By David N. Leff

If outlaws in Western movies could be cut off at the pass and captured before they held up the stagecoach - instead of being arrested long after their criminal enterprise - this would help the sheriff uphold law and order.

A similar scenario attaches to infection with the AIDS virus - HIV-1 - which is uneasily and expensively arrested by the multidrug HAART, or highly active antiretroviral therapy. What the world needs is a strategy to cut HIV off at the pass - that is, forestall infection rather than holding it to a standoff.

What sounds like just such a too-good-to-be-true approach is described in the January 11, 2001, issue of Science Express, an online advance publication of papers in today's issue of Science, dated Jan. 12, 2001. Its modest title: "Protein design of an HIV-1 entry inhibitor." The article's senior author is structural biologist Peter Kim at the Whitehead Institute of Biomedical Research in Cambridge, Mass. Its principal author is biophysicist Michael Root, a postdoctoral fellow in Kim's lab.

He and his co-authors have designed a small but potent molecule they call "5-Helix," which binds to a region in the HIV envelope glycoprotein - gp41. As the virus gets set to contact and penetrate its target T cell, gp41 launches a harpoon-like projection, which hooks that target's membrane. This leaves the momentarily exposed gp41 stretched out end to end, from C-terminal to N-terminal - and vulnerable to counterattack by drug therapy. Then those two ends snap together, pulling the viral envelope and the T-cell membrane into contact. When their membranes fuse, HIV can enter and infect the cell. (See BioWorld Today, Oct. 4, 1999, p. 1.)

The 5-Helix molecule that Kim and his co-authors fashioned closely mimicked the structure that gp41 assumes during those climactic moments of fusion. That configuration consists of six twisted, supercoiled amino acid sequences, which fold together to resemble three interconnected hairpins. 5-Helix comprises five of those six alpha-helices, leaving an unrequited sixth that gp41 needs to fill in order to accomplish its T-cell break-and-enter mission.

"After Kim solved the gp41 X-ray structure," Root recounted, "which showed that gp41 formed a trimer of hairpins, of which the core was a bundle of six alpha-helices, our team investigated C-peptides in the outer core as effective inhibitors of HIV entry. C-peptides are about 34 to 40 amino acids long. They're derived from the C-terminal region of gp41's extracellular domain. Some C-peptides are very potent inhibitors of HIV entry, which work by binding to N-terminal regions of that domain, and thus preventing proper formation of the hairpins."

5-Helix Padlocks Access To HIV's T-Cell Target

"5-Helix was a challenging molecule to make," Root went on. "It's made up of N-peptides and C-peptides that are joined in a specific pattern. When you have a folded 5-Helix, this molecule will bind C-peptides. An unfolded 5-Helix contains two of these C-peptides. We subcloned this artificial protein in E. coli bacteria. I constructed and expressed the gene using genetic segments from what corresponded to the N-peptide and C-peptide regions of gp41.

"We isolated and synthesized this peptide," he recounted, "and showed that it displayed nanomolar inhibition against gp41-mediated membrane fusion. That result validated the N-terminal region of the gp41 extracellular domain as a viable target for designing drugs to inhibit HIV entry."

One potent C-peptide inhibitor called T-20 recently entered Phase III clinical trials in 1,000 HIV-infected patients. It's being-conducted by Trimeris Inc., of Durham, N.C., jointly with Hoffmann-La Roche Inc., of Nutley, N.J. (See BioWorld Today, Oct. 4, 2000, p. 1.)

"T20 has shown promise in Phase II clinical trials," Kim observed, "but it has to be injected into patients in large quantities." He made the point that drugs based on 5-Helix, which targets a different part of the HIV coat protein than T20, also would need to be injected, but could be self-administered much the same way as insulin, for example.

Kim added, "The 5-Helix protein could also serve as a basis for prophylactics that would be injected in hospital settings immediately after inadvertent needle pricks, to prevent HIV from infecting cells."

His paper in Science Express allows, "Three potential obstacles to using 5-Helix as a therapeutic include proteolysis, renal filtration and immune clearance." All of these are potentially correctable, as Kim's press statement suggested: "5-Helix is very stable, so it is less likely to be degraded by the body's enzymes. It can be made larger to avoid elimination by the kidneys, and modified to escape the body's immune response."

That response, he intimated, can be exploited to develop an effective AIDS vaccine. "The 5-Helix results suggest a new strategy for generating antibodies against HIV, which may be useful in vaccine development."

On a broader horizon, Kim sees the new entry inhibitor as particularly promising to confront the AIDS pandemic in Africa, and afflicted populations on other continents.

"The American public may have become complacent about the HIV epidemic," Kim observed, "largely due to the success of the triple-cocktail drug therapies. However, worldwide, HIV continues to be a public health menace, infecting more than 33 million people, many of whom in poorer countries can afford no treatment options. Even more alarming, the wily virus is mutating to form variants, which escape treatment by the existing drugs, making it important to continue to find new targets and therapies for stopping HIV."

Next Pre-Human Step: Monkeys

Moreover, Kim continued, "The 5-Helix approach may have broader applications to a wide range of human viruses that use a similar fusion membrane strategy to enter cells. These include the influenza and Ebola viruses and HRSV - human respiratory syncytial virus - a leading cause of infant mortality in developed countries.

"The next step," he said, "is to find out whether or not 5-Helix has antiviral activity in an animal model of HIV. If it does work in animals, then the difficult and arduous process of developing 5-Helix for humans could begin to take place. We may only be a few steps away," he concluded, "from seeing whether 5-Helix works in monkeys."