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 Biomedical Developments in AIDS Research
By
Suzanne Jurriaans
  
Keywords:  Aids; Disease diagnostics (human); Drugs (human). 
Correct citation: Jurriaans, S. (1997), "Biomedical Developments in AIDS Research." Biotechnology and Development Monitor, No. 30, p. 2­5. 

Huge scientific efforts aim at understanding AIDS and its subsequent prevention and treatment. However, the high cost of diagnostics and therapies prevent their worldwide use. Moreover, differences in HIV variants make some technologies less applicable to people in developing countries.

In 1981 the first reports on Acquired Immunodeficiency Syndrome (AIDS) appeared.  Three years later, Human Immunodeficiency Virus (HIV) was identified as the cause of AIDS. Until then no human life threatening diseases were linked to a virus. The route of HIV infections differs between industrialized and developing countries. HIV infection in the developed countries spreads mainly by anal intercourse among homosexual men and by contaminated blood among intravenous drug users. In the days prior to heat treatment of blood products and donor screening, many haemophiliacs were infected. For example, between 1979 and 1985, over 10,000 haemophiliacs were infected in Europe and North America. In addition, infections occur among children born to HIV­infected mothers. In Africa and Asia, the main route of HIV transmission is heterosexual intercourse. Central and East African countries are suffering from an AIDS epidemic; it is estimated that 20 per cent of sexually active adults are infected.
While the rate of developing AIDS upon infection is very high, its clinical manifestation is slow to develop. Over 95 per cent of HIV­infected individuals develop AIDS. Of this group, only one per cent may develop AIDS after one to two years. Another one to two per cent do not develop AIDS even after 15 years, while the majority of HIV­infected persons develop clinical manifestations of AIDS within 10 to 12 years. This implies that people who carry HIV may look and feel healthy for a number of years, but meanwhile they can transmit the virus to others.
With AIDS, virology has become an important field in the management of prevalent infectious diseases. Most of the research is performed in the industrialized countries such as USA, Canada, United Kingdom, France, Italy, the Netherlands and Belgium. The research is largely funded by pharmaceutical industries. Governments, universities and non­governmental organizations (NGOs) are also funding a number of research projects. The budget for AIDS research in Western countries far exceeds the total budget for medical care in developing countries.

Diagnosis of HIV infection
There are different methods to detect the HIV infection:

  • Immunology. A characteristic of HIV infection is the depletion and functional impairment of CD4+ lymphocytes (See box). CD4+ cells in blood can be counted and their level reflects a person's immune status. The decline of CD4+ cell count is the most significant laboratory parameter. CD4+ cell count starts to decline relatively soon after HIV infection. However, it declines at different rates in different individuals. Another problem with this test is that it often detects the HIV infection late, when clinical manifestations of AIDS have already appeared.
  • Serology. The first available tests for HIV infections consisted of the detection of antibodies in the blood. This involves repeated blood screening for HIV antibodies by enzyme­linked immunosorbent assay (ELISA). This is confirmed by a similar test, the Western blot analysis. The HIV screening tests have also been used to identify risk factors for the acquisition of HIV.  The test for HIV antibodies has greatly contributed to the control of the HIV epidemic in Europe and the USA. However, in many developing countries, blood supplies are often not screened for HIV.

