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Although first generation recombinant adenoviruses, deleted of sequences spanning E1a and E1b, have been useful for in vivo applications of gene therapy, expression of the recombinant gene has been transient and often associated with the development of inflammation. We show that with first generation adenovirus-mediated gene transfer to the mouse lung, viral proteins are expressed leading to destructive cellular immune responses and repopulation of the lung with nontransgene containing cells. Second generation E1 deleted viruses further crippled by a temperature sensitive mutation in the E2a gene were associated with substantially longer recombinant gene expression and less inflammation. Stable expression of human CF transmembrane conductance regulator has been achieved in lungs of CF mice instilled with a second generation virus.

An important limitation that has emerged in the use of adenoviruses for gene therapy has been loss of recombinant gene expression that occurs concurrent with the development of pathology in the organ expressing the transgene. We have used liver-directed approaches to gene therapy in mice to study mechanisms that underlie the problems with transient expression and pathology that have characterized in vivo applications of first-generation recombinant adenoviruses (i.e., those deleted of E1a and E1b). Our data are consistent with the following hypothesis. Cells harboring the recombinant viral genome express the transgene as desired; however, low-level expression of viral genes also occurs. A virus-specific cellular immune response is stimulated that leads to destruction of the genetically modified hepatocytes, massive hepatitis, and repopulation of the liver with nontransgene-containing hepatocytes. These findings suggest approaches for improving recombinant adenoviruses that are based on further crippling the virus to limit expression of nondeleted viral genes.

Chickens injected intravenously (i.v.) with human adenovirus type 6 (Ad6) reveal a 2-17-fold increase in the number of plaque-forming cells producing antibody (Ab) against sheep red blood cells (SRBC) 2-6 days after virus infection. Further, polyclonal B-cell activation has been demonstrated by the quantitation of immunoglobulin-producing cells (IgPC) and cells producing immunoglobulin (Ig) of IgM isotype (IgPC mu) in the spleen of chicken inoculated with Ad6. Ad6 infection in chicken results in immunosuppression against SRBC when this unrelated antigen is given after virus infection. It seems that coincidence occurs between the B-cell mitogenic activation and the immunosuppression caused by Ad6, as the most pronounced change in both activities appears on the fourth day following virus infection. These findings suggest that the B-cell mitogenicity of the virus contributes to the impairment of the humoral immune response to SRBC.

Spleen cells from mice infected with human adenovirus type 6 (Ad6) showed defective interleukin 2 (IL2) production 3-5 days after the infection. The response of spleen cells to exogenous IL 2 was also deficient. The impaired capacity of concanavalin A--(Con A)-activated spleen cells from Ad6--infected mice to utilize IL 2 seemed to be related to the depressed capacity of the infected splenocytes to express IL 2 receptors. The immunologic dysfunction following infection with Ad6 may be a consequence of deficiencies in both the elaboration of and response to IL 2.

CFLP and BALB/c mice inoculated intraperitoneally with large doses of adenovirus type 6 (Ad6) showed a decreased humoral immune response to sheep red blood cells (SRBC) and circulating interferon was detected in their serum. The timing of infection was critical. Infection of mice 3--11 days before SRBC administration led to depression of the 19 S haemolytic plaque forming cell (HPFC) response in the spleen. When mice were given Ad6 and SRBC simultaneously on Ad6 14 days before or 1 day after SRBC, there was no decrease in the number of HPFC. The suppressive effect was dependent on the dose of virus and antigen. Heat and UV treatment completely abolished the immunosuppressive effect of the virus, suggesting that a great amount of infectious adenovirus is needed to induce immunosuppression in mice.

The effect of intraperitoneally (i.p.) inoculated human adenovirus type 6 (Ad6) was tested for humoral immune response against sheep red blood cells (SRBC) in normal and indomethacin-treated mice, with the aim to elucidate the mode of the virus action in immunosuppression. The results indicate that inhibition of prostaglandin synthesis slightly influences the immunosuppressive effect of the virus. It is very likely that also other mechanisms are involved in the immunosuppression observed.