Recent Scientific Contributions to Understanding HIV / AIDS from the Multicenter AIDS Cohort Study

1 <i> University of California, Los Angeles, School of Public Health, U.S.A.</i> 2 <i>Northwestern University Medical School, U.S.A.</i> 3 <i>Johns Hopkins University School of Hygiene and Public Health, U.S.A.</i> 4 <i>University of Pittsburgh Graduate School of Public Health, U.S.A.</i> 5 <i>National Institutes of Health, National Institute of Allergy of Infectious Diseases, U.S.A.</i> 6 <i>National Institutes of Health, National Cancer Institute, U.S.A.</i> This study was supported by NIH Research Contracts NOI-AI-72631, 72632, 72634, 72676, and 32535. * The Multicenter AIDS Cohort Study (MACS) includes the following: Baltimore: The Johns Hopkins University School of Hygiene and Public Health: Alfred J. Saah, Principal Investigator; Homayoon Farzadegan, Neil Graham, Joseph Margolick, Justin McArthur. Chicago: Howard Brown Memorial Clinic-Northwestern University Medical School: John P. Phair, Principal Investigator, Joan S. Chmiel, Bruce Cohen, Steven M. Wolinsky. Los Angeles: University of California, Los Angeles-Schools of Public Health and Medicine and the UCLA Jonsson Comprehensive Cancer Center: Roger Detels, Principal Investigator; Barbara R. Visscher, John L. Fahey, Jan Dudley, Janis V. Giorgi, David T. Imagawa, Otoniel Martinez-Maza, Jeremy Taylor. Pittsburgh: University of Pittsburgh Graduate School of Public Health: Charles R. Rinaldo, Jr., Principal Investigator; Lawrence Kingsley, Phalguni Gupta, James Becker, Monto Ho. Data Coordinating Center: The Johns Hopkins School of Hygiene and Public Health: Alvaro Muooz, Principal Investigator; Noya Galai, Donald Hoover, Lisa P. Jacobson, Curtis Meinert, Sol Su,Steve Piantadosi. NIH: National Institute of Allergy and Infectious Diseases: Lewis Schrager, Project Officer, Richard A. Kaslow, Sten Vermund, Mark J. VanRaden; National Cancer Institute: Iris Obrams, Daniela Seminara


INTRODUCTION
The Multicenter AIDS Cohort Study (MACS) was established in 1984 when four groups of investigators from the Johns Hopkins School of Hygiene and Public Health (Baltimore), the Northwestern University School of Medicine (Chicago), the University of Pittsburgh School of Public Health, and the University of California, Los Angeles (UCLA) School of Public Health elected to collaborate in the conduct of the cohort studies which they were initiating in each of these cities.In 1986 the Data Coordinating Center was established at the Johns Hopkins School of Hygiene and Public Health.The MACS has been funded through September 1995 by the National Institute of Allergy and Infectious Diseases with the collaboration of the National Cancer Institute.
In the first years of the study, the MACS identified the risk activities for transmission of HIV-1 among homosexual men(1.2.3) and the relationship of CD4 cell levels to risk of AIDS(4.5.6.) .In the last several years the MACS has concentrated on examining the nature of the immune response to infection by HIV-1, the factors enhancing or predicting rapid progression of infected individuals to clinical AIDS, the incidence of malignancies and factors associated with their occurrence, and the impact of therapy on the course of AIDS.Since 1985 the MACS has published over 200 papers in peer-reviewed journals.Because it is impossible to summarize all of these papers in this presentation, we shall present only selected observations and findings of the MACS in recent years.The success of the MACS has been largely due to the commitment and generosity of the men participating in the study who have given willingly and frequently of their time, responded to the most personal of questions, and provided repeated biologic specimens.

