Gene Therapy

Inder Verma L. Naldini, U. Blomer, P. Gallay, D. Ory, R. Mulligan, F.H. Gage, I.M. Verma, D. Trono, "In Vivo Gene Delivery and Stable Transduction of Nondividing Cells by a Lentiviral Vector," Science, 272:263-7, 1996. (Cited in more than 185 papers since publication) Comments by Inder M. Verma, American Cancer Society professor of molecular biology at the Salk Institute for Biological Studies, La Jolla, Calif. In the world of gene therapy, there's a fine line between friend and foe. HIV attacks cells with unparalleled efficiency. But, with a little genomic tweaking, the deadly virus could become an excellent, safe gene courier that might one day effectively eliminate cancerous cells. The authors of this paper were the first to report the use of a retroviral vector system based on HIV. Unlike other retroviruses, HIV, which is classified as a lentivirus, can target both dividing and nondividing cells such as neurons and macrophages (P. Smaglik, The Scientist, 12[10]:1, May 11, 1998). Prior to the advent of the HIV lentivirus vector, scientists could only infect with retroviruses ex vivo--a procedure that's not possible with tissues such as brain and lung. "More importantly, the efficiency following transplantation was very poor," explains coauthor Inder M. Verma, the American Cancer Society professor of molecular biology at the Salk Institute for Biological Studies, La Jolla, Calif. CUTTING AWAY: Blackened areas indicate the portions of the HIV genome that have been removed in the most recently developed HIV lentivirus vectors. Subsequently developed adenoviral vectors, based on the viruses that cause the common cold, do have the ability to infect genes in vivo. However, infected cells are too often recognized as foreign and are destroyed by the immune system. Verma and his colleagues wanted a virus that could readily integrate into nondividing cells without provoking immunological consequences. "The argument was that if these HIV viruses can infect nondividing cells we can take that property, take advantage of it, and ... convert it into a vector," notes Verma. To date, Verma and his team have not found any evidence of adverse immunological reactions to the virus in mouse and rat models. He does acknowledge, however, that more work needs to be done, possibly using higher titers of the virus. In order to make HIV a more efficient vector, Verma and his colleagues drastically altered its genome. Embedded in the virus membrane, the gp120 glycoprotein seeks out only cells that have CD4 receptors. To construct a vector that could infect cells other than CD4 cells, the Verma team removed the envelope (env) gene that regulates CD4 binding and membrane fusion and replaced it with one from another virus. "We ... made the virus far more useful by giving it a broad host range," explains Verma. In studies since this landmark paper, steps have been taken to deactivate the virus. Two teams, one led by Verma and Fred H. Gage, a professor of genetics at the Salk institute (T. Kafri et al., Nature Genetics, 17:314-17, 1997), and another led by Luigi Naldini, a scientist at Cell Genesys in Foster City, Calif., and Didier Trono, a professor of microbiology and genetics at the University of Geneva Medical School, (R. Zufferey et al., Nature Biotechnology, 15:871-5, 1997), have manufactured HIV vectors in which nearly all of the accessory proteins have been deleted. Only Rev, which regulates structural gene expression, was left intactm (See figure). In a more recent study still in press, the labs of Verma and Gage disabled long terminal repeats--elements controlling gene expression--on both ends of the genome. Verma plans to produce a third generation of vectors that will contain only a gag gene from the HIV, the reverse transcriptase gene from HIV (pol), and Rev--all of which together constitute about 25 percent of the total genome. "I can't tell you if every cell type in the body will be infected," cautions Verma. "For example, it is possible that lung ... may require all the accessory proteins for maximum infection. ... But in the tissues we have tested in the brain, in the liver, in the muscle, [and] retina, it doesn't seem to make a difference." Verma and his team now plan to use hemophilia as a model system in mice and dogs. They expect that the lentivirus will efficiently deliver the blood-clotting factors missing in hemophiliacs. "It's a very nice model system," Verma says of his current project, "and if it works, then I think the human will be the next choice."

Gene Therapy

The Scientist Staff

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