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Vaccination of Vaccinia-Naive Adults with Human Immunodeficiency Virus Type 1 gp160 Recombinant Vaccinia Virus in a Blinded, Controlled, Randomized Clinical Trial Vanderbilt University School o] Medicine. Nashville. Tennessee: St. Louis Universitv School of Medicine. St. Louis. Missouri: Universit v o]' Rochester School of Medicine and Dentistrv. Rochester. New York: Bristol-Myers Squibb Pharmaceutical Research Institute and University of Washington School (1' Medicine. Seattle; Johns Hopkins School of Hygiene and Public Health and School of Medicine. Baltimore. Georgetown University. Rockville. EMMES Corporation. Potomac. and National Institutes of Health, Bethesda. Maryland
The safety and immunogenicity of a human immunodeficiency virus type 1 (HIV-l) gp160 recombinant vaccinia virus (HIVAC-le) vaccine was evaluated in vaccinia-naive, healthy adults at low risk for acquiring HIV-l infection. Volunteers (n = 36) were randomized to receive HIVAC-l e or control vaccinia virus at two dosages by bifurcated needle puncture at 0 and 2 months; 12 HIVAC-le and 6 control vaccinia virus recipients received either 106 or 107 pfujmL at each inoculation. There was no significant difference in lesion size, level of viral replication, or systemic symptoms after vaccination with HIVAC-le or control vaccinia virus. Of 22 HIVAC-le recipients with lesion formation, 16 developed low-titer gp 160-specific antibody responses detectable by Western blot. The peak response occurred between days 70 and 120 and was still detectable at day 365 in 9 of 18 vaccinees. gp 160-specific Iymphoproliferative responses were detected in 5 of 10 vaccinees. Vaccination with HIVAC-le was safe in vaccinia-naive, healthy adults and could induce both humoral and cell-mediated gp 160-specific immune responses.
The precedent for using vaccinia virus in a successful, worldwide immunization program makes it an attractive vehicle for presentation of novel antigens. Therefore, the capability to engineer vaccinia virus to express foreign genes [I, 2] was an important breakthrough in vaccine development. Recombinant vaccinia viruses expressing the appropriate foreign genes have since been shown to provide protective immunity against experimental challenge with influenza virus in hamsters [3], rabies virus in rabbits and mice [4],
Received 29 January 1992: revised 12 March 1992. Presented in part: 3rd annual meeting of the National Cooperative Vaccine Development Groups for AIDS. Clearwater Beach. Florida. October 1990; VII International Conference on AIDS. Florence. Italy. June 1991 (abstract F.A.I). All volunteers underwent an extensive informed consent process and were required to comprehend fully the purpose and details of the study. All protocols and consent forms were approved by local Institutional Review Boards and Institutional Biosafety Committees. The study was conducted according to the human experimentation guidelines of the US Department of Health and Human Services. Grant support: National Institutes of Health (AI-82500. -05061. -05062. -05063. -05064. -05065. -15106). Reprints or correspondence: Dr. Barney S. Graham. Department ofMedicine. Division of Infectious Diseases. A-331 0 MCN. Vanderbilt University School of Medicine. Nashville. TN 37232.
The Journal of Infectious Diseases 1992;166:244-52 © 1992 by The University of Chicago, All rights reserved. 0022-1899/92/6602-0003$01.00
herpes simplex virus in mice [5], vesicular stomatitis virus in mice and cattle [6], rinderpest virus in cattle [7], hepatitis B virus in chimpanzees [8], and simian AIDS caused by a type D retrovirus in macaques [9]. Vaccinia recombinant vectors can induce both humoral antibody responses and cellular immune responses specific to the inserted gene product(s). Induction of cytotoxic T lymphocytes (CTL) is thought to be particularly important in containing human immunodeficiency virus type I (HIYI) infection. Expression of the influenza hemagglutinin by recombinant vaccinia virus vectors has been shown to induce hemagglutinin-specific major histocompatibility complex (MHC) class I-restricted CTL in inbred mice [10]. Simian immunodeficiency virus (SIV) genes expressed in vaccinia recombinant viruses have been shown to induce Sl V-specific MHC class I-restricted CTL in monkeys [II, 12]. The envelope gene of HIV-l, expressed in vaccinia, has induced gp 160-specific antibody in mice and rhesus monkeys [13, 14]. Vaccination of chimpanzees with vaccinia recombinant virus expressinggp 160 induced HIV-specific lyrnphoproliferative responses and MHC class II-restricted CD4+ CTL [15]. Although there is extensive clinical experience with native vaccinia, there is limited experience with recombinant vaccinia viruses in humans in terms of either clinical behavior of the recombinant viruses or their ability to effectively express
