Integrase: Maybe this time?

The first scientific meeting on HIV integrase was held in the United States in 1995. By the time agents that safely and efficiently inhibit reverse transcription and protein formation were developed, there was talk in specialist publications and on websites about integrase as the third target, it being hailed as a promising new kind of antiretroviral. But it was not so. Years went by without finding a candidate that would yield positive results in humans. Recent data on new molecules are restoring some hope in what already seemed like an unattainable goal. And this time it may be achieved. As knowledge about the interactions between HIV and cellular mechanisms increases, over ten pharmaceutical companies are showing an increasing interest in developing an integrase inhibitor. And some candidate drugs are already being tested in humans.

The life cycle of HIV
Since they cannot do it on their own, viruses need to replicate in fresh cells in order to reproduce themselves. Like reverse transcriptase and protease, integrase is a viral enzyme (or protein) that plays a specific role in the complex HIV replication process in the infected cell. This process is known as the life cycle of HIV (see graph).

The process by which the virus infects a cell involves different stages. The first of these is when HIV gains entry into the cell, and it is this process that is prevented by entry inhibitors (for example, experimental co-receptor inhibitors or fusion inhibitors such as T-20, already on the market). Subsequently, the virus releases into the cell cytoplasm its genetic material and the enzymes that will be used in replication. The genetic material of HIV is in the form of RNA (ribonucleic acid), which must then be converted into DNA (deoxyribonucleic acid) in order to be able to replicate.

RNA is comprised of a single strand of genetic material while DNA is a double strand sequence. At this point, reverse transcriptase intervenes and converts the single RNA strand into a double stranded DNA sequence. There are currently two kinds of drugs that inhibit a successful completion of this stage: nucleoside analogue reverse transcriptase inhibitors (abacavir, AZT, ddI, ddC, d4T, FTC, 3TC; plus the nucleotide analogue reverse transcriptase inhibitor tenofovir) and non-nucleoside analogue reverse transcriptase inhibitors (nevirapine and efavirenz).

What is integrase?
Integrase is originally found in the HIV cell along with the other two enzymes that are fundamental for HIV replication. Following virus-cell fusion, it enters the cell along with the rest of the genetic material of the virus. The new viral DNA formed after reverse transcription is attached to the DNA of a human cell. This sets in motion a reaction that alters cell programming and leads to the production of HIV.

The double stranded sequence is then divided into two parts to form new RNA from which genetic material is drawn to form new particles that will create new versions of the virus. The different parts of the new virus are formed by these divided blocks of proteins. Protease is the enzyme that divides these blocks into chunks. This is the point at which protease inhibitors work (indinavir, ritonavir, saquinavir, nelfinavir, fosamprenavir, lopinavir, atazanavir, tipranavir).

As far as integrase is concerned, it can be regarded as having a brief, although highly complex effect. Integration occurs in various steps that can be grouped into two broad stages. Firstly, the viral DNA is prepared for integration. The integrase is capable of recognising the new DNA by the molecules at its ends, and exerts catalytic activity after binding to it. The first step is the removal of the GT dinucleotide molecules from each end.

The result of this process is transported to the cell nucleus where it is broken down and gives rise to a second reaction. DNA strand transfer takes place at various moments: the new ends of the viral DNA are then joined to the host DNA at two specific sites while two additional nucleotides at the end of the viral DNA are removed, thus allowing it to become fully integrated or fixed to the cell’s DNA.

An old target
The first integrase inhibiting candidate drugs acted during the first step of the integration process, and the initial findings were presented in the mid 1990s. Merck (MSD) pioneered research into therapeutic strategies and offered the first in vitro and in vivo efficacy data. It became clear, however, that the brief action of integrase in the HIV replication process made it a difficult target to block. Nevertheless, research continued.

In 2000 an MSD team led by Hazuda published a study on small molecules that could act during the second stage of integration, while transferring and fixing viral DNA to host DNA. These molecules are known as DiKetoAcids (DKA). The first candidates of this kind were Merck’s L-708,906 and L-731,988, followed by others patented by the U.S. National Institutes of Health: Shionogi’s S-1360 and a compound by BMS. However, success remained elusive. The development of Shionogi and GSK’s S-1360 was halted in 2003 while animal testing was being conducted.

Depending on their structure and activity, the integrase inhibitors studied or currently being studied can be grouped into three kinds. The ones described here belong to the class of benzopyruvic acid derivatives. Other derivatives are the 8-hydroxyquinolines, with MSD, Shionogi and Japan Tobacco candidates, and polyphenols, a type of molecule found in nature in certain plants such as sage, endives and lettuce, and of which synthetic versions are currently being studied by some companies.

When is it expected?
MSD, with over thirty registered molecules, is the company with most registered patents, followed by Shionogi with around ten and BMS, GSK and Gilead Sciences, which all have smaller numbers. As usual, some universities in the U.S. are also playing a role in this field by having patented some of these compounds.

The large number of registered patents, particularly in recent years, has already produced a series of candidate drugs for clinical trials. The results of such large research efforts are not in doubt, but will an integrase inhibitor ever become a reality? Only time will tell.

MSD’s MK-0518 recently completed Phase II and began Phase III (the last step before approval). GS-9137, a molecule developed by Japan Tobacco and later acquired by Gilead Sciences is now in Phase II and undergoing optimal dosage research. If these studies are successful the first integrase inhibitor may be available before the end of the decade.

Since they act at a different stage of the HIV replication cycle, and provided research is completed and they are eventually made available, these drugs could prove useful for people who are resistant to current ARVs. A negative aspect, however, as occurs with many new medicines which are launched in record time, is that toxicity in the long term will not be known until thousands of people worldwide are on a prescription. This aspect is becoming all too familiar in the history of HIV treatments, and has sparked debate on the periods of research required for new compounds.This issue, despite being of vital importance for people who have no other alternatives, can also pose unnecessary risks for individuals who already have therapy options that have proved to be safe and efficacious.

Xavier Franquet

References
Cotelle Ph. Patented HIV-1 Integrase Inhibitors (1998-2005). Recent Patents on Anti-Infective Drug Discovery, 2006, Vol 1, No 1.

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Hazuda DJ, Felock P, Witmer M, et al. Inhibitors of strand transfer that prevent integration and inhibit HIV-1 replication in cells. Science 2000; 287: 646-50

EATN - European AIDS Treatment News, Volume 15, I – Spring 2006

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