To assess the potential impact of vaccine on plasma viral RNA and proviral DNA, subjects were stratified at entry according to treatment with antiretroviral compounds (for number of subjects taking or not taking antiretrovirals see Table 1). As shown in Fig. 3, there was no significant change over time in either HIV RNA or proviral DNA copy numbers in volunteers taking or not taking antiretroviral compounds. Among the volunteers taking antiretroviral compounds, there was a trend towards increasing proviral DNA (and higher plasma RNA on one occasion) with the placebo group. However, this did not achieve statistical significance. Vaccination by itself did not increase viral RNA or DNA (assessed 4 weeks after administration).
Although this study was not designed as a phase III efficacy study, sufficient AIDS-defining events occurred in group B (CD4 cell count 200-500¥106 cells/l) relative to group A (CD4 cell count >500¥106 cells/l) to be significant after 24 months (data not shown). However, no impact of vaccine could be determined in this time, nor was there a trend. Among the 103 vaccinated volunteers, there were 19 with one or more AIDS-defining events and among the 105 placebo recipients, there were 18 with one or more AIDS-defining events (c2, 0.06, P=0.86). Results of time-to-event analyses were similar (data not shown).
Similarly, when stratified by baseline CD4 cell count, no effect of vaccine on CD4 cell count decline was observed. Within 24 months 132 of 208 subjects had experienced a decline in CD4 cell count of ≥33% from baseline or to <200¥106 cells/l. Among group A, 58 (32 vaccine versus 26 placebo), and in group B 74 (34 vaccine versus 40 placebo) developed this immunologic deterioration. There was no statistically significant difference between vaccine and placebo recipients in the probability of CD4 decline in either group A or group B (Fig. 4a). When these subjects were stratified according to viral RNA or proviral DNA at baseline, a significant difference in probability of CD4 decline was observed. However, no significant difference between vaccine and placebo recipients was found in these analyses (Fig. 4b and 4c).
Despite induction of new lymphoproliferative responses by vaccination with rgp160 IIIB and rgp160 MN, no clinical benefit was found in this trial. This was true for subjects with relatively intact immune systems as shown by baseline CD4 cell counts of >500¥106 CD4 cells/l, as well as subjects with moderate CD4 cell decline (200-500¥106 CD4 cells/l) at study entry. The results of the trial also imply, therefore, that CD4 cell count as the only marker for patient selection is not sufficient in future vaccination trials as evidenced by a comparatively high plasma viral RNA and by a considerable proportion of patients progressing to AIDS even in group A. Clinical outcome was predicted by either viral RNA or proviral DNA at baseline as has been shown by others [21-25]. When subjects were stratified into high or low plasma RNA or proviral DNA baseline levels, vaccine had no effect on CD4 cell count (see Fig. 4) or clinical events (data not shown). Although there was no benefit from vaccine, neither was there an adverse effect. The vaccine was safe and well tolerated, and use of vaccine was not associated with any change in CD4 cell decline, viral RNA in plasma or proviral DNA.
Previous studies of vaccines in infected persons have included phase I studies of this and other gp160 preparations [14,26-31], gp120 , HIV-1 core proteins [33-35], whole inactivated HIV-1 [36,37], live vector vaccine including canarypox  and polynucleotide vaccine . Larger studies to evaluate some of these vaccines for efficacy are currently ongoing, but to date the results on efficacy have not been reported. Although the present trial was not designed as a phase III efficacy trial, the long duration of follow-up (2 years) in both vaccine and placebo groups resulted in sufficient AIDS-defining events to suggest that this type of vaccine was not highly efficacious.
New immune responses to HIV antigens among infected persons who have received an HIV vaccine have included the induction of lymphocyte proliferative responses to HIV Env [27-31], as found in the present study. Although lymphocyte proliferation responses to Env would be an expected response to HIV-1-infection, this is not usually the case. Vaccination clearly is adding this type of immune response to the infecting agent. In previous studies, gp160 has been a much more powerful inducer of lymphoproliferation than has gp120  and the vaccine used in this study was shown to have significant immunogenicity in uninfected persons, particularly with regard to lymphocyte proliferation [10,11,13]. Recent observations suggest that HIV-1-specific lymphoproliferative responses develop after treatment of acutely infected persons with potent antiretroviral drugs . This has lead to speculation on the importance of HIV-1-specific T helper cell responses in controlling HIV infection. However, despite these immune responses that were induced in our study, the clinical outcome observed to date does not support this hypothesis.
Identification of immune parameters that correlate with slowing disease progression may refocus approaches on therapeutic vaccine in HIV. High levels of neutralizing antibodies, high frequency cytotoxic T-cell precursors, or other parameters may emerge as correlates of slow disease progression. Therapeutic vaccine to induce one or more of these parameters in conjunction with chemotherapy may provide significant immune control over the virus. Future trials should examine the effect of vaccine on persons who were vigorously treated with chemotherapy and responded with reduction in viral load.
