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Soluble markers of immune system activation

Part 1: The correlation between HIV-1 infection and ß2-microglobulin

Keeping up with the rapid progress of clinical research is essential for manufacturers seeking to create therapy-guiding AIDS assays.

David A. George

Predicting the clinical progression of disease in individuals infected with the human immunodeficiency virus-1 (HIV-1) is often difficult with the routinely used markers of disease progression.¹

Historically, CD4 T-cell lymphocyte (CD4) count has been used to monitor the clinical progression of disease, and a strong correlation exists between a low CD4 count and the development of acquired immunodeficiency syndrome (AIDS) and death.^2–6 CD4 count has also been routinely used to classify and assess the risk of HIV-infected patients. In general, patients who maintain a CD4 count of at least 500 cells/µl for a prolonged period of time (8 years or longer) are deemed long-term nonprogressors (LTNPs) and are at a reduced risk for the development of AIDS. Those with a count of 200 cells/µl or less are at increased risk and are candidates for aggressive prophylactic or antiretroviral therapy.^7–9 CD4 count, however, does not always correlate with disease stage—especially in early-stage disease—or with progression to AIDS. Frequently, patients with similar CD4 counts do not follow the same clinical course.^1,7

Three-dimensional structure of the class I major histocompatibility complex (purple), ß2-microglobulin (red), and the alpha-3 domain from human histocompatibility leukocyte antigen B2705 (other colors). Illustration Courtesy Glaxo Wellcome Experimental Research

The advent and refinement of assays for HIV-1 viral load (the number of copies of HIV-1 RNA found per milliliter of plasma or serum) have substantially improved the ability to stage HIV-infected individuals and monitor the clinical course of disease. In certain patient populations, however, HIV-1 viral load determinations have not correlated well with the course of disease. Two such populations are the LTNPs described above, which represent approximately 5–8% of HIV-infected patients, and those patients with advanced disease and very low CD4 counts.^9,10

As a result, researchers and clinicians are looking to soluble markers of immune system activation to augment the data provided by CD4 and HIV-1 viral load determinations. They hope to find additional prognostic data by which to determine the best course of treatment for an individual—for example, whether or not the patient should begin receiving antiretroviral therapeutic agents. Three immune system activation markers under consideration are ß2-microglobulin (ß2M), neopterin, and tumor necrosis factor receptor type II (TNFR-II). ß2M is the light chain moiety of the class I histocompatibility leukocyte antigen (HLA) complex; neopterin is a product of guanosine triphosphate catabolism and is primarily accounted for in serum by interferon--stimulated macrophages; and TNFR-II is a cell-membrane-bound receptor specific for the cytokines TNF- and lymphotoxin- and is released into circulation upon proteolytic cleavage of its extracellular component. In many of the studies discussed here the data provided by ß2M, neopterin, and TNFR-II were of approximately equivalent clinical value, and although neopterin and TNFR-II deserve consideration and will be briefly discussed, the remainder of this article will focus on ß2M, for which clinical immunoassays are currently available.

ß2M belongs to the ß-globulin family of human plasma proteins. It is bound noncovalently to the heavy chain subunit of the class I HLA complex, and is found on the cell surface of all nucleated cells.¹¹ Overproduction of the HLA complex can cause ß2M to be released into circulation. The lymphocytes are largely responsible for the overproduction of this solubilized ß2M, which is normally filtered out of serum through renal glomeruli and ultimately reabsorbed and catabolized by epithelial cells. Serum elevations of ß2M occur, however, in several clinical conditions such as multiple myeloma, nasopharyngeal carcinoma, and a variety of lymphoproliferative disorders.^12–15 In addition, ß2M has been studied extensively as a marker of immune system activation in HIV-infected individuals in an effort to identify and monitor those patients at high risk for progressing to AIDS.^16–18

ß2M Correlation with HIV-1 Infection

The association between ß2M and HIV-1 infection is well documented, with ß2M serum levels rising in correlation with disease progression and reaching a peak just before death.^18–22 Some early investigations suggested a prognostic role for ß2M, as patients with higher ß2M serum levels tended to develop AIDS sooner than patients with lower levels.^23,24 Recent studies have confirmed the strong correlation among ß2M, CD4 count, and HIV-1 infection (see Table I).²⁵

