmid CpG sequences to any plasmid-induced immune response, or the effect of this immune response on transgene expression.


METHODS
In the present study, coding or noncoding plasmids were delivered to the intrathecal space of the lumbar spinal region in rats. Lumbosacral CSF was then collected at various time points afterwards for monitoring of cytokines and transgene expression.


RESULTS
This work demonstrates, for the first time, increased tumor necrosis factor-alpha and interleukin-1 in response to intrathecal plasmid vector injection and provides evidence indicating that this response is largely absent in a CpG-depleted vector. Transgene expression in the CSF is not significantly affected by this immune response. Expression after intrathecal plasmid injection is variable across rats but correlates with the amount of tissue associated plasmid and is increased by disrupting normal CSF flow.


CONCLUSIONS
The data obtained in the present study indicate that plasmid immunogenicity may affect intrathecal plasmid gene therapy safety but not transgene expression in the CSF. Furthermore, the development of methods to prevent loss of plasmid via CSF flow out of the central nervous system through the injection hole and/or natural outflow routes may increase intrathecal plasmid gene delivery efficacy.
Hughes
Travis S
TS
Department of Molecular, Cellular and Developmental Biology, University of CO, Boulder, CO, USA.
Langer
Stephen J
SJ
Virtanen
Salla I
SI
Chavez
Raymond A
RA
Watkins
Linda R
LR
Milligan
Erin D
ED
Leinwand
Leslie A
LA
D000818:Animals; D002460:Cell Line; D018899:CpG Islands / Q000235:genetics*; D016207:Cytokines / Q000378:metabolism*; D015870:Gene Expression; D015316:Genetic Therapy; D006801:Humans; D007278:Injections, Spinal; D008297:Male; D010957:Plasmids; D051381:Rats; D017207:Rats, Sprague-Dawley; D051217:Toll-Like Receptor 9 / Q000235:genetics*; D014162:Transfection; D019076:Transgenes
D016207:Cytokines; C495371:TLR9 protein, human; D051217:Toll-Like Receptor 9