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Barney S. Graham, Robert B. Belshe, Mary Lou Clements, Raphael Dolin, Lawrence Corey, Peter F. Wright, Geoffrey J. Gorse, Karen Midthun, Michael C. Keefer, Norbert J. Roberts, Jr., David H. Schwartz, Jan M. Agosti, Bruce F. Fernie, Donald M. Stablein, David C. Montefiori, John S. Lambert, Shiu-Lok Hu, Joy R. Esterlitz, Dale N. Lawrence, Wayne C. Koff, and the AIDS Vaccine Clinical Trials Network
JID 1992; 166 (August)
gpl60 Recombinant Vaccinia Vaccine
Materials and Methods Vaccine Product Preparation HIV AC-l e (Bristol-Myers Squibb, Seattle) is the product designation for the vaccinia recombinant containing the HIV-1 envelope gene [14]. The parent vaccinia virus was derived by plaque purification from a commercial preparation of smallpox vaccine (Dryvax, lot 321501 G; Wyeth Laboratories, Philadelphia) grown in BSC-40 cells (derived from BSC-l, ATCC CCL26). The master seed lot of the recombinant virus was then prepared in human diploid MRC-5 cells (ATCC CeL-171). The vaccine virus was prepared in MRC-5 cells by inoculation with master seed virus. The infected cell monolayer was washed twice with PBS; cells were scraped and pelleted by centrifugation at 1500 g. resuspended in PBS, and then ruptured by two cycles offreeze-thaw and sonication. After shipment and storage at -70°C, the resulting suspension had a titer of --2 X 108 pfu/ml, as determined at inoculation. A single lot ofcommercial native vaccinia product (Dryvax; Wyeth) prepared as lyophilized calflymph was reconstituted before use with a diluent containing 50%glycerin. The titer of the control vaccinia virus suspension was -- 2 X 108 pfu/rnl..
Subjects Inclusion criteria were that the subjects be healthy HIV-seronegative adults 18-60 years old with negligible risk for HIV-l exposure and with no evidence of prior smallpox vaccination. Volunteers were excluded if there were any indications of highrisk behavior for HIV-l infection (determined by history, questionnaire, and laboratory evaluation). Volunteers were excluded if there was any evidence of chronic illness or if they or their
Table 1. Clinical and virologic characteristics of vaccinia-naive adults vaccinated with different doses of human immunodeficiency virus type I gp 160 recombinant vaccinia virus (HIVAC-I e) or control vaccinia virus. Dosage 107 pfu/rnl.
106 pfu/ml.
Subjects Men Women Total Heterosex ual Homosexual Age (mean years) No. with lesion formation after Primary vaccination Secondary vaccination Total with lesion formation No lesion formation Maximum skin titer Day of maximum titer Last day of shedding
HIVAC-Ie
Control
HIVAC-Ie
Control
9 3 12 10 2 20.0
5 I 6 4 2 20.5
9 3 12 8 4 23.5
4 2 6 4 2 21.5
5
4
11*
6
5
2
10 2t 5.6 ± 0.4
6 0 6.1 ± 0.6
12 0* 5.0 ± 0.4
6 0 5.5±0.7
II (6-18)
10 (5-14)
8 (4-20)
14 (5-21)
21(14-22)
19 (9-21)
18 (6-26)
18 (9-21)
P
NOTE. Maximum skin titers are given as mean loglo pfu/rnl, ± SE. Days of maximum titer and shedding are given as median (range). * Two volunteers were vaccinated with stocks stored at - 35°C for> 12 months: virus titers in inocula were 1000-fold in titer from inside to outside (mean 10gIO pfu/ml. inside was 7.0 vs. 3.4 outside). Genetic stability of HIVAC-le. The last samples from which virus could be isolated in 5 HIV AC-I e recipients (days 8, 15, 15, 18, and 20) and one Dryvax recipient were evaluated for gp 160 expression. None of the Dryvax plaques (>500) had evidence ofgpl60 expression. Of> 1000 individual plaques from HIVAC-I e recipients, all showed gp 160 expression. Vaccinia-specific antibody responses to vaccination. ELISA (figure I) and plaque-reduction neutralization tests showed that vaccinia-specific antibody could not be detected until after an inoculation that resulted in lesion formation. The level of antibody was not augmented further in persons who had lesion formation after initial vaccination and then received a booster vaccination. HI V-specific antibody responses to vaccination. The antibody responses to vaccination were recorded from the time of the inoculation that resulted in lesion formation. Western blot assays were used to detect antibody responses to the HIV-I envelope glycoproteins gp 160, gp 120, or gp41. In 8 of 10 HIVAC-Ie 106-dose recipients and 8 of 12 HIVAC-Ie 107-dose recipients, antibody was detected. In 4 106-dose HIVAC-I e recipients, antibody to HIV-I envelope bands did
