Being Alive Newsletter, Being Alive/Los Angeles - July 1993
Ben Cheng

WHAT IS HIV PROTEASE?

HIV protease is a protein comprised of two identical structures 99-amino acids long, that are symmetrical in C shape (think of a boomerang). It is a member of the aspartyl-protease enzyme family that also include renin, used in making cheese, and pepsin, used to slice and dice antibodies. The HIV protease enzyme is essential for viral infectivity and replication. In in vitro (test tube) experiments, the activity of reverse transcriptase was impaired when normal protease was absent. Mutation in substrate cleavage sites (the places where the model parts are supposed to break off the frame) led to abnormal viral development (think of square wheels on your model car).

HIV protease breaks the gag polyprotein product into several infectious proteins, p6, p7, p17 and p24. (Gag is the gene that makes the protein parts used to build the internal egg-shaped capsid that protects the viral RNA.) If the HIV protease is chemically blocked, the formation of these core proteins is disrupted and the virions that are assembled are malformed, immature, and non-infectious.

DEVELOPMENT OF HIV PROTEASE INHIBITORS

Pharmaceutical companies and biotechnology companies are researching methods to produce a potent oral drug to inhibit HIV protease. Most of the research has focused on two basic approaches: 1) screening compounds that are known blockers of aspartyl proteases, such as renin and pepsin inhibitors; and 2) taking a structural approach. That means that researchers examine the 3-D shape of the HIV protease and use molecular tools to design and synthesize possible inhibitors of this enzyme. The specific drug must be active against the virus, but must also have minimal interaction with the normal cellular processes.

Some of the obstacles that have been encountered in the development of the protease inhibitor are that some compounds have been shown to be potent inhibitors of protease in vitro, but they have no anti-HIV affects in cell cultures. Another obstacle has been the poor oral absorption, poor stability, and rapid metabolism (breakthrough) of peptide-based drugs which have shown potent inhibition of viral maturation and a reduction of infectious particles in cell culture.

The structure-based approach to design HIV protease inhibitors has allowed researchers to identify several strong C2 symmetric inhibitors (think of an overhead ceiling fan). These drugs may only bind to HIV protease, because their shape allows them to fit tightly into the enzyme and prevent it from working while this same shape keeps them from interfering with more oddly shaped human enzymes.

A number of protease inhibitors have entered or are about to enter clinical trials. Hoffman LaRoche's RO 81-8959 was the first compound to be tried in humans. The Phase I safety and pharmacokinetic study was conducted in England, France and Italy. The drug is peptide-based so is not very well absorbed (peptides break down very quickly and easily in humans), but is being developed as an oral agent. The oral bioavailability is only 4% of the intravenously administered version; however, bioavailability is increased 18 times when it is taken with food.

Yet another problem is that this drug requires 21 steps to manufacture. However, data presented from these three studies has shown that the drug is very well tolerated at the highest dose tested, 600 mg three times a day, and there was an increase in CD4 counts and a reduction in viral load. Additionally the Italian study showed that the combination of AZT and RO 81-8959 was well tolerated with a greater increase in CD4 counts that either drug alone.

Hoffman-LaRoche is conducting a larger clinical trial in the US using the same 600 mg three times a day dose in combination with either AZT or ddC. Clearly, higher doses of this compound also need to be studied.

Abbott's first generation protease inhibitor, A-77003, has also been tested in humans. This drug has a C2 symmetry design, but again, this is an all peptide drug with very poor bioavailability so it has to be administered intravenously. Due to the early development of toxic side effects in humans, this drug will no longer be developed. Abbott's second generation protease inhibitor, A-80987, will be entering clinical trials in the near future. This drug also has C2 symmetry structure but is has a bioavailability of 20%. It can therefore be administered orally. Abbott also has a third generation protease inhibitor in development, A-84538, which in vitro is about ninefold more potent than the first two generations and is also orally bioavailable.

POTENTIAL CLINICAL USES OF PROTEASE INHIBITORS

The rapid emergence of viral resistance to the RT inhibitors has limited their long-term usefulness. It is hoped the development of resistance will be much slower in protease inhibitors, because they have activity against both HIV-1 and HIV-2. However, a mutant of HIV-1 that is partially resistant in vitro has recently been observed.

One obvious use of protease inhibitors would be to combine them with RT inhibitors. Protease inhibitors effectively prevent the production of infectious virus in chronically infected cells, whereas RT inhibitors do not affect the viral production in this kind of cells. In addition, additive to synergistic anti HIV interactions were seen in the test tube, when a protease inhibitor was combined with AZT, ddC or alpha interferon.

Another potentially useful strategy would be to combine two different protease inhibitors a C2 symmetry based compound and a non-C2 symmetry based compound. In so doing, researchers would hope to stop protease by hitting it with two differently shaped enzymes. Ultimately, a protease inhibitor could be used in combination with compounds that attack the virus at different sites, such as tat inhibitor, a nucleoside analog RT inhibitor, and a non-nucleoside RT inhibitor. The convergent therapy model that is currently being advocated for RT may also work for protease, but this experimental work has not been done yet.

COMMENTARY

The overall effectiveness of protease inhibitors as a potential treatment for HIV is still unknown. However, given the limited usefulness of RT inhibitors, it is critical that this class of compounds be developed and tested rapidly in people at all stages of disease. Many researchers believe that protease inhibitors must be combined with other antiretroviral drugs, since they have demonstrated synergistic effects. Many researchers also believe that it may require testing three or four generations of protease inhibitors before a truly effective one is found.

While resistance to the protease inhibitors remains a critical question, many people feel that this class of drugs is currently the most promising approach for the treatment of HIV.

(Ben Cheng is an AIDS treatment activist now with Project Inform in San Francisco, 415.558.8669.)
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