Important note: Information in this article was accurate in January 2002. The state of the art may have changed since the publication date.
American Foundation for AIDS Research, January 2002
Anne-christine d’Adesky
Three million people throughout the world died of AIDS last year, of whom 2.3 million were Africans. The enormous scale and relentless pace of [quote]the HIV epidemic has fueled activists’ demands for wider access to HIV treatment. The Indian generic drug producer Cipla has dropped the cost of a generic three-drug anti-HIV regimen - d4T, 3TC and nevirapine - to a record low price of $38 (1,800 rupees) a month.
With affordable therapy almost at hand, tremendous pressure exists to deliver HIV drugs quickly to Africa as well as to Asia, where a simmering epidemic threatens to explode. The equation has a major new element: political will at the highest echelons of world government, backed by money, or talk of money. In the wake of last summer’s United Nations Special Session on AIDS, a new Global Fund for HIV, Malaria and TB was created to help buy drugs for poor countries.
Though drug prices have fallen, most tests considered routine for HIV management in the West are simply out of reach elsewhere. Viral load tests cost an average of $100, excluding equipment and laboratory facilities. The cheapest flow cytometers for measuring CD4 T-cell counts cost $75,000 to $100,000, while other machines are for sale in the $40,000 to $80,000 range. Added to this expense is money needed to train and hire technical staff, maintain and repair broken equipment, ship and store samples. Serge Diagbouga, an HIV doctor at the Centre Muraz in Burkina Faso, remarked, “There’s no question about our ability to afford it: we just can’t. We need alternatives and, unfortunately, we need them soon.”
Diagbouga continued, “Now that treatment is becoming available, the government is making a lot of effort to decrease the price of [anti-HIV drugs]. But we need to know the criteria before we put the patients on treatment. I don’t think we can permit ourselves to distribute anti-HIV drugs to patients across Africa the way we give out aspirin. What we need is some kind of consensus for us in developing countries with limited money and resources, to get a panel of techniques that work not only for CD4 T-cell count, but also for viral load, that we can work to distribute and make accessible in our country, so that we can correctly follow our patients.”
Diagbouga was among a handful of African physicians attending a recent meeting entitled “Monitoring and Diagnostic Tools for the Management of Antiretroviral Therapy in Resource-Poor Settings,” held in Bethesda, MD last November 11-13. It was cosponsored by Gay Men’s Health Crisis and Project Inform. In attendance were frontline field workers and HIV researchers from around the world, including U.S. government scientists and representatives of equipment manufacturers who are developing an array of immunologic, virologic and clinical monitoring techniques for HIV care.
There were no arguments at the meeting concerning the need for cheaper and simpler laboratory assays for monitoring HIV and its treatment. These tools are needed in settings of abject poverty, where there is frequently no electricity, refrigerators or running water, and no technician or doctor to administer the tests.
Doctors in Africa can be few and far between in rural areas, where the majority of people with HIV live, making it imperative that minimally trained staff be able to perform the tests. Test results would be available on the same day as the visit to the testing center, since rural Africans usually live far from these centers. Finally, with few laboratories in place, samples that do require shipping and storage must be capable of withstanding heat and the rigors of travel without decomposing or otherwise compromising a test’s accuracy.
Overall, frontline researchers advocate a more comprehensive approach to HIV clinical testing in impoverished, medically challenging settings. In an ideal world, the next generation of laboratory technology should be capable of monitoring the impact of multiple chronic infections and therapeutic regimens at once. Tests should also be able to follow overlapping drug toxicities, side effects, and drug resistance on the part of HIV and other pathogens. Instead of a one-size-fits-all approach, veterans like Diagbouga advocate a more individualized approach that allows countries to adopt cost-effective technologies best fitted to their specific capabilities and health conditions.
In moving ahead, it is also important to consider the goal of lab monitoring and the use of a particular tool at a particular time, in order to assess its relevance, a point underscored at the conference by Donna Mildvan of Beth Israel Medical Center. HIV monitoring is used not only for disease management, but also in epidemiology, to screen blood, and for research purposes. Reviewing the state of lab technology in most of Africa, Dr. Charlie Gilks of the Liverpool School of Tropical Medicine and the World Health Organization was blunt about the need for researchers and manufacturers to engage in innovative thinking when developing new testing methods. “Most of the discussion about these tools is skewed [by an assumption] that there will be laboratory technologies to do the testing,” he said. “But there is a relative scarcity of trained cadres and technologists. Bear in mind that there are very fragile health systems that are being overwhelmed by the HIV epidemic.” As he noted, HIV has not spared health professionals in the worst hit African countries. “In a country that has a 20% prevalence rate for the population, 20% of the medical and laboratory staff will be infected as well. It’s a double whammy for the health center, having to deal with the burden of disease and the consequences of a shrinking work force affected and infected with HIV.”
