Naked DNA transfer of a high-expressing @entity1 factor IX ( @entity4682 ) plasmid yielded long-term (over 1 1/2 years) and therapeutic-level (0.5-2 microg/ml) gene expression of @entity4682 from @entity19 livers. The expression cassette contained a hepatic locus control region from the @entity30560 gene locus, an alpha1-anti-trypsin promoter, @entity4682 cDNA, a portion of the @entity4682 first intron, and a @entity291 growth hormone polyadenylation signal. In contrast, a @entity4682 plasmid containing the expression cassette without effective regulatory elements produced initially low-level gene expression that rapidly declined to undetectable levels. Southern analyses of the cellular DNA indicated that the majority of the input genome from either vector persisted as episomal forms of the original plasmids. Together with RT-PCR analyses of the transcripts, these data indicated that at least two processes are critical for sustained gene expression: persistence of vector DNA and transcriptional/posttranscriptional activation. @entity409 after partial hepatectomy resulted in a significant decline in transgene expression, further suggestive of decreased episomal plasmid maintenance rather than transgene integration. Transaminase levels and liver histology showed that rapid intravenous plasmid injection into @entity19 induced transient focal @entity99 (< 5% of hepatocytes), which was rapidly repaired within 3 to 10 days and resulted thereafter in histologically normal tissue. No significant differences were observed between rapid injection of plasmid and saline control solutions. Transient, very low level antibodies directed against @entity4682 did not prevent the circulation of therapeutic levels of the protein. Gene transfer of @entity4682 plasmid DNA into liver elicited neither transgene-specific cytotoxic effect nor long-term @entity137 . These results demonstrate that long-term expression of @entity4682 can be achieved by nonviral plasmid transfer and suggest that this occurs independent of integration.

Long-term and therapeutic-level hepatic gene expression of XXXX factor IX after naked plasmid transfer in vivo.

@entity1 :: ('9606', 'Species') :: ['human'] @entity137 :: ('MESH:D064420', 'Disease') :: ['toxicity'] @entity19 :: ('10090', 'Species') :: ['mice', 'mouse'] @entity409 :: ('MESH:D008107', 'Disease') :: ['Liver regeneration'] @entity4682 :: ('2158', 'Gene') :: ['hFIX'] @entity30560 :: ('281004(Tax:9913)', 'Gene') :: ['ApoE'] @entity99 :: ('MESH:D056486', 'Disease') :: ['acute liver damage'] @entity291 :: ('9913', 'Species') :: ['bovine']
@entity1 :: (9606,Species) :: ['human']