METHODOLOGY
The original objectives of the MACS have been described in two papers by Kaslowl( 7) Chmiel (8), and their colleagues.Briefly, homosexual/bisexual men in Los Angeles, Chicago, the Pittsburgh.tri-statearea, and the Baltimore/Washington area were invited to participate in the MACS in 1984 and 1985.Men were recruited through public service announcements in the media, presentations to homosexual organizations, recruitment by early participants, and visits to gay bars and baths.No effort was made to recruit a "representative" group of men.Although whites represent a minority in the four cities, the majority of the men recruited into the MACS were white and had at least some college education (see Table 1).
Men were eligible to participate in the MACS if they did not have signs or symptoms of AIDS at baseline and were 18 years of age or older.Men are considered to have a diagnosis of AIDS if they met the 1987 CDC criteria" and are reported by their health care provider as having AIDS, or if AIDS is indicated on their death certificate.Eight years after the initiation of the MACS, the health status of eighty-five percent of the men is known , an unusually high rate of compliance which reflects the commitment of the men in the MACS to contribute to the advancement of scientific knowledge of HIV/AIDS.
Every three to six months, men participating in the MACS respond to an interview schedule soliciting information on demographic factors, habits, disease history (including history of sexually transmitted diseases), and past and present sexual activities (including numbers of partners and frequency of specific sexual acts) .Each man is given a brief physical exam to identify signs of HIV-1 infection and, at the same time, provides a blood specimen .Cells and serum are divided and those not used for determination of HIV-1 antibodies and levels of T cells are stored in both a local and a national repository .The occurrence of AIDS and clinical conditions such as malignancies are documented every six months and verified by a physician's report and by tissue specimens whenever possible.
The antibody status of the men at entry and changes in their antibody status are determined by the enzyme-linked immunosorbent assay (ELISA) test (Genetic Systems and Dupont) .A blood specimen is considered positive if the ELISA test gives a reaction which is above the cutoff value for absorbance and is confirmed by the presence of a positive band in at least two of the three gene product regions of the Western blot (Biorad) represented by the gag, env and pol proteins (10).
Levels of CD4 and CD8 cells were determined on whole blood samples using flow cytometric procedures and monoclonal antibodies (11).Numbers of CD4+ and CD8+ cells were determined by obtaining total and differential white-cell counts and multiplying by the appropriate factor obtained on flow cytometry.
Neopterin is a product of stimulation of macrophages.B2M is a product of generalized lymphocyte activation.IgA is a marker of B cell activation.Levels of sIL-2R reflect activation of CD4 cells.
Standard National Institute of Health microcytotoxicity techniques were used to type peripheral-blood mononuclear cells for gene antigens (15).

RESULTS
A total of 4,954 men have been followed in the multicenter cohort (see Table 1).At baseline, 1,809 men were antibody positive and 3,145 were antibody negative.As of September 1991, 393 men had seroconverted and 790 had developed AIDS, including 69 of the men who seroconverted after the study began (Table 2).Seroconversion rates and AIDS rates were similar in the four centers.Different patterns of change in CD4 cell levels over time have been observed.The majority of men who seroconverted experienced a decline in their levels of CD4 cells (16,17).Few of these men experienced a return to pre-seroconversion levels of CD4 cells.Three years after baseline over half of the antibody-positive men had not experienced a significant decline in their levels of CD4 cells and less than one percent had experienced a slow steady decline in the number of CD4 cells (Table 3).Of those who had lower levels of CD4 cells after three years, the majority had experienced a shift from steady levels to a downward level suggesting that they had experienced activation of infected CD4 cells with a commensurate release of HIV-1 virions.
Further evidence of the importance of activation of cells in the natural history of HIV-1 infection was observed by correlating the levels of the markers of cell activation with risk of developing AIDS (Table 4) (12).Although levels of CD4 cells were the best predictors of AIDS, elevated levels of serum neopterin and serum B2M were also independently associated with an increased risk of AIDS.Elevations in levels of IgA and sIL-2R and the presence p24 were also predictive of AIDS, but to a lesser degree.The n all r the negative value, the greater the predictive value (e.g., -432 is more predictive than -464).</i>Estimates of incubation period Methods to combine information from the seroincident and seroprevalent subcohorts were developed by MACS investigators for the purpose of estimating the incubation period of AIDS (18.19) .Briefly, longitudinal data on seroconverters were used to identify markers of maturity of infection (i.e., variables with monotonic changes after seroconversion).Using the values of these markers measured in the seroprevalent individuals at baseline, their unknown times since seroconversion were imputed.Direct extensions of the Kaplan-Meier estimator were used in conjunction with these estimated dates of seroconversion in the seropositive cohort to estimate the survival function.Combining the seroconverter and seropositive cohort (with estimated dates of seroconversion) provided a sample large enough to estimate the hazard or incidence of AIDS by non-parametric methods.Results of the analysis indicated that, in cohorts which have been infected for more than three and one-half years, the incidence of AIDS is approximately five percent every six-month period.The estimates of the cumulative percent with AIDS by year for the first six years after seroconversion were zero percent; two percent; six percent; twelve percent, nineteen percent, and twenty-seven percent.