“I can design the best toys out there, but what’s the point if no one can use it?” asked Dr. Helen Lee of the University of Cambridge, who is regarded by her peers as a master lab test designer. Surveying the super-tech assays now used in the West - such as phenotypic resistance testing - she remarked, “We’ve all got a million ideas and tricks up our sleeves that may not help anyone in Africa. The question is, what can we do that will actually make a difference?”
Lee drew knowing laughs from colleagues by comparing the qualities of her ideal lab test for poor settings to a lover doing the Kama Sutra: it has to act quickly (results in 30 minutes); be simple (requiring 10 minutes to train a nontechnician to use), cheap (costing 50 cents at most), stable (having a one-year shelf life), noninvasive (does not use syringes to draw fresh blood), and multiplex (capable of testing more than one sample at a time). Once the tool is ready, it typically takes years and a lot of money to test for efficacy in human trials, win FDA approval, and bring to the open market. “Now,” she grinned, “which one of you is ready to fund me to do that?”
The existing clinical guidelines for managing patients on HAART (highly active antiretroviral therapy containing three or more anti-HIV drugs) derive from clinical studies showing that a three-drug regimen can effectively lower HIV plasma viral loads below detectable levels, using polymerase chain reaction (PCR) to indicate HIV virion concentrations. In many patients on HAART, CD4 T-cell counts rise as viral load drops. An algorithm based on these two surrogate markers is now used to guide physicians on the impact of HAART on a patient’s health, risk of developing OIs, and prospect of disease-free survival.
For now, low-tech realism is taking precedence over loftier high-tech dreams. There was general agreement at the meeting - and a lot of debate - that clinical monitoring is the new Holy Grail. African doctors are hoping for a “no-tech” minimum standard for diagnostic and treatment evaluation that lays out basic criteria and clinical markers for monitoring patients on HAART, and beyond that, how and when to incorporate more expensive laboratory tests should they become available.
The simplest approach, “syndromic” management of HIV, relies on physical symptoms rather than laboratory values to gauge the impact of therapy on patients’ disease status. The appearance or disappearance of physical signs of disease [quote]can help doctors in the field determine when to start or switch a given therapy and to monitor side effects, drug resistance, and drug failure. Syndromic management can be used as an adjunct to or even a substitute for more expensive viral load or CD4 T-cell tests.
As Brazilian researcher Mauro Schechter pointed out, such a low-tech monitoring strategy for Africa or Asia does not rule out future use of the gold standard of viral load. “The perfect is not the enemy of the good,” he said, an opinion echoed by others at the meeting. There is little solid information on how well clinical monitoring stacks up to the laboratory gold standard. Advocates cite the few existing comparative studies as promising evidence of a viable alternative.
In 1994, Gilks and colleagues in Uganda initiated a natural history HIV study in Entebbe that compared disease symptoms (based on the CDC clinical definition of HIV disease) to CD4 T-cell counts as a prognostic marker of disease-free survival or death in 201 HIV-positive individuals. They were enrolled in the study from October 1994 to January 1995 and were followed up until the end of 1997. In a December 1999 Journal of AIDS report, the Uganda team observed, “Baseline CD4 T-cell count distribution showed clear and appropriate associations with clinical stage in the 201 [Ugandan] participants. Clinical monitoring has proven to be an effective alternative to CD4 T-cell testing, based on the 201 participants. Both provided equivalent prognostic information.”
With these results and others in mind, Gilks put forth what he calls “the roughest guidelines” for implementation of HAART that rely on the CDC clinical staging criteria. CD4 T-cell counts are used only to assess how well therapy is working. Viral load tests are not used at all. Instead, markers like increased body weight, decrease in OIs and restoration of energy are taken as signs of effective therapy. Forced to choose, most physicians at the Bethesda meeting admitted they would use a CD4 T-cell test over a viral load, and might even save that money. “If I have a few rupees to spend, I’d rather spend them on liver enzyme than viral load or CD4 T-cell,” said Indian researcher Dr. N.M. Samuel of The Tamil Nadu M.G.R. Medical University in Chennai, India. “In reality, a clinician will see a patient after the first week, then every two weeks. If the patient is not doing well, they will do liver enzymes, then look at jaundice. Most patients have been managed that way.” In South Africa, Mark Cotton of Tygerberg Children’s Hospital in Western Cape Province remarked, “We could treat a patient for almost a month for the cost of a viral load test.”
Those pushing for an alternative standard assume that most patients in poor countries will be given non-protease HAART regimens and will have a very limited choice in switching regimens. “I’d say one initial course of treatment, then one switch,” predicted Gilks. “And maybe a T-cell test somewhere down the line.”
