Scientific American, 7/98, John W. Mellors, page 70
Viral-Load Tests Provide Valuable Answers

In the early 1990s tests that could accurately detect the amount of HIV in a patient's blood finally became available. These viral-load assays have since revolutionized understanding of HIV's behavior and have helped define new principles of therapy. The assays directly measure viral RNA per milliliter of blood plasma; each HIV particle contains two strands of RNA, so the level of actual virus is half the RNA count.
The tests enabled researchers to show that HIV never undergoes a period of slow growth. From the start, the collective viral population in a patient generates many billions of new HIV particles a day, resulting in destruction of millions of CD4 T lymphocytes-the immune cells most depleted in infected patients. The body tries to compensate for the loss by making new CD4 T cells, but the immune system remains under constant siege and eventually fails to keep up.
In 1996, as part of the federally supported Multicenter AIDS Cohort Study (MACS), my colleagues and I measured virus in stored plasma samples collected from about 1,600 untreated HIV-infected men and traced the fate of those patients. We found striking differences in prognosis, depending on the level of virus (graph). For example, 70 percent of the men whose viral load was greater than 30,000 copies per milliliter died within six years of the test, the average survival time being 4.4 years. In contrast, fewer than 1 percent of patients whose viral load was below 500 copies per milliliter died in six years, and the average survival time was more than 10 years. These results established that viral load critically influences the rate of disease progression. They also suggested that lowering viral levels as much as possible for as long as possible with therapy is essential to prolonging life.
Subsequent studies using viral-load tests have both confirmed this concept and changed how new therapies are evaluated. Until recently, investigators assessed potential treatments by comparing the incidence of AIDS or death in test subjects and a control group, often having to wait years for definitive results. After the viral-load assays became available, several large studies demonstrated that measures of viral load, often after just weeks of treatment, were valid indicators of whether a therapy could slow progression of HIV infection. For example, treatments that lowered the load by 75 to 90 percent within eight to 24 weeks reduced by 50 to 65 percent the likelihood of progressing to AIDS within a year. Viral-load measures have therefore replaced assessment of clinical outcome in therapeutic trials, and routine monitoring of viral levels has been incorporated into medical practice.
Recent studies have suggested a refinement of the admonition to keep viral levels low. Current treatment guidelines aim to maintain viral load below 500 copies per milliliter (the limit of detection for the tests usually used today). Failure to reach that level is associated with breakthrough of drug-resistant virus and loss of control over HIV replication. Yet trials applying more sensitive tests indicate that depressing viral load below 50 copies per milliliter offers better insurance against resistance. Moreover such a reduction is probably necessary to halt viral replication everywhere-in lymph nodes (where the replication rate is higher than in the blood) and also in other body compartments. I believe, therefore, that viral loads below 50 should become the new goal if the more sensitive tests (now used only in research) are made readily available. Not all physicians agree, however, in part because this target may be more difficult to reach, particularly for patients in whom initial therapy has failed.
A major new challenge for therapy is finding ways to eliminate HIV from infected resting CD4 T cells, which do not produce viral particles but harbor the genetic blueprints for doing so in the future. Current antiretroviral drugs cannot eliminate these HIV reservoirs. To develop such therapies and monitor their effects, investigators will undoubtedly require a new generation of viralload tests-ones that can accurately measure the virus hiding in resting cells. Fortunately efforts to create those urgently needed assays are under way.
Although the "field" results have not matched those of the clinical trials, a SO percent success rate is still 100 percent better than could be achieved just a few years ago. And that imperfect level of success apparently has been enough to slow disease progression in many cases. Since HAART was introduced, the HIV clinics at San Francisco General and at Johns Hopkins have witnessed a 50 to 80 percent decrease in hospitalizations for HIV related problems and a SO to 70 percent drop in the incidence of major AIDS-related opportunistic infections.
The less stellar results in the community are not surprising. Patients cared for in clinics are much more heterogeneous than are trial participants. Many begin therapy in a relatively late stage of HIV infection, when their viral burden may be too large to overcome with existing therapy and when their immune system may be beyond rescue. Most clinic patients, moreover have had prior exposure to antiretroviral drugs and so probably harbor virus resistant to one or more drugs in the regimen.
The Down Side of HAART
What is more, HAART has many drawbacks that can hinder adherence unless a patient is very committed and organized. Patients must swallow at least eight, and often 16 or more, anti-HIV pills a day (along with any other needed medicines, such as those meant to prevent specific opportunistic infections or to control pain). They must remember which ones have to be consumed with food, which on an empty stomach and which cannot be taken together or with other kinds of pills. Even the most efficient individuals can become confused or forget doses; a number of clinic patients are homeless, demented or dependent on illicit drugs or otherwise lacking in the supports that would help them adhere strictly to a medication schedule.