  • The detection of antibodies to the HIV core proteins does not only diagnose HIV infection. It can also predict the clinical course of the infection. Antibodies to the core and envelope proteins of HIV become detectable in serum within two to six weeks after primary infection. The emergence of antibodies reflects the effort of a person's immune system to eliminate the virus. When antibodies to viral proteins disappear from a person's blood, s/he is no longer able to elicit an antibody response and this indicates rapid clinical progression.
    Before the appearance of antibodies, the HIV p24 antigen, one of the viral core proteins, can be detected in serum. The presence of HIV p24 antigen indicates rapid disease progression. Hence, it is a relatively early marker. Unfortunately, a substantial number of HIV­infected individuals do not have detectable levels of p24 antigen. They progress to AIDS in the absence of this prognostic marker, making the screening of p24 antigen an unreliable indicator.
  • Nucleic acid based diagnostics. In recent years improvements in the field of diagnostic assays have expanded from immunoassays to nucleic acid amplification­based assays. The most significant advantage of nucleic acid amplification­based assays is its extreme level of sensitivity. Amplification technologies are based on multiplication of viral nucleic acids to a quantity that is easily visible. The development of molecular amplification procedures has enabled the specific detection of HIV genetic material (instead of proteins) within the host. Moreover, direct measurement of viral activity and detailed study of viral characteristics are now possible. The uniqueness of the viral genetic sequence is responsible for the high level of specificity obtained with amplification technologies. A number of amplification principles have evolved and are commercially available. They are: Polymerase Chain Reaction (PCR) Amplicor HIV Monitor developed by Swiss Roche Molecular Systems, NASBAHIV­1 QT­RNA developed by Organon Teknika of the Netherlands, and Branched DNA Signal Amplification (bDNA) developed by the US Chiron Corporation.  While these assays are generally available worldwide, the cost certainly limits their use in developing countries. The assays cost US$ 100 to 150 per test. Moreover, the commercial PCR and NASBA tests are developed and optimized for HIV variants circulating in Europe and North America. Due to the different HIV variants, these test are less reliable when, for example, detecting African HIV virus types. The bDNA assay is currently the only amplification­based assay that detects all HIV variants with equal efficiency. Roche Molecular Systems and Organon Teknika are also trying to modify their assays to detect all HIV subtypes with equal efficiency.

  • The detection of HIV nucleic acids provided insight into the most fundamental aspects of HIV infection. Conventional immunoassays often fail to detect viral components in the blood of many infected individuals. Moreover, the test affirmed that HIV is the cause of AIDS. Through nucleic acid amplification techniques, it was shown that HIV production occurs during all phases of HIV infection. It even occurs during the so­called clinical latency or inactive period. The turnover of virus and infected cells was found to be much greater than previously thought. Millions of virus particles are produced daily within the body of an infected individual. This technology established that the decrease in CD4+ cell counts occurs as a result of viral replication.

    The HIV particle and the viral life cycle

    The HIV is a very small sphere, which contains the viral genetic material surrounded by viral proteins (the so­called core). The outside of the virus consists of a lipid bilayer (which is derived from the infected cell) spiked with viral proteins (together called the envelope). HIV attacks cells of the immune system called lymphocytes, specifically its subset known as a helper T­cell. Present on the outside of these T­cells is a protein called CD4. These CD4+ lymphocytes have an important role in almost all immune responses. The loss of these cells causes a severe defect in the host defense mechanism against pathogens. This renders a person susceptible to all kinds of infections. 
    As a parasite, the virus is incapable of independent reproduction. HIV is a member of the retrovirus family. The virus has to copy its RNA into DNA in order to survive in the cellular environment. To reproduce, HIV is able to bind to the CD4 receptor through its envelope proteins and penetrate the cell. Once inside the cell, the virus releases its genetic material, the viral RNA. This is subsequently transcribed in viral DNA by reverse transcriptase, a protein which is also present in the virus particle. The viral DNA copy is then inserted into the cellular DNA. During this transfer process, mistakes are being made. Through these mistakes, the virus changes a little during every round of replication. These mistakes have both a disadvantage and an advantage for the virus. The disadvantage is that some mistakes results to 'crippled' viruses, which are unable to replicate and consequently 'die'. The advantage for the virus is that some mistakes results in changes wherein the virus becomes unrecognizable and hence escape from the immune response. This could also render the virus less susceptible to antiviral drugs. 

    During the acute phase of infection, HIV replicates at high levels in the blood. Following the emergence of an immune response to the virus, there is a decline in the level of viral nucleic acids in the blood. In most individuals, a further reduction in the number of HIV particles is observed during the asymptomatic phase of infection. Two distinct groups of HIV­infected people have been identified. The first group seems to be able to reduce the number of circulating virus particles to a very low level for a prolonged period. These individuals were found to be long­term asymptomatic. The second group are those HIV­infected persons who lack the capacity to further reduce the number of circulating virus particles. These individuals could be stratified as being rapid progressors. This is based on the HIV RNA count in the blood. This occurs before the presence of any symptoms or other markers (CD4+ cell count, presence of p24 antigen) direct towards disease progression. The stratification of patients on the basis of viral RNA levels in the blood is predictive of their progression to AIDS or death.