Under the proposed “no-to-low” tech guidelines, HIV patients would be screened and treated for TB before starting HAART to avoid negative drug interactions between drugs that affect the liver. At that time, hemoglobin, liver and renal tests would be done, too. Based on CDC staging criteria, those with AIDS or clinical symptoms of HIV disease, or with no symptoms but with CD4 T-cell counts below 200, would qualify to begin HAART. Clinical evaluation of patients would take place at month 1, with basic biochemistry and CD4 T-cell counts at month 4, and when possible, a viral load at month 6. Others advocate a similar model, with slight variations in the timing and schedule of follow-up tests.
Some researchers, including Gilks, have proposed using basic blood tests, including a total lymphocyte count, or TLC, as an alternative to a CD4 T-cell count to assess the impact of therapy on survival. The TLC measures non-T-cell lymphocytes (B-cells and natural killer cells) as well as T-cells, with the latter making up the biggest portion of the TLC. This approach requires only a light microscope, whereas CD4 T-cell assays require expensive flow cytometry (see box). Although major hospitals in Africa have flow cytometers, it is a relatively expensive technology and requires a trained technician to operate.
How good is TLC? “Although total lymphocyte count doesn’t predict who does badly [on treatment], it did reflect on survival,” Gilks said, referring to his own studies. “It is a pretty good prognostic that also correlates with CD4 T-cell count.” In a presentation on surrogate markers at the Bethesda meeting, Victor De Gruttola of the Harvard University School of Public Health also weighed in on the adjunctive value of TLC: “Longitudinal monitoring of lymphocytes could have value for disease progression,” he remarked. In small studies of people with HIV, De Gruttola found that, “the mean value of TLC drops 12% a year, so it is meaningful in people after they develop AIDS.”
In Senegal and Ivory Coast, clinical staging has been used successfully in small pilot projects, along with the use of “Dynabeads” to gauge CD4 T-cell counts. This technology uses magnetic beads or particles that are coated with antibodies to T-cells’ CD4 receptor. The results can be read by standard light microscopy. In a study done in seven African laboratories, Dynabead results correlated well with those from CD4 T-cell tests performed by flow cytometry.
Like TLC, Dynabeads are a labor-intensive technology that requires time-consuming manual counting. But it beats having a more sophisticated machine that requires steady electricity and someone to operate and maintain. The Dynabead system is far cheaper than flow cytometry - only $2,000 for the equipment and $5 per test. It takes only three days to train a technician to use it.
Even if feasible testing technology exists, African researchers are a bit worried about accepting guidance from lab assays whose standards derive from the blood or immune profiles of non-Africans. Little is known about what constitutes “normal” blood levels or T-cell levels among different African populations, including those with HIV. Chronic endemic diseases like TB, malaria and parasites can suppress immune function on their own to a certain extent. TB, for one, can cause a 20% drop in total lymphocyte count that does not completely rise to baseline levels after successful TB treatment. Similarly, parasites can lower hemoglobin levels.
What about alternatives to a PCR viral load test? This virologic tool is currently used to evaluate the success of HAART. Insufficient suppression of HIV indicates the appearance of drug resistance or some other problem that reduces treatment effectiveness. PCR tests are too pricey in developing countries to be used outside of research purposes, even at the current specially reduced price of $100 per test. A cheaper choice may be the Retina HIV-1 Rainbow “real time” NASBA assay that uses a different technology than PCR and can detect all HIV-1 subtypes.[quote] It yields results in just two hours, and in early studies, it was comparable to the standard but more expensive NASBA - the NucliSens HIV 1 assay by Organon Teknika, which delivers results within 60 minutes and is FDA-approved. The Retina assay will be offered at only $20 to $25 outside the U.S. market. This is still too expensive for many countries, unfortunately.
The TaqMan PCR assay is a cheaper option that involves a simple, one-step test similar to standard HIV PCR assays. Its advantage is that it can measure 90 specimens at once, with results in two to three hours. The main drawback is the large initial investment needed to buy the system.
A promising alternative to viral load is a heat-denatured p24 antigen test, in which HIV levels are inferred from the detected amount of free HIV core protein. This assay costs $6 to $8 in Africa. Jörg Schüpbach of the University of Zurich found that the sensitivity and specificity of the heat-treated p24 antigen test is comparable to existing PCR viral load tests. In a study of nine adults on HAART, the p24 antigen disappeared as quickly as viral RNA. In a larger comparative study of 155 patients on HAART who were followed for 154 days, the p24 antigen test was better at predicting survival than HIV PCR and is a good predictor of subsequent disease progression.