Patients missing the encouragement and support provided in clinical trials may also be more likely to skip doses or to give up on therapy when they encounter unpleasant side effects. The unwanted effects of antiretrovirals can range from rash, nausea, diarrhea and headache to anemia, neuropathy (painful or numb feet), hepatitis and possibly diabetes. Certain effects, if severe enough, can be dangerous or intolerable for anyone, but others are potentially bearable.
Whatever the reasons for missing doses, the consequences can be incomplete suppression of viral replication and the emergence of resistance. HAART, as some say, is unforgiving of nonadherence. Once resistance sets in, the patient cannot "start fresh" with the same plan; it will not work. Regrettably, incomplete adherence to treatment plans accounts for about half of treatment failures. No one knows the minimum number of missed pills needed to promote resistance.
We should note that in some patients who apparently follow therapy strictly, the virus fails to reach, or to stay at, undetectable levels. This virological failure may occur because the prescribed drug cocktail is not potent enough to halt viral replication fully in those particular individuals. Their bodies, for whatever reason, may break down the drugs too rapidly or may block their complete dissemination, resulting in too little medication reaching infected cells. Also, when viral-load measures are below 500 copies per milliliter, some people will truly have no viral replication occurring, but others will have some unknown degree of activity. This small amount of activity may be enough to allow resistance to arise.
Unsettled Issues
Experience with HAART in the past few years has been instructive, but it has yet to resolve a number of questions-among them, Should everyone be treated as soon as they are diagnosed?
Experts agree that the ideal time to institute therapy is during the acute stage, when patients have the best chance of preserving their immune defenses. Few patients are diagnosed at that stage, however At the other end of the spectrum, people who already have symptoms, or who have a CD4 T cell count below 200 per cubic millimeter, need treatment, too; they probably will not live long without intervention.
Authorities on HIV care also favor offering HAART to a third group: symptomless patients with a CD4 T cell count between 200 and 500 cells per cubic millimeter or a viral-load measure exceeding 10,000 to 20,000 RNA copies per milliliter Without treatment, patients in this third category have at least an 8 percent chance of progressing to an AIDS-defining opportunistic infection within three years and at least a 25 percent chance of reaching that unwelcome benchmark within six years.
The T cell and viral-load thresholds, however, are fairly arbitrary. Because patients in the third group feel perfectly well, certain of them may be quite reluctant to embark on a demanding treatment plan, with its side effects and constant reminder of the disease. Further, if resistance arises and treatment fails, the patients will be left with restricted options later on. Some will therefore choose to delay therapy until tests show signs of progression or until simplified regimens or new drugs with fewer side effects become available. Most patients with better numbers - CD4 T cell counts over S00 and viral burdens below 10,000 to 20,000 - will decide to wait as well.
For patients in whom HAART does not quash detectable viral replication, the issue becomes, What next? Unfortunately, no second-string, or salvage, strategy seems to succeed as frequently as does HAART given to patients who have never been treated with antiretroviral drugs. Physicians generally respond by helping patients who had difficulty following the initial therapy to overcome obstacles to adherence. They then switch the entire regimen to one composed of all new drugs, taking care to avoid agents that are likely to encounter resistance.
Experience also offers limited sage advice for what to do when salvage regimens do not halt detectable viral replication. In people whose immunity has collapsed, persisting with antiretroviral treatment might do little good. For others, continuation of the regimen may be worthwhile. As the clinics at San Francisco General and Johns Hopkins have found, virological failure (incomplete viral suppression) is not the same as clinical failure (the emergence of HIV related complications). And a few investigations have shown that even when viral levels are climbing, combination therapy can sometimes restrain HIV replication somewhat; a technically failing therapy, then, may nonetheless help maintain or elevate CD4 T cell levels and thus might buy patients valuable time.
Even. those who do very well on HAART from the beginning pose puzzles for investigators. Scientists remain unsure whether apparently successful therapy can fully reconstitute the immune system. So far only partial restoration seems to be common, with treatment raising CD4 T cell levels by an average of 100 to 200 cells per cubic millimeter. The mix of CD4 T cell subsets may remain abnormal as well-that is, the expanding population may not recognize as many pathogens or may be less effective at combating infections than was the original T cell population.
Can Reservoirs Be Eliminated? Looking ahead, everyone hopes that initially successful HAART regimens will keep working year after year. When HAART fully stops HIV replication, it seems to prevent the development of resistance and thus should work indefinitely. But whether fact will bear out theory remains to be seen.
