From Soup to Nuts - Drug Discovery and Development

HIV drug developers try to include every possible novel target and MOA on the menu.

click to enlarge Panacos’ novel maturation inhibitor, bevirimat, acts by inhibiting cleavage of the viral protein Capsid from its linker protein SP1. This results in immature, noninfectious virions, which are rapidly cleared from the body. (Source: Panacos Pharmaceuticals; Illustration: Tami J. Tolpa)

Human immunodeficiency virus (HIV) has been a challenge to drug developers since it was first discovered over 25 years ago. And although successful drugs have entered the marketplace since then, HIV remains a challenge. HIV’s constant exploration of its own genome in an attempt to improve its own biological fitness has lead to resistance of conventional HIV drugs and has been the major obstacle in the eradication of this virus. Thus, drugs with new mechanisms of action (MOAs) and with new targets—both in HIV itself as well as in human cells—are necessary to win the battle against HIV drug resistance. Luckily, there are drug researchers who are willing to fight. This article describes some of the latest HIV drug developments.

Mutating HIV
“Every time the virus mutates to escape the selective pressures of an HIV drug or the host immune system, its reproductive capacity falls off,” says Stephen Becker, MD, vice president of clinical development and chief medical officer at Koronis Pharmaceuticals, Inc., Redmond, Wash. According to Becker, it is because of this reduction in fitness with increasing number of mutations that caused Nobel Laureate Manfred Eigen to propose that if researchers could mutate this virus population, they could potentially drive HIV to extinction. It will not take a lot to bring HIV to extinction. “If you actually look at the [HIV] viral load, 97% of the virus is already dead. Actually, that observation is very important because it suggests that HIV—and this is true for HepB and HepC as well—is pretty close to an error catastrophe,” says Becker. And it was with that evidence that the founders of Koronis Pharmaceuticals attempted and eventually succeeded in developing KP-1461, a drug that selectively induces mutations in HIV.

Conventional HIV drugs are capable of reversibly suppressing viral replication. But because they are reversible inhibitors, conventional Highly-Active Anti-Retroviral Therapy (HAART) regimens must be taken for life. According to Becker, no other approved HIV drug works by the same MOA as KP-1461. “Unlike those drugs, KP-1461 irreversibly extinguished HIV in vitro,” says Becker, who adds that after the drug was removed, the virus did not replicate.

KP-1461 is a deoxycytidine analogue containing a modified base, and it competes with dCTP for incorporation. Tautomerization of this modified base between cytidine and thymidine brings about base-pairing errors. Moreover, HIV cannot repair these errors, which may bring HIV closer to extinction. Currently, KP-1461 is in Phase 2a, where dosing in humans is up to four months. Koronis is also conducting animal toxicology studies to investigate the possibility of longer-term dosing in humans. As per FDA guidance, the patient population used for this clinical trial has advanced HIV disease and few, if any, available treatment options. At the start of KP-1461 monotherapy, patients were taken off their HAART regimens.

Blocked entry

click to enlarge The first event in HIV-1 infection is the association of the viral envelope glycoprotein gp120 with the cell surface receptor CD4. This event facilitates binding to a second receptor, the membrane chemokine CCR5 or CXCR4, which triggers the sequence of events that lead to the fusion of viral and cell membranes. A major research effort is aimed at identifying and optimizing gp120/CD4 inhibitors. Two different classes of inhibitors have been identified: competitive inhibitors that block the association of gp120 to CD4, and allosteric inhibitors that block the signaling process without impeding gp120/CD4 binding. (Source: Ernesto Freire, PhD)

Another researcher interested in HIV drug resistance is Ernesto Freire, PhD, Henry Walters Professor of Biology and Biophysics, Johns Hopkins University, Baltimore, Md. A major area of Freire’s research deals with HIV entry inhibitors. HIV gp120 is a viral envelope protein that interacts directly with CD4 molecules on the cell surface of T-cells (HIV’s host cell) and thereby allows for HIV to enter these cells. Essential viral proteins that interact with human proteins are attractive targets for drug development. According to Freire, there is a limit to the number of mutations HIV can make in its gp120 protein because the virus still must be able to interact with human proteins to infect the cell. Freire is developing entry inhibitors that inhibit this interaction between gp120 and CD4.

The gp120 project is at the pre-clinical stage, is a collaborative effort between research groups at multiple universities, and is sponsored by the National Institutes of Health (NIH). For this project, Freire and collaborators are developing two different types of gp120 inhibitors. One type, which he calls competitive inhibitors, directly block the binding of gp120 to CD4; the other type, the so-called noncompetitive or allosteric inhibitors, block the gp120-induced signaling events that occur subsequent to binding and are necessary for HIV infection. Freire and collaborators hope to eventually license these drugs to big pharma so that they might be developed in the clinic.

Tackling Mitochondrial Toxicity
One major problem with long-term use of Highly-Active Anti-Retroviral Therapy (HAART) therapy for HIV is drug-induced toxicity. Varsha Desai, PhD, research biologist, Center for Functional Genomics, Division of Systems Toxicology, FDA-National Center for Toxicological Research (NCTR), Jefferson, Ark., is one researcher studying the underlying causes of this problem. Desai, who is speaking for herself, not on behalf of the FDA, says that “this work is being performed in collaboration with the National Toxicology Program and the principal investigator is Dr. Julian Leakey who works at NCTR.” The aim of this research is to determine the mechanism of toxicity and carcinogenicity of perinatal exposure to the HIV drugs AZT and 3TC.

