One of the reasons why adenovirus has short term expression is due to the strong immune response that it generates. This anti-vector immunity usually limits the expression to ~1-2 weeks. It is my understanding that the short duration of expression is due to CTL mediated destruction of the transduced cells. Is this correct? I have read some papers stating that Ad can be used for short term expression of transgenes, but not mentioning that the reason for the short term expression is destruction of expressing cells.
Short term expression is one thing, short term expression because the expressing cells are dead is another. Can anyone shed any light on this?
Thanks,
John
Dear John,
Certainly, immune elimination of cells displaying viral epitopes is a major contribution to loss of expression, but isn't the only factor. There is also the contribution of cell turnover (depending upon tissue) since adenovirus vector is non-integrating and remains episomal. There is also the issue of genetic silencing, which depends upon the promoter and vector genome configuration, as well as cell type. "1st" and "2nd" generation adenovirus vectors carry a lot of the original viral genome, so viral genes can continue to be expressed. Helper dependent (i.e gutless) adenovirus vectors are less immunogenic and have been shown in some instances to achieve more prolonged expression, partly because they express no viral genes; the only bits of virus they carry into the cell, in theory, are the non-genetic components, which will be degraded. In this way, helper-dependent adenovirus vectors are more like AAV.
So, I guess, in summary, loss of expression is dependent upon the version of adenovirus vector, immune status of the recipient, cell type transduced, and payload configuration.
BW Simon
Hi John,
we compared AAV and Adenovirus-5 (2nd gen.) transduction in the lung of mice and found that Adenovirus not only induced classical inflammatory cytokines (IL1, IL6, TNFa...) but also upregulated MyD88 and TLR signaling components. We also found a significant increase in CD8+ T-cells in the lung lavage samples of AdV-treated animals, suggesting that indeed cytotoxic T-cells might be responsible for the loss of AdV-mediated transgene expression. We haven't looked into direct effects of the T-cells on AdV-transduced cells, though...
The paper is entitled "Modeling Pulmonary Disease Pathways Using Recombinant Adeno-Associated Virus 6.2."
Best wishes,
Ben
Thanks for the information. I know that Cardium is using an Ad vector (not HD-Ad) for short term expression of FGF4 in the heart, but I am not clear on whether the expression is short term because the cells are eliminated, or the expression is shut off for some other reason (methylation?). Does anyone know why the expression is short term?
Checking the website of Taxus Cardium it looks like they may use balloon catheter-isolated delivery to stretches of cardiac vasculature. Seems unlikely they'd be prepared to lose myocardium through cytotoxic T cell response or that would be majorly bad news Looks like the preclinical stuff was kicking around a decade ago (http://content.onlinejacc.org/article.aspx?articleid=1132725).
CMV promoter - maybe shutdown there?
HI, there has been a lot of literature on this, from the early gene therapy days, it was initially thought to be immune mediated (I am sure it has a contribution) but the majority of the effect is shutdown of the promoter. There was a really nice study from Bart de geest showing if you use an endogenous human promoter and intron you can get more than 6 months expression from 1st generation Ads in the liver, another in the brain showing the same thing with the synapsin promoter from James Uney. I wrote a review on it some years ago now which is a good place to get a handle on some of these (now old) papers. (it is in my publication list - Promoters and control elements: Designing expression cassettes for gene therapy - sorry not trying to promote myself, it concisely summarises other people's work) I have also created construct for the vasculature. I think it is possible to extend expression from 1st generation Ads, by adding introns to ensure correct nuclear processing, and by using human promoters. I have just made a composite endothelial promoter that I think is about as strong as CMV as a bit of a side project, not fully tested it out yet. Hope this helps...
Hi, Colleagues
All of the above are correct: (a) Immune system kills infected cells since they express foreign proteins even if E1 is deleted, also the capsid protein counts, (b) Promoter turns off; try a promoter that is tissue specific. (c) The episomal DNA lasts longer in cells that have low turnover (hepatocytes longer). (d) HD (gutless) ADv with a liver specicific promoter lasts for one year and (e) you can increase espression by blunting the immune system some 3 days before injecting de AdV ( cyclosporin like). An editorial in Molecular Therapy refers to this issue. Best wishes
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