Engineered Virus Targets HIV-Infected Cells: Interview with Garry Nolan, Ph.D.
Two research teams, one at Yale and the other at the Federal Research Center for virus Diseases of Animals in Tübingen, Germany, have produced viruses which can enter and, in certain cases destroy, only HIV-infected cells. Although human use of such viruses will need more research and extensive ethical review, the same mechanism could be used almost immediately to target liposomes, which are already in widespread use for drug delivery -- potentially improving existing AIDS drugs by delivering them directly into infected cells, not to the body as a whole. The new research was published September 5 in the journal Cell; articles appeared in The New York Times and other newspapers on September 6.How was a virus engineered to enter only HIV-infected cells? The targeting works by exactly the same mechanism that HIV itself uses to enter human CD4 cells -- only in reverse. HIV enters human cells when a viral protein, gp120, which is on the surface of the HIV particle, interacts with the CD4 receptor (a protein which is normally on the surface of human T-cells, and certain other human immune-system cells); this interaction causes the virus to merge with the cell, infecting that cell. [It is now known that the CD4 receptor is not enough to allow HIV to enter a human cell; there must also be another protein called a co-receptor. Several co-receptors have been discovered, including CCR5, and CXCR4 (also called fusin); others may be found in the future. Differences in co-receptors cause some strains of HIV to infect T-cells, while others infect macrophages, or other human immune cells.
The team at Yale created an anti-HIV virus by starting with the vesicular stomatitus virus (VSV), which causes a disease in animals. This virus was genetically engineered to remove the protein which normally allows it to enter certain animal cells. Instead, the virus was given the genetic machinery to produce the human CD4 receptor, and also one of the human co-receptors which is used by HIV.
This virus cannot enter any normal cell. But if a T-cell or other human cell has been infected by HIV, the HIV protein gp120 will eventually appear on that cell's surface. Then the genetically engineered virus can fuse with and infect that cell, because of the interaction between gp120 (on the cell this time, not on the virus) with the human receptor proteins (which have been placed onto the engineered virus).
The other research team in Tübingen started with the rabies virus, and made it able to enter HIV-infected cells. This virus did not kill the cells efficiently, although it could be engineered to do so.
So far all these experiments have been done in laboratory cell cultures. Clearly there are major ethical questions to address before the virus could be tried in humans. But as Dr. Nolan points out, the same techniques could also be used to target liposomes, which are already widely studied as drug-delivery vehicles -- making existing drugs such as protease inhibitors more effective, by delivering them only to the infected cells.
On September 14 AIDS Treatment News spoke to Garry P. Nolan, Ph.D., Assistant Professor in the Department of Molecular Pharmacology at Stanford University, who wrote the review of this work which appeared with the research reports in Cell..
AIDS Treatment News: The virus produced by the Yale group was able not only to kill the infected cells it first entered, but to reproduce and kill more HIV-infected cells?
DR. Nolan: Yes it could replicate. Then it would seek out the other infected cells. Apparently this virus lived long enough in these experiments that even low levels of HIV production could be targeted. We know this because HIV replication appeared to start up again every couple of weeks or so in the cell culture, then be suppressed by the engineered virus. The HIV probably remained in rare cells in a latent form, periodically becoming active and then being suppressed again.
The concern now is that one must be very careful of any replicating system. The researchers are going ahead with monkey studies, I understand, not with people. One possible problem is that this kind of virus might be infectious not only to the person you provide it to, but also to any sexual partners that HIV-infected person might have contact with, who also have HIV. Anybody who is HIV-positive could become an outlet for this virus, which is why I believe we need research on how one could control the virus, and tone it down. The medical and ethical review will take some time.
What could be done now is to use liposomes, instead of viruses. You could put CD4 and CXCR4 onto the surface of liposomes, to target them only to HIV-infected cells. Rather than delivering a virus, this technique would deliver packets of drugs, using the receptors to target them much more effectively than conventional liposomes. This approach might greatly improve the effectiveness of drugs like protease inhibitors, while reducing their toxicity.
It is unlikely that the liposome technology would be very useful for drugs that attack the virus early in its life cycle. Reverse transcriptase inhibitors such as AZT, for example, must be in the cell before the virus is integrated into the human genome. At that time the cell will not have enough gp120 on its surface to make it a target for the engineered viruses.
ATN: Why did these studies use CXCR4 (fusin), instead of other co-receptors?
You are not just limited to CXCR4. You could have used one of the other HIV co-receptors as well. It might be possible to design liposomes that include all the co-receptors, to target them against all of the different kinds of cells that HIV infects. And one could do the same thing with viruses.
Avoiding Viral Resistance
The advantage of using receptors in this way, to target viruses or liposomes to particular cells, is that HIV does not have a way around it. If HIV mutates so that the engineered virus cannot bind with it, then that HIV could no longer enter human cells [because it needs the interaction between gp120 and the human receptors to do so]. This is one of the first times we can design something that the virus probably cannot escape.
HIV can mutate quickly against a drug which attacks it directly. But if you design a treatment which prevents the virus from using the cellular protein which it must use, then the virus may be blocked. This strategy has not received enough attention in HIV drug development. Existing drugs target the virus, rather than its interaction with what the virus takes advantage of in the body.
Next Steps
ATN: What has to happen first to develop such a liposome for treatment?
DR. Nolan: Companies need learn how to scale up the production of these proteins. CD4 will be relatively easy. But the co-receptor will probably pose some difficulty, because it is an integral membrane protein. Someone will have to learn how to purify these co-receptors, and then incorporate them into liposomes.
ATN: Wouldn't it be faster with the viruses, since one already exists in the laboratory, and can reproduce itself?
DR. Nolan: There are major issues. You do not want to scare everybody that you will be releasing new viruses. Certainly people are going to be thinking about the safety issues. What happens if you give a replication-competent pathogen to an immunocompromised patient? Somebody is going to have to volunteer. Don't be surprised if it does not go quite the way you want it to.
ATN: One science-fiction thought is that if this approach did work, it might be possible to treat the worldwide epidemic by treating one person. Of course that would be a long time from now.
DR. Nolan: People will test the virus idea with animals fairly soon.
But meanwhile, what can be done now is to look at the liposome technology for delivering drugs, and ask what drugs could be encapsulated. There is lots of technology already about liposomes, since they are being researched heavily for standard medical treatment. The trouble has always been that liposomes are not specific enough, and also that they do not have the ability to fuse with cells. But adding the CD4 and a co-receptor basically provides a self-loaded trigger. It pulls the trigger for the virus, when it binds next to the cell. Then the liposome will fuse with the cell and deliver the drug.
What would we want to deliver? The obvious approach is a drug to kill the infected cell. But it might also be possible to deliver something which specifically seeks out the virus in the DNA, and does not kill the cell but shuts the virus down. People are talking about what is called "triplex DNA"; it is a way of basically delivering a clamp to a target cell, to the cell's DNA. It clamps down on a piece of the viral DNA within the cell's genome, and never lets go, basically locking the DNA in an 'off' position. So the virus could still be there, but it would be permanently inactive.
Besides triplex DNA, some other technologies are being developed -- for example, for creating small molecules that will bind to specific DNA sequences. Using receptor proteins (to target either viruses or liposomes) may help solve the difficult problems of delivering these potential treatments into the appropriate cells.