Pregnant mice were treated with AZT alone or in combination with 3TC from gestational day 12 to 18. And then, they continued the same treatment in pups from postnatal day 1 through 28. These experiments are designed to model the FDA’s recommendation of AZT in HIV-positive pregnant women to reduce the transmission of HIV to the fetus, in which this treatment is continued in new born babies for six weeks postnatal.

Although these drugs are very effective in suppressing HIV infection in these pregnant women (as well as in other HIV-infected individuals), long-term use is associated with severe side effects. An underlying cause of the drugs’ side effect profile is believed to be mitochondrial dysfunction. “Some investigators believe that mitochondrial toxicity in Nucleoside Reverse Transcriptase Inhibitor (NRTI)-treated patients is due to reduced mtDNA copy number. But certain studies have shown that this is not the only type of mitochondrial dysfunction caused by NRTIs; they also affect the electron transport chain protein complexes as well as other mitochondrial targets,” says Desai, whose research is aimed at determining the exact mechanism of NRTI-induced mitochondrial dysfunction.

To study this mitotoxicity, Desai and her colleagues have recently developed a new genomic tool, a mitochondrial-specific oligonucleotide microarray they call the MitoChip. This array, developed for the mouse model, has 542 genes—both nuclear and mitochondrial—and can determine interaction between pathways that occur within mitochondria and involve both sets of genes. The hope is that by understanding the underpinnings of NRTI-induced mitotoxicity, less toxic drugs can be developed.

Inhibiting maturation
For their HIV drug development strategy, Panacos Pharmaceuticals, Gaithersburg, Md., is focusing on HIV maturation, the target for their lead compound bevirimat, a first-in-class HIV maturation inhibitor that is currently in Phase 2 clinical trials, with Phase 3 trials scheduled to begin in late 2008. Maturation, one of the last steps in the HIV life cycle, involves the processing HIV gag protein, a polyprotein that must be cleaved by HIV protease to form separate proteins; this step must occur during maturation. “There are drugs on the market now that inhibit the protease enzyme by binding to the enzyme, but what bevirimat does is distinct from that,” says Graham P. Allaway, PhD, chief operating officer at Panacos. Bevirimat works by binding to the gag substrate itself, and by doing so, prevents the release of a core protein of the virus called capsid, a part of the gag polyprotein that must be released in order to function. Bevirimat prevents the release of capsid from gag and thereby inhibits HIV maturation and the subsequent spread of infection throughout the body.

Other salient features of bevirimat include oral delivery and long half-life, a strong safety profile, and lack of drug interactions. “We have treated nearly 500 patients and healthy volunteers for the most part up to 14 days, with a handful of patients who have gone out to 24 weeks. And there really is no signature adverse event associated with bevirimat treatment. Its safety profile is relatively comparable to placebo,” says Scott McCallister, MD, chief medical officer at Panacos.

Multi-target mechanisms
Another major trend in HIV drug development is to find compounds that inhibit multiple targets. In particular, a novel target class, cyclin-dependent kinases (CDKs), holds promise for circumventing drug resistance. CDKs regulate the cell cycle; blocking them induces HIV-infected cells to commit suicide but leaves uninfected cells unharmed. “We can avoid inducing HIV resistance because CDK inhibitors do not target components of the HIV virus but kinases of the host cell on which HIV has hitched a ride. Targeting virus-carrying cells and not the virus itself may be a novel strategy of combating viral diseases,” says Spiro Rombotis, president and chief executive officer, Cyclacel Pharmaceuticals, Inc., Berkeley Heights, N.J. Investigators at George Washington University, Washington, D.C., showed that seliciclib, a Cyclacel CDK inhibitor, when applied to HIV-infected cells could “induce a cell cycle-like apoptotic outcome in HIV-1-infected cells, but at the same time spare uninfected cells.” Seliciclib inhibits CDKs 2/A, 2/E, 7/H, and 9/T and is being explored in Phase 2 trials in nasopharyngeal cancer patients.

Other novel multi-target compounds are designed to inhibit more familiar targets in HIV. “We are just designing compounds that are dual inhibitors where one molecule can inhibit two different enzymes. And there is a lot of interest in this type of work lately, but no one has successfully designed a molecule that inhibits two different enzymes of the AIDS virus,” says Robert Vince, PhD, director, Center for Drug Design and professor of medicinal chemistry, University of Minnesota, Minneapolis, Minn. “We try to design the drug so that one half of it can fit into one enzyme while the other half can bind to another enzyme.” Vince has one series of dual-target compounds designed to inhibit HIV reverse transcriptase and HIV integrase, while another series is designed inhibit reverse transcriptase and HIV protease. “What we are trying to do at this point is really prove the concept without coming up with some toxic molecule or something that has disadvantages over using two drugs.”

In summary, the discovery of new HIV drug targets and the concomitant novel MOA has a history of boding well with HIV drug development. And as the aforementioned research efforts detail, the hope is that rich history will continue long into the future.

This article was published in Drug Discovery & Development magazine: Vol. 11, No. 4, April, 2008, pp. 32-34.