Many HIV experts are excited about using dried blood spots and heel sticks (a pinprick to the heel to get a drop of blood) rather than larger syringes that require refrigeration. This well-established, simple technology is routine in pediatric care and uses a dipstick, or filter paper. Specimens can be put into a sealed plastic bag and shipped and remain stable at room temperature for up to a year. Heel-stick methods are less invasive and may be especially useful for monitoring pediatric HAART patients.
According to Susan Fiscus of the University of North Carolina, the dried blood spot technique is also applicable for HIV antibody measurements as well as actual virus levels. It is applicable for both gene sequencing and resistance testing, too. Fiscus explained, though, “The only thing that is less about the dried blood spot is the collection and storage. There’s no need for trained phlebotomists or freezers. In order to do a viral load, you still have to do a commercial viral load assay, so it gets to be $50 to $100. The best thing would be to elute the p24 from the dried blood spot and use it in Schüpbach’s enhanced p24antigen assay. We are working on that now.”
And what about tests to monitor potential resistance to HAART? Here, the picture is less hopeful while the threat is serious. One recent U.S. report found that half of patients on treatment for at least three years harbor HIV with drug-resistance mutations (see page 6). The Darwinian fact remains that HIV constantly mutates, and the presence of drugs favors the HIV containing mutations for drug resistance. Given what has occurred in the U.S., it is likely that many people in poor countries will develop HIV that is refractory to treatment. Without cheap tools to monitor resistance, how can we directly assess the scale of this threat?
Given resistance tests’ cost - $400 to $1,000 - most clinicians confronting limited resources restrict their use to population-wide surveillance of emerging drug-resistance patterns. For individual patients, doctors stress adherence to dosing schedules so as to maintain optimum drug levels in the body. Adherence can be improved by prescribing simplified regimens and following the Directly Observed Therapy (DOT) model used in TB control. The DOT model for HIV is gaining support (see the article on the Partners in Health program in rural Haiti that appeared in the December, 2001 Treatment Insider). But the hard reality is that, despite the availability of DOT, multidrug resistance to TB is spreading wildly across many parts of the world.
Dr. Salif Sow of Senegal acknowledged, “We need to look at the failure of TB control. [Laboratory tests] must be seen in the overall context of health care needs. It is only a tool in a very complex puzzle.” Clinical studies to determine how best to monitor and manage multidrug-resistant HIV in poor communities are now critically important.
Despite these myriad challenges, the prevailing attitude among frontline doctors and testing experts has shifted to optimism. In Burkina Faso, Diagbouga said, “We are no longer talking about whether[quote] to try to treat with [anti-HIV therapy], but how, and when, and where, at what cost.” As he sees it, significant ethical issues face African governments as they move to implement treatment in the absence of high quality monitoring technology. But they are not a reason to delay treatment. “It’s true that you have to make choices: Do I [start treating] to provide the person with the hope that they can live a few more years? Or do I avoid intervening in order to wait for a complete battery of tests? In terms of human ethics, we can’t cross our arms and wait. We simply have no act, because too many of our people have been dying.”
Gilks agreed, adding a note of caution, “The knowledge gap shouldn’t stop us but should just make us be very clear as we move forward. We have to very carefully evaluate what works and what doesn’t work. We need to be very clear that we will make mistakes and that is going to be very uncomfortable for some international organizations. And we have to be very open about that.”
If test designers have their way, existing CD4 T-cell count and viral load tests will soon seem as bulky and outdated as a CD compared to an LP. They will become smaller, faster, and involve cheap, mass-produced parts. Boston-based immunologist Dr. Howard Shapiro wants to use inexpensive light-emitting diode lasers, plastics, digital technology and new antibodies to this immunofluorescent equipment that is cheaper to use and easier to maintain than flow cytometers. He predicts that a prototype could be built in less than a year.
Another new option is multiplex microgene chip technology. It is behind new products like the HIV diagnostic “Taste Chip” under development by John McDevitt at the University of Texas at Austin. These gene chips allow for simultaneous detection of many infectious microorganisms, for example hepatitis B, C, HIV, CMV, and mumps. McDevitt is now working on chips for diagnosing acute and chronic HIV. Best of all, the tests will be cheap, costing pennies, not dollars, and the equipment will run on a simple battery.
But who will fund and mass-produce such products? “Is the industry ready to market products for countries that don’t have much money?” wondered Shapiro while challenging U.S. lab test manufacturers to do just that. Several years ago, Shapiro points out, the TRAx CD4 enzyme immunoassay was produced by Avant Immunotherapeutics as another cheap, validated alternative to flow cytometry for CD4 T-cell counts. Avant sold the product to Immunogenetics. Today it is off the market, an abandoned, but still promising technology.
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