However, conflicting results have been reported regarding the prognostic value of ß2M, that is, whether the determination of ß2M serum levels early in the course of HIV-1 infection can be used to predict the time-course of progression to AIDS or death. A recent prospective investigation involving 34 asymptomatic HIV-infected individuals evaluated the prognostic value of ß2M and several viral markers, including HIV-1 viral load.²⁶ For each subject, all markers were measured upon study entry and every eight weeks thereafter for a period of three years. Those subjects who progressed to AIDS or to an HIV-related disease were placed into the progressor (P) group, and those who remained asymptomatic were placed into the nonprogressor (NP) group. Table II shows that at study entry, ß2M serum levels were higher in the P group than in the NP group, but the difference was not significant enough to predict clinical progression in asymptomatic individuals. Only HIV-1 viral load at study entry showed a significant correlation between the NP and P groups (p <0.003, two-sided Mann-Whitney test).

The values reported for ß2M in this study are discrepant from those reported in Table I, in which the ß2M serum levels of all HIV-infected groups were significantly elevated above normal. Further investigation would likely reveal factors that might have contributed to this discrepancy, such as the small size of the recent study's population, differences in data interpretation, or differences in the ß2M assay methodology (e.g., selection of an appropriate upper reference limit).

A similar but retrospective study evaluated HIV-1 viral load, ß2M, neopterin, and TNFR-II in HIV-infected individuals who were matched by baseline CD4 count, rate of CD4-count decline, and other demographic factors such as age and ethnic origin.¹ The 90 study subjects were categorized into one of three groups—slow, moderate, and rapid progressors—and were evaluated for up to a 10-year period. Samples were obtained and assayed upon study entry (baseline), at six months, and again at the time closest to AIDS onset, death, or at the end of the follow-up period; a total of three samples were evaluated from each subject. For slow and moderate progressors, the third sample was obtained at a visit chronologically closest to that of the clinical progression in the rapid progressor group. At baseline, levels of all markers increased incrementally from the slowest to the most rapid progressor groups, but only HIV-1 viral load and TNFR-II showed a statistically significant difference between each group (see Table III). In addition, univariate and multivariate matched conditional regression models of the baseline data indicated that only HIV-1 RNA and TNFR-II were significant predictors of progression to AIDS in the rapid progressor group.

It is interesting to note that in both of the studies discussed above, ß2M tended to correlate with disease progression, although the increases were not statistically significant. In the prospective study, ß2M was more elevated in the progressor group than in the nonprogressor group. In the retrospective study, in which HIV-1 viral load was significantly predictive of clinical disease progression, ß2M increased across all three progressor groups and correlated significantly with HIV-1 viral load. The authors of the latter study offered no hypothesis to explain why ß2M levels would correlate with HIV-1 viral load but lack statistical significance as a predictor of disease progression. However, it could be that the preclassification of their study subjects by CD4 count and CD4-count decline biased the interpretation of ß2M measurements.

Conclusion

The studies discussed here reveal a distinct correlation between HIV-1 infection and elevated serum levels of ß2M. The question remains, however, just how ß2M and the other markers of immune system activation can best be used in monitoring disease progression or in predicting the prognosis of HIV-infected individuals. These and other issues will be addressed in the second installment of this article, which appears in the March/April 1999 issue of IVD Technology.

References

1. DS Stein et al., "Predicting Clinical Progression or Death in Subjects with Early-Stage Human Immunodeficiency Virus (HIV) Infection: A Comparative Analysis of Quantification of HIV RNA, Soluble Tumor Necrosis Factor Type II Receptors, Neopterin, and ß2-Microglobulin," Journal of Infectious Disease 176 (1997): 1161–1167.

2. AR Moss et al., "Serum Positivity for HIV and the Development of AIDS or AIDS-Related Condition: Three-Year Follow-Up of the San Francisco General Hospital Cohort," British Medical Journal 296 (1988): 745–750.

3. AN Phillips et al., "Serial CD4 Lymphocyte Counts and Development of AIDS," Lancet 337 (1991): 389–392.

4. AJ Mocroft et al., "Staging System for Clinical AIDS Patients," Lancet 346 (1995): 12–17.

5. GD Mills and PL Jones, "Relationship between CD4 Lymphocyte Count and AIDS Mortality, 1986–1991," AIDS 7 (1995): 1383–1386.

6. JM Colford, L Ngo, and I Tager, "Factors Associated with Survival in Human Immunodeficiency Virus-Infected Patients with Very Low CD4 Counts," American Journal of Epidemiology 139 (1994): 206–218.