Factors associated with rapid progression of HIV-1 infection
A comparison of the thirty-two men who developed AIDS within five years after seroconverting with men who seroconverted but did not develop AIDS within the same time period revealed several significant differences between the two groups (Table 5) (20).Men who developed AIDS were more likely to report two or more HIV-1-related symptoms within six months of seroconversion such as fever, unintentional weight loss; diarrhea, and/or thrush.The men progressing rapidly to AIDS also had a significantly greater number of different This observation in compatible with the evidence of greater cell activation in men progressing to AIDS observed in the study of CD4 levels and of markers of cell activation reported above.Genetic profiles associated with more rapid progression to AIDS Genetic typing of peripheral blood mononuclear cells of men with rapid and men with slow declines in the levels of CD4 cells revealed an association of several HLA antigens with rapid decline"".The most significant association with rapid decline was observed with the Al-CW7-B8-DR3 haplotype suggesting a link between gene products and course of disease.Since change in CD4 levels is a very strong predictor of impending AIDS, the gene product presumably also plays a role in determining risk of developing clinical AIDS.Studies are currently underway to confirm the association with increased risk of AIDS.

Studies of use for clinical management of HIV-1 infection
Two recent studies in the MACS have provided information which will be useful for physicians responsible for managing HIV-1-infected individuals who have not yet developed AIDS or Pneunzocystis carinii pneumonia (PCP).The first of these studies looked for factors which predicted the likelihood of developing PCP in HIV-1 antibody-positive men (22).The men in this study who had less than 200 CD4 cells per cubic millimeter had a significantly higher probability of developing PCP within one to four years than men with more than 200 CD4 cells per cubic millimeter.Men who also were more likely to develop PCP had thrush or persisting fever for more than two weeks regardless of their level of CD4 cells.This study suggested that prophylactic treatment for PCP should be initiated in men with less than 200 CD4 cells per cubic millimeter and in men who have persisting fever and/or thrush.
The second of these two studies examined the impact of taking AZT and PCP prophylaxis on the risk of developing AIDS in HIV-1 antibody-positive men (23).A significant reduction in progression to AIDS within twelve, eighteen, and twenty-four months was observed in men with less than 350 CD4 cells per cubic millimeter who received AZT.A non-significant trend was also noted in those men with more 350 CD4 cells per cubic millimeter.After adjusting for effects of AZT treatment, a beneficial effect was also noted for PCP prophylactic treatment in reduction of AIDS at six, twelve, eighteen, and twenty-four months.This study suggests that early primary PCP prophylaxis is effective in preventing first episodes of PCP and that AZT is effective in delaying the onset of AIDS particularly in those men with less than 350 CD4 cells per cubic millimeter.

DISCUSSION
The studies presented above, of course, represent only a few of the many studies which are being conducted as part of the MACS.They, however, provide an example of the types of important information which are being generated by this large cohort study, the first to be developed specifically to elucidate the natural history of HIV-1 infection and AIDS.These investigations include basic immunologic and virologic studies of the alterations in the human system in response to HIV-1 and the response of HIV-1 to the human immune response, as well as studies which can be directly translated into medical management of infected individuals with and individuals with and without a diagnosis of AIDS.
Despite the many advances in the ten years since AIDS has been recognized, many questions remain to be answered before HIV/AIDS can be brought under control.We still do not know what factors determine the likelihood that an individual will develop AIDS once infected.Once we have a better understand of the biologic processes which determine, promote, and/or trigger the deterioration of the immune system leading to AIDS, we need to investigate how those processes can be altered to promote containment of HIV infection in the body and, thus, delay or prevent progression to AIDS.
The Global Program on AIDS of the World Health Organization has estimated that only one in a thousand or more sexual intercourse exposures results in successful infection by HIV.Although the presence of coexisting sexually transmitted diseases enhances the probability of infection, few exposures result in infection.Although some of this variability may be due to the actual number of virions which enter the body, the nature of the immune response of the individual and/or the presence of coexisting infection may also promote the likelihood of infection.Investigation of these possible factors may reveal alternatives to vaccination in preventing HIV infection in people at risk.
Increased survival as a result of more and better drugs will increase the incidence of other outcomes of HIV infection such as lymphomas and other malignancies.The MACS can provide information not only about what these other outcomes are, but may also provide information about the factors which are associated with their occurrence.
Although the MACS has provided much important information about the natural history of HIV/AIDS, much more needs to be done before the epidemic will be brought under control.Because of its unique nature and the outstanding commitment of its participants and staff, the MACS will continue to provide key information in the fight against HIV/AIDS.

Table 1 Participants
Enrolled in the Multicenter AIDS Cohort Study

Table 3
Patterns of Change in CD4 Levels Among Seropositive Men* <i> *This table has been adapted from Detels et al. 17</i>

Table 5
Factors Associated with Rapid Progression to AIDS* *This table has been adapted fromPhair et al .20