BOSTON--Harvard Medical School researchers, working in collaboration with scientists at other institutions, have made a discovery that could help unlock one of the more perplexing mysteries surrounding the AIDS virus: What causes the immune system of AIDS patients to suddenly cave in after years of valiant struggle?
Immune system collapse--and progression from HIV-infection to full-blown AIDS--has been shown to coincide with the rise of a mutated, and apparently more virulent, strain of HIV in the bloodstream of patients. Yet it has been unclear how, exactly, the late-comers might be causing such calamitous destruction.
It now appears that later (or T-tropic) strains of virus may exploit a different population of immune cells than do earlier (or M-tropic) strains. The conclusion, reported in the March 4 Proceedings of the National Academy of Sciences, was reached by looking at which immune cells express coreceptors for the two strains of virus.
Although both M-tropic and T-tropic strains begin entering immune cells through the same front door, the CD4 receptor, researchers at HMS and other institutions have recently discovered that each relies on a different chemokine coreceptor to complete its entry. In a series of experiments with scientists at Cambridge-based LeukoSite Inc. and the University of Pennsylvania, the HMS researchers found that subsets of cultured human immune cells express these recently-discovered chemokine coreceptors in a largely reciprocal fashion: Mature Oactivated1 T cells--which are T cells that have been taught to spot invaders--express the M-tropic receptor, CCR5. Untutored or Onaive1 cells express the T-tropic receptor, CXCR4.
If this occurs in vivo, then the rise of T-tropic virus observed in AIDS patients could coincide with a point at which HIV is expanding its base to include naive as well as mature activated T cells. In fact, the researchers experiments suggest that even immature activated T cells may admit T-tropic viruses for a brief period.
When naive T cells were artificially activated, they expressed very high levels of the T-tropic receptor, CXCR4, for the first three to six days, at which point expression of the receptor dropped off. Expression of the M-tropic receptor, CCR5, followed an opposite pattern: downregulation over the first three to six days, followed by gradual increase over the next 12 days.
The findings could help explain a mystery that has recently bedeviled AIDS researchers. To make T cells more responsive to HIV in their experiments, researchers would often artificially stimulate T cells. To their surprise, the activated T cells became resistant to M-tropic infection. The enigma can now be explained as a consequence of downregulation of the M-tropic coreceptor immediately after activation.
"I think the impact this paper will have on the HIV field is going to be pretty big," says Conrad Bleul, HMS research fellow in pathology at the Center for Blood Research, and lead author of the study. "Obviously, if you design your experiments now, and you1re using a certain subtype of virus [M-tropic or T-tropic], it will be very important to be aware of what the coreceptors do under your experimental conditions," says Bleul.
More significantly, the new findings promise to shed light on that shadowy brink--when HIV infection is turning into full-blown AIDS. "Before this was all really phenomenology. Nobody really had any way of relating this to a basic mechanism--a molecule, a coreceptor. Now, we can frame all of the really big questions about what happens in the disease process in terms of what happens to specific molecules," says Timothy Springer, Latham Family Professor of Pathology at HMS and the Center for Blood Research.
Charles McKay, senior director of immunology at LeukoSite and a coauthor on the PNAS paper, is currently exploring the feasibility of designing molecules to render the coreceptors uninfectable by HIV. "Because the T-tropic coreceptor plays a fundamental role in the immune system, blocking it could do damage," he says. However, preliminary experiments suggest that blocking the M-tropic coreceptor, which plays a less critical role in the immune system, can reduce the amount of virus in the bloodstream.
The discovery--which marks the HMS researchers1 third contribution over the last year to the burgeoning field of chemokine receptors--is the culmination of a rather circuitous and serendipitous journey. In 1995, Springer and Bleul, who are immunologists, came across an unusually powerful chemokine. (Chemokines are chemical signals that call immune cells to an infection). Normally, chemokines attract about 5 percent of cells. Bleul found that the new chemokine, called SDF-1, was attracting 50 to 80 percent of the cells in his sample. „We'd never seen any activity like that before," Springer says. (The findings were reported in the September issue of Journal of Experimental Medicine.)
Bleul set out to find a receptor that would respond to SDF-1. He found a protein, called LESTR, that responded to SDF-1. Meanwhile, AIDS researchers Robert Gallo and Ed Berger were discovering that chemokine receptors could also serve as entry points for HIV. Springer came upon a paper by Berger describing one of the new HIV coreceptors, called fusin. "Tim called me on a Saturday night, at home, and said OI have something very, very interesting here?maybe you should come in and read this paper," says Bleul.
Not only did LESTR turn out to be identical to fusin (now called CXCR4), but the researchers, working with Joseph Sodroski, HMS associate professor of pathology, also found that blocking it with SDF-1 powerfully inhibited infection by HIV. Their findings were reported in the August 29 Nature.
"Obviously we were very interested in finding what subset of T cells express the CXCR4 receptor," says Bleul. The researchers --using antibodies to CXCR4 and to CCR5 created by Jim Hoxie of the University of Pennsylvania and McKay, respectively--stained both naive and activated T cells. "The two subsets were pretty much mutually exclusive," says Bleul.
In a surprising twist, the researchers found that, in addition to naive cells, newly activated T cells stained for CXCR4 for three to six days after activation.
"People who are immunocompromised often get many infections, and these go hand in hand with activating T cells," says Bleul. If newly activated T cells express CXCR4 in vivo, then they may be providing a window of opportunity for infection by T-tropic strains.
It is still not clear whether the rise of T-tropic strains is the cause or the consequence of the collapse of the immune system. It could be a little of both, according to the researchers. "What people think is that late in infection, if the immune system is not able to clear virus anymore, then the virus can basically do what it wants," says Bleul. "One idea is that maybe they1re mutating toward T-tropic virus that can use a broader spectrum of target cells. So it may be that they can target naive as well as memory (or activated) cells," he says.
Adds Springer: "And T cells may not be the whole story. The differential expression of these T-tropic coreceptors on other immune cell types--such as dendritic cells--could also be very important, providing an additional reservoir of infectable cells."
Though speculative, such a scenario intimates a new breed of explanation for the critical transition from HIV-infection to AIDS: "Now we really know enough about what kinds of cells the [HIV] coreceptors are expressed on, how activation of cells affects coreceptor expression and we know what kinds of viruses use these coreceptors," says Springer. "We're in a position to frame the question of what happens at that crucial point in the host-virus relationship when the virus starts switching over to another receptor," he says.
While therapies designed to block CXCR4 do not appear feasible given its critical role in normal immune system functioning, McKay and his colleagues at LeukoSite are looking at the effectiveness of blocking CCR5. They have designed antibodies to the receptor and, once they have produced enough, plan to test them to see if they reduce viral load in monkeys and humans.