AIDSWEEKLY Plus; Monday, January 18, 1999
Daniel J. DeNoon, Senior Editor

A new treatment strategy would do just that. The strategy uses a novel technique to rig a cell's own self-destruction mechanism so that it is triggered by HIV protease - effectively making the cell a death trap for the virus.

And the strategy could also be adapted to kill other pathogens (e.g., hepatitis C virus, cytomegalovirus, and malaria parasites) as well as some malignant cancers.

"We report here a new 'Trojan horse' strategy to treat HIV infected cells that exploits the HIV protease to kill the infected cell while leaving uninfected cells unharmed," wrote Adita M. Vocero- Akbani, Steven F. Dowdy, and colleagues of the Washington University School of Medicine, St. Louis, Missouri.

Vocero-Akbani et al. reported their findings in the journal Nature Medicine ("Killing HIV-Infected Cells by Transduction with an HIV Protease-Activated Caspase-3 Protein," Nature Med, 1999;5(1):29-33).

The researchers noted that current attacks on HIV protease employ small-molecule inhibitors of the crucial viral enzyme. However, inhibition of virus replication by these drugs actually extends the lifetime of infected cells, during which time drug-resistant viral variants may emerge.

They therefore sought an alternative treatment that would eliminate or reduce virus production. To achieve this they took advantage of the cellular enzyme caspase-3 (Casp3). Casp3 activation of begins a chain of events that lead to programmed cell death or apoptosis. The enzyme normally waits quietly within cells in a proenzyme or zymogen form, awaiting conversion to its active, lethal state by apoptotic mechanisms. This conversion involves removal of the zymogen's N-terminal Pro domain.

By replacing the Pro domain with a cleavage site that reacts only with HIV protease, Vocero-Akbani et al. created a modified Casp3 protein (dubbed TAT-Casp3) that initiates apoptosis upon interaction with the HIV protease enzyme.

A major problem with this approach is getting TAT-Casp3 into target cells. But a recent breakthrough by the Washington University research team may have solved this problem (Nagahara, H. et al., Nature Med, 1998;4:1449-52).

"Bacterially produced, misfolded fusion proteins containing an in- frame N-terminal protein transduction domain from HIV Tat are capable of transducing, in a rapid and concentration dependent manner, about 100 percent of all target cell types, including peripheral blood lymphocytes, all cells present in whole blood, diploid fibroblasts, fibrosarcoma cells, hepatocellular carcinoma cells, and leukemic T cells," they wrote.

In vitro experiments showed that the anti-HIV TAT-Casp3 protein could indeed transfect nearly all T cells in the culture without harm to the cells. But when TAT-Casp3 was added to cell cultures infected with HIV there was "substantial loss of HIV positive cells from the culture." Pretreatment of HIV infected cells with the protease inhibitor ritonavir - which inactivates the viral protein - protected the cells from TAT-Casp3-induced apoptosis.

Whether the Vocero-Akbani transduction technique will work in vivo remains to be seen. But the researchers are already working on ways to improve and/or broaden the use of their strategy:

• TAT-Casp3 is being modified to resist binding by cellular apoptosis-inhibiting enzymes.

• Instead of the Casp3 enzyme, the researchers are exploring use of an enzyme that makes infected cells susceptible to the otherwise noncytotoxic drug ganciclovir.

• Other apoptosis-inducing zymogens modified for activation by pathogen proteases may be added.

• Substitution of HIV cleavage sites used as the modified Casp3 trigger would permit treatment of HIV variants or resistance mutations.

"In addition to HIV, infectious pathogens such as hepatitis C virus, cytomegalovirus, and malaria are dependent on pathogen-encoded proteases for their life cycles," Vocero-Akbani noted. "Thus, the generation of modified TAT-Casp3 proteins containing these pathogen- specific protease cleavage sites may prove helpful in combating other infectious diseases and human malignancies that have either upregulated cellular proteases or express specific cellular proteases."

This work was supported by the NIH and the Howard Hughes Medical Institute.

The corresponding author for this study is Steven F. Dowdy, Howard Hughes Medical Institute, Washington University School of Medicine, St. Louis, Missouri 63110. Email: <dowdy@pathology.wustl.edu>.

990118
AW990107

Copyright © 1999 - Charles Henderson, Publisher. All rights Reserved. Permission to reproduce granted to AEGIS by Charles W. Henderson. Authorization to reproduce for personal use granted granted by C. W. Henderson, Publisher, provided that the fee of US$4.50 per copy, per page is paid directly to the Copyright Clearance Center, 27 Congress Street, Salem, Massachusetts 01970, USA. Published by Charles Henderson, Publisher. Editorial & Publishing Office: P.O. Box 5528, Atlanta, GA 30307-0528 / Telephone: (800) 633-4931; Subscription Office: P.O. Box 830409, Birmingham, AL 35283-0409 / FAX: (205) 995-1588 http://www.newsrx.net

Copyright ©1990, 2000. AEGiS & the Sisters of Saint Elizabeth of Hungary. All materials appearing on AEGiS are protected by copyright as a collective work or compilation under U.S. copyright and other laws and are the property of ÆGIS, or the party credited as the provider of the content.