News and Publications

Feline Immunodeficiency Virus

J.H. Elder, A.P. de Parseval, U. Chatterji, Y.-C. Lin, D.L. Lerner, Z.Q. Beck, A.J. Olson, C.-H. Wong,* A. Wlodawer,** C.D. Stout, S. Prasad

* Department of Chemistry, TSRI
** Frederick Cancer Research and Development Center, Frederick, MD

Feline immunodeficiency virus (FIV) causes an AIDS-like syndrome in domestic cats. It is a lentivirus closely related to HIV, the causative agent of AIDS in humans. Thus, FIV infection in cats is a natural animal model for study of the lentivirus life cycle and for the development of antivirals that block viral infection and spread. The goal of our research is to define the molecular structure of FIV proteins, with emphasis on determining regulatory mechanisms that control the rate of production of FIV and dictate the ability of the virus to infect specific cellular targets. Studies include both defining the functions of virus-encoded proteins necessary for replication and determining host-cell proteins that influence viral expression and spread. We wish to use information gained from these studies to develop intervention strategies that will be effective against both feline and human lentiviruses.

A major area of emphasis continues to be elucidation of the structure and function of FIV protease. This enzyme is responsible for accurately processing both itself and other enzymes and structural proteins from long polyproteins translated from virus-encoded mRNAs. Protease is necessary for the production of infectious virus particles, so blocking its function effectively blocks viral spread. By developing antiprotease drugs effective against FIV and HIV type 1 (HIV-1), we hope to avoid the development of drug resistance, a major problem that has limited the usefulness of currently available medications.

As reported previously, we have developed a C-2 symmetrical inhibitor (TL-3) that not only inhibits FIV and HIV-1 proteases in vitro but also inhibits the replication of FIV, HIV-1, and simian immunodeficiency virus in tissue culture. The inhibitor is also effective against 3 different drug-resistant HIV isolates obtained from patients undergoing treatment with several antiprotease drugs. Animal studies are in progress to determine whether TL-3 will be effective against viral infection in vivo. Additionally, we are carrying out both tissue culture and in vivo analyses to determine if drug resistance develops against TL-3. We hope that the broad efficacy of this compound will make it difficult for either FIV or HIV-1 to circumvent the drug without losing protease function. In a 9-month period, we did not detect the development of any drug resistance to TL-3 in FIV in tissue culture.

We are also working to define transcriptional control mechanisms for FIV. In particular, we are examining the role of a small open reading frame, termed Orf2, found in the middle of the virus. Recent findings now support the notion that this gene segment encodes a transcriptional transactivator (Tat) that is required for efficient viral replication in certain cells, particularly primary T cells. Coexpression of the gene for Orf2 in combination with a reporter plasmid, whose transcription is controlled by elements of the FIV long terminal repeat, results in a marked enhancement of transcription driven by the long terminal repeat. Studies of deletion mutants that lack binding elements for specific transcription factors suggest that a binding site for activator protein-1 may be required for Orf2 function. Further analyses are in progress to map Orf2-responsive elements and to define the nature of the interaction. Interestingly, certain cell types complement an Orf2 defect, a finding that implies the presence of a cellular homolog. This information may aid in ultimately understanding the mechanism of action of Orf2.

Studies are continuing to elucidate the cellular receptors that facilitate FIV infection of host cells. Understanding receptor interactions may allow the development of ways to block viral infection. Good evidence indicates that certain FIVs use the chemokine receptor CXCR4 to aid in entry into certain cell types, as does a subset of HIV-1 isolates. However, we have not found usage by FIV of the CCR5 chemokine receptor used by most clinical HIV isolates. Whether other chemokine receptors are involved in FIV infection is the subject of ongoing investigations. Currently, we are using recombinant FIV envelope protein as a probe and as an affinity matrix to bind and isolate cell-surface molecules that interact with the FIV envelope. Specific proteins bind to the recombinant protein, and efforts are under way to identify these molecules.

Studies are continuing to molecularly characterize the enzyme deoxyuridine triphosphatase, which is encoded by the pol gene of FIV. We previously reported the crystal structure of this enzyme. Studies during the past year have led to the 3-dimensional definition of a C-terminal P-loop--like motif that is required for dUTP cleavage by the enzyme. This motif was disordered in previous crystal structures. Further characterization of deoxyuridine triphosphatase will increase our understanding of the catalytic mechanism of this unique pyrophosphatase and, we hope, will yield generalities helpful to the study of all pyrophosphatases. We are also continuing collaborative studies to define the mechanisms by which FIV induces lesions in the central nervous system (see the reports of Drs. Phillips, Henriksen, and Fox). We hope that all these studies will lead to a better understanding of the life cycle of FIV and that this information will direct the development of effective interventions.

PUBLICATIONS

de Parseval, A., Lerner, D.L., Borrow, P., Willett, B.J., Elder, J.H. Blocking of FIV infection by a monoclonal antibody to CD9 is via inhibition of virus release, rather than interference with receptor binding. J. Virol. 71:5742, 1997.

Elder, J.H., Dean, G.A., Hoover, E.A., Hoxie, J.A., Malim, M.H., Neil, J.C., North, T.W., Tompkins, M.B., Tompkins, W.A.F., Pedersen, N.C., Miller, R.H. Lessons from the cat: Use of feline immunodeficiency virus (FIV) as a means to develop intervention strategies effective against human immunodeficiency virus type 1 (HIV). AIDS Res. Hum. Retroviruses 14:797, 1998.

Jacobsen, S., Henriksen, S.J., Prospero-Garcia, O., Phillips, T.R., Elder, J.H., Bloom, F.E., Fox, H.S. Cortical neuronal cytoskeletal changes associated with FIV infection. J. Neurovirol. 3:283, 1997.

Laco, G.S, Fitzgerald, M.C., Morris, G.M., Olson, A.J., Kent, S.B.H., Elder, J.H. Molecular analysis of the feline immunodeficiency virus protease: Generation of a novel form of the protease by autoproteolysis and construction of cleavage resistant proteases. J. Virol. 71:5505, 1997.

Lee, T., Laco, G.S., Torbett, B.E., Fox, H.S., Lerner, D.L., Elder, J.H., Wong, C.H. Analysis of the S3 and S3´ subsite specificities of feline immunodeficiency virus (FIV) protease: Development of a broad-based protease inhibitor efficacious against FIV, SIV, and HIV in vitro and ex vivo. Proc. Natl. Acad. Sci. U.S.A. 95:939, 1998.