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Threshold, Schmeshold

Long-Awaited Research Paper Takes on the Viral Diversity Threshold, Cannibalistic CTLs and NSI-to-SI Switch

Shining Star of the New Guard

July 1996

AIDS research is only fifteen years old. A few years ago, the field was largely dominated by and comprised of scientists who had moved from other areas of inquiry to colonize a new metier for investigation. Like colonists before them, many had made the voyage into AIDS research to seek their fortune in the new pot of grant money available for this new disease, to re-start their professional lives after little success in their native specialities, or for the sheer adventure of exploring an uncharted scientific wilderness.

The first generation of AIDS researchers primarily trained in the new field, however, has now come of age. The leaders of this new generation are a precocious bunch. Most of them studied with the early pioneers who established the discipline, but have already far surpassed their elders. Some have a loose formal association, as many of them are part of the Pediatric AIDS Foundation's Ariel Project to Prevent Transmission of HIV from Mother to Infant. A few of the new wonderkids include David Ho of the Aaron Diamond AIDS Research Center, Steve Wolinsky of Northwestern University, George Shaw and Beatrice Hahn of the University of Alabama, Jim Mullins of the University of Washington and J. "Mike" McCune of the Gladstone Institute of Virology and Immunology at UCSF.

Steve Wolinsky's recent paper in the weekly journal Science (26 April 1996), "Adaptive Evolution of Human Immunodeficiency Virus Type 1 During the Natural Course of Infection," is the latest example of the work of this talented group. In this remarkable paper, Wolinsky and colleagues set out to elucidate the factors that lead to the variable rate of disease progression in HIV infection and in the process boldly demolish three existing theories which have tried to explain the phenomenon. Wolinsky's investigation is based on semiannual blood samples from six men from the Chicago site of the Multicenter AIDS Cohort Study (MACS).

The first theory of the correlates of HIV progression to be debunked by Wolinsky and his team is the notion that disease progression is due to increasing antigenic diversity which leads to the crossing of a threshold beyond which the immune system of an HIV-infected individual can no longer control the virus. In other words, this hypothesis claims that HIV's enormous capacity to mutate and present slightly different versions of itself to the human immune system becomes, at some fateful moment, greater than the ability of the immune system to effectively recognize and subdue all these viral variants. At first, the immune system goes toe to toe with the virus, sending one after another antigen-specific T-cell and its clones to battle their retroviral antigens until HIV dons a form for which the immune system has no T-cell with which to respond. HIV then flourishes unfettered rampaging like a band of Visigoths across the body.

Wolinsky deconstructs this idea by tracking the development of genetic diversity over time in the six men from the MACS--two individuals that progressed rapidly, two that progressed at a moderate pace and two that have remained stable. Wolinsky did this by sequencing DNA coding for a part of the viral envelope over the course of the study and charting the rate of accumulation of mutations that would result in amino acid changes (changes that could confer structural and/or antigenic changes to the virus). Contrary to what the existing theory predicted, Wolinsky showed that the slower progressors in his study had greater genetic diversity than those who progressed more rapidly. The genetic changes in his slower progressors were also more likely to result in amino acid changes pointing towards a selective pressure driving the virus towards mutations that changed its appearance to the immune system. With the genetic data from his subjects, Wolinsky, grouping together similar amino acid sequences, was also able to show that his slower progressors also had greater antigenic diversity than the rapid progressors.

Wolinsky and his research team then move on to jettison a long-standing fancy that a pathological immune response, particularly by cytotoxic T-lymphocytes (CTLs), may be responsible for pushing disease progression along. Looking at both antibody and cell-mediated responses, Wolinsky's team saw no association between antibody response and disease progression, but did show that a vigorous HIV specific CTL response was associated with a slower rate of disease progression. Clearly, CTLs are not chomping away on CD4+ T-cells in a cannibalistic frenzy: they are delaying progression rather than inciting it.

Finally, after clearing away two theories of AIDS pathogenesis in one paper, Wolinsky moves onto skewer a third. The most recent theory of AIDS progression embraces the hypothesis that a change in viral virulence is what's responsible for progression (and acceleration) toward symptomatic disease. This theory claims that as people with HIV progress to AIDS, the predominant viral population shifts from strains with a predilection for macrophages and monocytes to those that seek out T-cells. In addition, the monocyte tropic strains do not generally have the ability to induce the deadly clumping together of T-cells (known as syncytia), while the T-cell tropic variants do induce this phenomenon. Wolinsky found no evidence for a shift from non-syncytium-inducing, monocytotropic to syncytium-inducing, T-cell tropic strains in any of their subjects. In fact, none of the subjects studied harbored the dreaded T-cell clumping viral strains. In addition, viral strains from all subjects replicated equally well and destroyed T-cells at equivalent rates when introduced into SCID-hu mice.

So what sends the HIV+ person down that stony, stony path to disease progression in Wolinsky's study? As David Ho, another superstar of the new generation of AIDS researchers and a co-author on the Wolinksy paper has said: "It's the virus, stupid!" The rate of T-cell depletion correlated with the amount of viral RNA in the blood at around 9 months after primary infection in Wolinsky's subjects, as well as with the ratio of unspliced to multiply-spliced messenger RNAs at this time. The average viral burden over the course of the study also correlated with these two factors. In Wolinsky's world, the more virus you have at 9 months post-infection, the faster you will experience CD4+ T-cell loss. In addition, the larger the ratio of unspliced RNA (It is the job of unspliced RNA to code for HIV's structural proteins pol, gag and env) to multipy-spliced RNA (Multiply-spliced RNA code for HIV's regulatory proteins: rev, nef and tat.) you have at that earlier time, the worse you can expect your prognosis to be. Structural proteins (the building blocks of new viruses) are produced later in the viral life cycle than the regulatory proteins like tat, nef, and rev. If you have a lot of structural proteins being produced in comparison to regulatory proteins, it is likely that your T-cells are churning out new virions by the bucketful: not a good sign early in the course of infection, at least according to Wolinsky.

Wolinsky's universe is a Darwinian system where a powerful (and, ideally, adaptive) immune response exerts continual selective pressure and drives viral diversification. While Wolinsky debunks three theories and establishes several firm correlates of disease progression, he offers nothing at all as to his explanation for the mechanism by which the final immune collapse in AIDS occurs. In an accompanying commentary, Frank Miedema and Michel Klein of the Central Laboratory of the Netherlands Red Cross Blood Transfusion Service, propose a few possibilities. As David Ho has said, HIV may simply exhaust the lymphopoietic system by the enormous numbers of T-cells that it destroys and that get replaced each day. At some point, the capacity of the body to produce T-cells just burns out. Miedema and Klein hypothesize that the final decline in AIDS may occur through "the gradual perturbation of cellular immunity," as CD4+ cells necessary in the regulation of the immune response are picked off by HIV and as the remaining immune system cells become functionally impaired.

Wolinsky's lab and those of his colleagues are producing some of the finest scientific work ever on AIDS. The leaders of this new generation of researchers are men and women working at the peak of their talents. Hope for people with HIV is often solely conceived as a new drug coming down the pike. However, hope is also knowing that there are scientists out there that are slowing chipping away at HIV's secrets, uncovering the wily ways HIV infects us and destroys our immune systems, opening up new avenues in understanding which may help us to stop this epidemic in in tracks.