  • Diagnosis of neonatal infection. HIV infection in newborns can only be confirmed through direct detection of viral components, and not antibodies, in their blood. This is because HIV antibodies are transferred from the mother to the infant before birth. The HIV antibodies remain detectable in infected and in uninfected infants of HIV­positive mothers more than a year after birth. Fortunately, HIV nucleic acid based diagnostics can be done shortly after birth. It can detect HIV RNA in venous blood of an infant from an infected mother. This is important so that infected individuals may benefit from early treatment.

  • Antiretroviral therapy
    Besides developments in diagnostics, there is also progress in antiretroviral therapies and their evaluation. HIV RNA is a good surrogate marker to evaluate the effectiveness of antiretroviral therapy. The first clinically available compound with anti­HIV activity was zidovudine (AZT). It was developed and is sold exclusively by UK Glaxo­Wellcome. The therapy aims to disrupt the HIV reproduction. AZT chemically resembles one component of the DNA. Hence, AZT can 'trick' the virus into incorporating AZT into the DNA molecule during the conversion of RNA into DNA. However, the other components necessary to build the DNA cannot be attached to the AZT molecule. This impairs the reverse transcriptase of RNA to DNA. A number of trials demonstrated that zidovudine monotherapy had a beneficial effect to people who had already developed AIDS. It has been shown to postpone death by one to two years. However, the drug failed significantly to benefit asymptomatic HIV­infected people.
    Recently far more potent drug regimens have been introduced consisting of three or more antiviral components. These protease inhibitors work in a similar way to AZT. However, the combination therapy does not only arrest the reverse transcription of the virus. These drugs further block the HIV protease, an enzyme needed to produce the viral proteins. In some cases, it has been shown that under combination therapy, HIV RNA levels are dramatically reduced by as much as 100 to 1000 times within one to two weeks after the start of treatment. However, if there is no significant decline in this period, this indicates that the drug regimen is not effective for the individuals involved.
    The recent regimens have significantly decreased the levels of HIV RNA in the blood of most infected people. In addition, it is most probable that these individuals now have a good clinical prognosis. However, time will prove whether this assumption is true, and whether these individuals will still progress to AIDS despite the low HIV RNA level.
    These combination regimens are commercially available. Examples of protease inhibitors are: Ritonavir produced by US Abbott; Indinavir produced by US Merck, Sharpe and Dohme; and Saquinavir, produced by Roche Molecular Systems. Examples of reverse transcriptase inhibitor are: 3TC produced by Glaxo­Wellcome; D4T produced by US Bristol Meyers Squibb; and Nevirapine produced by Boehringer Ingelheim of Germany. However, the high cost of these drugs and tests greatly limits their usage. For example, even for a developed country like the Netherlands, funding for the evaluation of the therapy, such as  HIV RNA load testing is already difficult. Moreover, the efficacy of antiviral treatment on different HIV variants remains to be established. Due to the recent introduction of combination therapies and the limited availability of antiviral drugs in developing countries little is known  about this subject.

    Conclusion
    In a relatively short period of time (about 15 years), the world has learned a lot about AIDS: the cause, the pathogenesis, and even (in part) the treatment. At present there is reason believe that AIDS may eventually become a treatable disease. However, due to the cost and intensity of medical care (i.e. treatment regimens must be tuned to each individual) this hope remains mainly restricted in the industrialised world. Therefore, research must continue to develop a vaccine against HIV, which can be supplied to the developing countries on a large scale and at much lower cost. V
    Suzanne Jurriaans

    Department of Human Retrovirology Academic Medical Centre, University of Amsterdam
    Meibergdreef 15, 1105 AZ Amsterdam, the NetherlandsFax (+31) 20 691 6531.

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    S. Jurriaans, B. van Gemen, et. al. (1994), "The Natural History of HIV­1 Infection: Virus load and virus phenotype independent determinants of clinical course?". Virology, Vol. 204, pp.223­233.

    S. Jurriaans, G. J. Weverling, et. al. (1995), "Distinct Changes in HIV type 1 RNA Versus p24 Antigen Levels in Serum During Short Term Zidovudine Therapy in Asymptomatic Individuals With and Without Progression to AIDS". AIDS Res Hum Retroviruses, Vol. 11, pp. 473­479.

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