1. Girard M, Kieny M-P, Pinter A, et al. Immunization of chimpanzees confers protection against challenge with human immunodeficiency virus. Proc Natl Acad Sci USA
2. Berman PW, Murthy KK, Wrin T, et al. Protection of MN-rgp120-immunized chimpanzees from heterologous infection with a primary isolate of human immunodeficiency virus type 1. J Infect Dis
3. Hu S-L, Abrams K, Barber GN, et al. Protection of macaques against SIV infection by subunit vaccines of SIV envelope glycoprotein gp160. Science
4. Hu S-L, Polacino P, Stallard V, et al. Recombinant subunit vaccines as an approach to study correlates of protection against primate lentivirus infection. Immunol Lett
5. Mossman SP, Bex F, Berglund P, et al. Protection against lethal simian immunodeficiency virus SIVsmmPBj14 disease by a recombinant Semliki Forest Virus gp160 vaccine and by a gp120 subunit vaccine. J Virol
6. Lutz H, Hofmann-Lehmann R, Leutenegger C, et al. Vaccination of cats with recombinant envelope glycoprotein of feline immunodeficiency virus: decreased viral load after challenge infection. AIDS Res Hum Retroviruses
7. Hosie MJ, Dunsford TH, de Ronde A, et al
. Suppression of virus burden by immunization with feline immunodeficiency virus Env protein. Vaccine
8. Mannhalter JW. Acquired immunodeficiency syndrome vaccines: current concepts and future prospects.
In Symposium in Immunology VII. Vaccination.
Edited by Eibl MM, Huber C, Peter HH, Wahn U. Berlin Heidelberg: Springer Verlag; 1998:137-153.
9. Dolin R. Human studies in the development of human immunodeficiency virus vaccines. J Infect Dis
10. Belshe RB, Clements ML, Dolin R, et al. Safety and immunogenicity of a fully glycosylated recombinant gp160 human immunodeficiency virus type 1 vaccine in subjects at low risk of infection. J Infect Dis
11. Gorse GJ, Schwartz DH, Graham BS, et al. HIV-1 recombinant gp160 vaccine given in accelerated dose schedules. Clin Exp Immunol
12. Gorse GJ, Rogers JH, Perry JE, et al. HIV-1 recombinant gp160 vaccine induced antibodies in serum and saliva. Vaccine
13. Gorse GJ, McElrath MJ, Matthews TJ, et al. Modulation of immunologic responses to HIV-1MN recombinant gp160 vaccine by dose and schedule of administration. Vaccine
14. Schwartz D, Clements ML, Belshe R, et al. Interim results of rgp160 vaccine trial in HIV + volunteers. IX International Conference on AIDS/IV STD World Congress.
Berlin, June 1993 [abstract PO-A28-0668].
15. Barrett N, Mitterer A, Mundt W, et al. Large-scale production and purificaiton of a vaccinia recombinant-derived HIV-1 gp160 and analysis of its immunogenicity. AIDS Res Hum Retroviruses
16. Piatak M, Luk K-C, Williams B, Lifson JD. Quantitative competitive polymerase chain reaction for accurate quantitation of HIV DNA and RNA species. BioTechniques
17. Hämmerle T, Himmelspach M, Dorner F, Falkner FG. A sensitive PCR assay system for quantitation of viral genome equivalents: Human immunodeficiency virus type 1 (HIV-1) and hepatitis B virus (HBV). Arch Virol
18. Bøyum A. Isolation of mononuclear cells and granulocytes from human blood. Scand J Clin Lab Invest
19. Mannhalter JW, Pum M, Wolf HM, et al. Immunization of chimpanzees with the HIV-1 glycoprotein gp160 induces long-lasting T-cell memory. AIDS Res Hum Retroviruses
20. AIDS Vaccine Evaluation Group: Protocol 13. 1995
21. Mellors JW, Rinaldo Jr CR, Gupta P, White RM, Todd JA, Kingsley LA. Prognosis in HIV-1 infection predicted by the quantity of virus in plasma. Science
22. O‚Brien TR, Blattner WA, Waters D, et al. Serum HIV-1 RNA levels and time to development of AIDS in the multicenter hemophilia cohort study. JAMA
23. Phillips AN, Eron JJ, Bartlett JA, et al. HIV-1 RNA levels and the development of clinical disease. AIDS
24. Mellors JW, Muños A, Giorgi JV, et al. Plasma viral load and CD4+ lymphocytes as prognostic markers of HIV-1 infection. Ann Intern Med
25. Chevret S, Kirstetter M, Mariotti M, Lefrère F, Frottier J, Lefrère J-J. Provirus copy number to predict disease progression in asymptomatic human immunodeficiency virus type 1 infection. J Infect Dis
26. Redfield RR, Birx DL, Ketter N, et al. A phase I evaluation of the safety and immunogenicity of vaccination with recombinant gp160 in patients with early human immunodeficiency virus infection. N Engl J Med
