The cellular and humoral immune responses to adenovirus (Ad) remain a major barrier to Ad-mediated gene therapy. We recently reported that mice deficient in tumor necrosis factor alpha (TNF-α) or Fas (APO-1, CD95) have prolonged expression of an Ad transgene expressing a foreign protein in the liver. To determine whether blockade of TNF-α or Fas would have the same effect in normal mice, we created transgenes that expressed soluble murine CD8 or CD8 fused to the extracellular regions of TNF receptor 1 (TNFR) or Fas and inserted into the left-end region of first-generation (E1/E3−) Ad vectors. Consistent with the results observed in TNF-deficient mice, expression of the TNFR-CD8 fusion protein was prolonged in vivo compared to that of control proteins. Not only did expression of TNFR-CD8 persist in the liver and the lung, but when coadministered with another first-generation vector, the protein provided “transprotection” for the companion vector and transgene. In addition, TNFR-CD8 attenuated the humoral immune response to the Ad. Together, these findings demonstrate that blockade of TNF-α is likely to be useful in extending the expression of an Ad-encoded transgene in a gene therapy application.