7. JJ Lefrére et al., "Even Individuals Considered as Long-Term Nonprogressors Show Biological Signs of Progression after 10 Years of Human Immunodeficiency Virus Infection," Blood 90, no. 3 (1997): 1133–1140.

8. AJ Mocroft et al., "The Relationship between ß2-Microglobulin, CD4 Lymphocyte Count, AIDS, and Death in HIV-Positive Individuals," Epidemiology and Infection 118 (1997): 259–266.

9. L Ashton et al., "Predictors of Progression in Long-Term Nonprogressors," AIDS Research and Human Retroviruses 14, no. 2 (1998): 117–121.

10. R Zangerle et al., "Serum HIV-1 RNA Levels Compared to Soluble Markers of Immune Activation to Predict Disease Progression in HIV-1-Infected Individuals," International Archives of Allergy and Immunology 116 (1998): 228–239.

11. P Cresswell et al., "Immunological Identity of the Small Subunit of HL-A Antigens and ß2-Microglobulin and Its Turnover on the Cell Membrane," Proceedings of the National Academy of Science USA 71 (1974): 2123–2127.

12. G Brenning, L Wibell, and R Bergstrom, "Serum ß2-Microglobulin at Remission and Relapse in Patients with Multiple Myeloma," European Journal of Clinical Investigation 15 (1985): 242–247.

13. W Shiu et al., "Expression of ß2-Microglobulin by Nasopharyngeal Carcinoma," British Journal of Cancer 66, no. 3 (September 1992): 555–557.

14. JA Child and MRS Kushwaha, "Serum ß2-Microglobulin in Lymphoproliferative and Myeloproliferative Diseases," Hematological Oncology 2 (1984): 391–401.

15. S DiGiovanni et al., "ß2-Microglobulin Is a Reliable Tumor Marker in Chronic Lymphocytic Leukemia," Acta Haematologica 81 (1989): 181–185.

16. JL Fahey et al., "The Prognostic Value of Cellular and Serologic Markers in Infection with Human Immunodeficiency Virus Type I," New England Journal of Medicine 322 (1990): 166–172.

17. RB Bhalia et al., "Abnormally High Concentrations of ß2-Microglobulin in AIDS Patients," Clinical Chemistry 29 (1983): 1560.

18. B Hofmann et al., "Serum ß2-Microglobulin Level Increases in HIV Infection: Relation to Seroconversion, CD4 T-Cell Fall, and Prognosis," AIDS 4, (1990): 207–214.

19. NA Harrison and SJ Skidmore, "Neopterin and ß2-Microglobulin Levels in Asymptomatic HIV Infection," Journal of Medical Virology 32, no. 2 (1990): 128–133.

20. G Cavalli et al., "Diagnostic and Prognostic Significance of ß2-Microglobulin during HIV Infection," Ricerca in Clinica e Laboratorio 20, no. 2 (1990): 105–111.

21. GA Garden et al., "ß2-Microglobulin as a Marker of HIV Disease Status in Nairobi, Kenya," International Journal of STD and AIDS 4, no. 1 (1993): 49–51.

22. M Radkowski et al., "Prospective Studies of T4 and T8 Lymphocyte Subpopulations and ß2-Microglobulin in Patients with HIV Infection," Polskie Archiwum Medycyny Wewnetrznej 86, no. 4 (1991): 247–253.

23. AN Phillips et al., "Serum ß2-Microglobulin at HIV-1 Seroconversion as a Predictor of Severe Immunodeficiency during 10 Years of Follow-Up," Journal of Acquired Immune Deficiency Syndromes 13, no. 3 (1996): 262–266.

24. M Thakar et al., "Serum ß2-Microglobulin Levels in HIV Seropositive Persons," Indian Journal of Medical Research 95 (1992): 168–170.

25. V Henne, P Frei, and P Bürgisser, "ß2-Microglobulin—A Rapid and Automated Determination for a Broad Range of Clinical Applications," Anticancer Research 17 (1997): 2915–2918.

26. SM Bruisten et al., "Prospective Longitudinal Analysis of Viral Load and Surrogate Markers in Relation to Clinical Progression in HIV Type I-Infected Persons," AIDS Research and Human Retroviruses 13, no. 4 (1997): 327–335.

David A. George is a product manager at Scripps Laboratories (San Diego).

Continue to part 2 of this article.