27. Loomis LD, Deal CD, Kersey KS, Burke DS, Redfield RR, Birx DL. Humoral responses to linear epitopes on the HIV-1 envelope in seropositive volunteers after vaccine therapy with rgp160. J Acquir Immune Defic Syndr Hum Retrovirol
28. Valentine FT, Kundu S, Haslett PAJ, et al. A randomized, placebo-controlled study of the immunogenicity of human immunodeficiency virus (HIV) rgp160 vaccine in HIV-infected subjects with ≥400/mm3 CD4 T lymphocytes (AIDS Clinical Trials Group Protocol 137). J Infect Dis
29. Kundu SK, Katzenstein D, Valentine FT, Spino C, Efron B, Merigan TC. Effect of therapeutic immunization with recombinant gp160 HIV-1 vaccine on HIV-1 proviral DNA and plasma RNA: relationship to cellular immune responses. J Acquir Immune Defic Syndr Hum Retrovirol
30. Leandersson A-C, Bratt G, Hinkula J, et al. Induction of specific T-cell responses in HIV infection. AIDS
31. Pontesilli O, Guerra EC, Ammassari A, et al. Phase II controlled trial of post-exposure immunization with recombinant gp160 versus antiretroviral therapy in asymptomatic HIV-1-infected adults. AIDS
32. Eron JJ, Ashby MA, Giordano MF, et al. Randomised trial of MNrgp120 HIV-1 vaccine in symptomless HIV-1 infection. Lancet
33. Klein MR, Veenstra J, Holwerda AM. et al. Gag-specific immune responses after immunization with p17/p24:Ty virus-like particles in HIV type 1-seropositive individuals. AIDS Res Hum Retroviruses
34. Peters BS, Cheingsong-Popov R, Callow D, et al. A pilot phase II study of the safety and immunogenicity of HIV p17/p24:VLP (p24-VLP) in asymptomatic HIV seropositive subjects. J Infection
35. Kelleher AD, Roggensack M, Jaramillo AB, et al. Safety and immunogenicity of a candidate therapeutic vaccine, p24 virus-like particle, combined with zidovudine, in asymptomatic subjects. AIDS
36. Trauger RJ, Ferre F, Daigle AE, et al. Effect of immunization with inactivated gp120-depleted human immunodeficiency virus type 1 (HIV-1) immunogen on HIV-1 immunity, viral DNA, and percentage of CD4 cells. J Infect Dis
37. Moss RB, Giermakowska W, Lanza P, et al. Cross-clade immune responses after immunization with a whole-killed gp120-depleted human immunodeficiency virus type-1 immunogen in incomplete Freund‚s adjuvant (HIV-1 immunogen, REMUNE) in human immunodeficiency virus type-1 seropositive subjects. Virol Immunol
38. Tubiana R, Gomard E, Fleury H, et al. Vaccine therapy in early HIV-1 infection using a recombinant canarypox virus expressing gp160MN (ALVAC-HIV): a double-blind controlled randomized study of safety and immunogenicity [letter]. AIDS
39. Calarota S, Bratt G, Nordlund S, Hinkula J, Leandersson AC, Sandström E. Cellular cytotoxic response induced by DNA vaccination in HIV-1-infected patients. Lancet
40. Rosenberg ES, Billingsley JM, Caliendo AM, et al. Vigorous HIV-1-specific CD4(+) T cell responses associated with control of viremia. Science
E. Tschachler, Universiäts Hautklinik, Vienna, Austria; B. Colebunders, Institute of Tropical Medicine, Antwerpen, Belgium; F. Black, Marselisborg Hospital, Aarhus, Denmark; A. Ranki, Deptartment of Dermatology and Venereal Diseases, Helsinki, Finland; J. Rockstroh, Department of Medicine, University of Bonn, Germany; R. Zimmermann, Kurpfalzkrankenhaus Heidelberg, Germany; I. Scharrer, University Hospital, Frankfurt, Germany; J. R. Bogner, Medizinische Poliklinik, University of Munich, Germany; C. F. Mantel, Landesinstitut für Tropenmedizin, Berlin, Germany; H. Jablonowski, Medizinische Universitätsklinik, Düsseldorf, Germany; W. Schramm, F. Rommel, Medizinische Klinik, University of Munich, Germany; W. Brockhaus, Zentrum innere Medizin, Klinikum der Stadt Nürnberg, Germany; S. S. Fröland, Rikshospitalet, Oslo, Norway; E. Sandström, Södersjukhuset, Stockholm, Sweden; E. Berntorp, Malmö General Hospital, Malmö, Sweden; M. Flepp, Universitätsspital Zürich, Switzerland; L. Stigendal, Sahlgrenska Sjukhuset, Göteborg, Sweden.
K. Krohn, V. Blazevic, A. Lagerstedt, Institute of Medical Technology, University of Tampere, Finland; B. Wahren, G. Gilliam, A. C. Leanderson, Karolinska Institut, Stockholm, Sweden; J. D. Lifson, M. Piatak, Genelabs Technologies Inc. Redwood City, CA, USA; B. Husch, L. Trawnicek, N. Barrett, T. Hämmerle, F. Dorner, G. Eder, IMMUNO AG, Vienna, Austria.