Investigations into whether currently available drug combinations can cure HIV infection are less encouraging. In resting (nondividing) CD4 T cells, HIV can apparently survive in a latent state, as an integrated provirus that produces few or no viral particles. These resting cells can be prodded to churn out new particles if something causes them to become active again. When HAART was first introduced, some scientists expressed hope that resting T cells would die out quickly on their own, thereby eliminating the perpetual threat of a HIV resurgence. New findings, however, suggest that certain of the cells may endure, and pose a threat, for years.
Not surprisingly, investigators are considering ways to incite the immune system to eliminate these reservoirs. To destroy the cells, the immune system has to "see" them. Normally when a cell is infected by a virus, the cell displays pieces of the microbe, or antigens, on the cell surface; then T cells that bear receptors for the antigens orchestrate an attack. But cells carrying a quiescent provirus in their DNA do not display HIV antigens.
One strategy for overcoming this problem is to administer substances that would purposely activate the resting cells, inducing them to make HIV particles, display antigens and arouse a killing immune response. Treatment with HAART, meanwhile, could prevent the virus from colonizing other cells. The risk, obviously, is that the strategy would backfire, allowing the virus to gain the upper hand. And even if the plan worked, other protected havens for HIV, such as brain neurons, might continue acting as lifelong sanctuaries for proviral HIV (White blood cells known as macrophages can serve as reservoirs, too, but they are relatively short-lived.) Because current treatment plans seem unlikely to cure HIV infection within several years and because of HAART's various drawbacks-including side effects, complicated regimens and resistance-the search is on for additional drugs. The first entries to reach the market will add new choices within existing drug classes, and some will combine existing drugs into a single pill or will otherwise reduce the complexity or toxicity of current regimens. And at least two custom-designed protease inhibitors in clinical trials appear to work against strains of HIV that are resistant to existing protease inhibitors.
Other Therapeutic Ideas
Other agents under consideration would stall HIV replication in new ways. Some would block the integrase enzyme from inserting HIV DNA into a cell's chromosomes. A different strategy attempts to knock zinc out of a protein that needs the metal in order to draw HIV RNA into new particles.
Teams are also exploring ways to stop infected cells from making critical viral proteins. One such approach deploys so-called antisense DNA to inactivate two genes (tat and rev) that normally give rise to proteins needed for the efficient manufacture of other viral proteins. In rhesus macaques exposed to the simian version of HIV, this therapy limited viral replication and CD4 T cell depletion.
Many investigators are attempting to interdict HIV's entry into cells. Recall that to fuse with, and gain entry into, a target cell, the virus must attach both to CD4 and to a co-receptor on the cell surface. Initial attempts to interfere with binding to CD4 were disappointing, but new possibilities have recently come to light, and many groups are examining compounds that might sheath the HIV docking site on co-receptors to keep HIV at bay [see "In Search of AIDS-Resistance Genes," by Stephen J. O'Brien and Michael Dean; Scientific American, September 1997].
A number of scientists are focusing on the other part of the equation: the immune system. They are trying to augment the body's remaining forces or to restore lost powers. For instance, some patients are receiving low doses of a biological molecule called interleukin-2, which enhances the proliferation of T lymphocytes. It is also hoped that interleukin-2 can help force immature "stem cells" to spawn a full repertoire of fresh new cells of the immune system, including T lymphocytes and antibody-producing cells that can recognize and eliminate HIV
Immune-reconstitution efforts are taking additional forms as well - such as harvesting stem cells from an HIV-infected patient, multiplying them in the laboratory and then returning the enlarged cell population. A twist on this scheme would supply the infused cells with a gene that would protect them from later succumbing to HIV
Other approaches being pondered might kill HIV infected cells without relying on the immune system. One would deliver a virus that had been genetically altered to enter only HIV producing cells, leaving HIV free cells untouched. It would enter by attaching to viral proteins displayed on a colonized cell and would then destroy the cell.
As time goes by, patients infected with HIV are sure to have more and more options for therapy. Specific recommendations may change from time to time, but the principles of therapy, now grounded in solid research, will remain steadfast: until there is a cure or a vaccine, controlling HIV replication offers the best chance for a long, productive life.
The fact that such insight and the tools to suppress HIV reproduction are available is both astonishing and wonderful. The therapeutic advancement achieved since late 1995 has few parallels in the history of medicine, save perhaps for the revolution sparked by the introduction of penicillin. Just three years ago those of us who cared for HIV patients mainly devoted our time and energy to easing symptoms and preparing patients to die. Now we help patients to live. The war against HIV is far from over, but the success of aggressive therapy is certainly a victory to be savored.

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