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not develop until day 120 after lesion formation, and in 3 of those 4 recipients, antibody persisted past day 365. Overall, 9 of 18 HIV AC-l e recipients had gp l60-specific antibody detectable by Western blot beyond day 365. The earliest development of antibody to HIV-I envelope bands was day 28. The responses were generally weak, with serum from only 1 volunteer showing a positive intensity against envelope bands on Western blot at one time point (figure 4). The delayed development ofantibody made interpreting the HIV antibody response to the second vaccination difficult. However, antibody did not appear to be augmented after revaccination. Evaluation of antibody responses by ELISA identified only 1 volunteer at one time point as positive, with a ratio of test sample 00 to cutoffof2.35. This serum, from day 210 after vaccination, was the same sample that tested strongly
positive by Western blot. Neutralizing antibody activity against the homologous HIV -1 IIIB strain was detected only in the day 180 serum from the individual with the strong positive Western blot and positive ELISA. Cell-mediated immune responses to vaccination. Lymphoproliferative responses specific for HIV-I gp 160 and responses induced by mitogen and another recall antigen were measured. Lymphoproliferative responses to phytohemagglutinin and Candida antigen remained stable beyond day 365. No control vaccinia virus recipients developed a significant Iymphoproliferative response (SI >5.0) to rgp 160, whereas 5 of 10 HIVAC-Ie recipients did (figure 5). The peak number of responders (5/1 0) occurred on day 120 after vaccination. Only I of 6 still had significant rgp 160 responses on day 365.
Discussion flit
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DAYS AFTER PRIMARY VACCINATION Figure 3. Absolute CD4 lymphocyte counts throughout followup after vaccination. There was no deleterious effect on CD4 cell numbers as a result of gp 160 expression.
The induction of immunity through presentation of antigen by live recombinant vectors is an important vaccination strategy. Vaccinia is among a number of viral and bacterial recombinant vaccine vectors in development; it offers many advantages as a vaccine vehicle including low cost, heat stability, and documentation of vaccination via a scar. Its ease of administration in developing nations is attested by its use in the global eradication of smallpox [22, 25]. However, there are drawbacks to consider. Spread of vaccinia to contacts is well recognized [26]. Transmission to healthy individuals is a concern, especially with regard to ocular infection and unanticipated acquisition of seropositivity to the foreign gene product. The greater concern, however, is accidental transmission to people with eczema or immunodeficiency [27]. Vaccinia virus has been reported to have caused a severe local reaction and dissemination in an asymptomatic but severely immunodeficient HIV-I-infected person [28]. This has implications for the
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DA YS AFTER PRIMARY VACCINATION Figure 4. Human immunodeficiency virus type I (HIV-I) envelope-specific Western blot responses in volunteers after vaccinations that led to lesion formation. Results are aligned from time of inoculation that resulted in lesion formation. Band intensity was graded according to kit directions as absent (-); present, but less intense than anti-p24 band on weakly reactive control strip (± or visible); at least as intense as anti-p24 band on weakly reactive control strip, but less intense than anti-p24 on strongly reactive control strip (+ or positive); or greater than or equal to intensity of anti-p24 on strongly reactive control strip (++ or strong positive). Percentage of volunteers with responses represented by bars with absolute number of positive responses per number tested recorded above bar.
mended for virus containment in large-scale trials or for routine vaccination. Nevertheless, for small phase I or II trials in which study personnel can change the dressing, the transparent dressing is an effective barrier. The thymidine kinase (TK) gene, which is interrupted in the HIV AC-l e product, is considered to be a determinant of virulence. TK+ vaccinia is more virulent than TK- vaccinia in mice when the inoculation is given parenterally [31] or into the lung [32]. It was therefore anticipated that HIV ACl e would be less virulent than control vaccinia virus. Analysis of viral replication, size and character of skin lesions, and serologic responses after vaccination, however, showed no differences between the HIVAC-Ie and control vaccinia virus products in this trial. Although there was no evidence of HIV AC-l e attenuation by these measures of virulence, HIV AC-I e was derived from a plaque-purified vaccinia stock that may not accurately reflect the composite effects of minor strain variants in the control vaccinia virus product. Also, in mice inoculated by scarification, there is no apparent attenuation of TK- vaccinia as measured by mortality, viral dissemination, or serologic responses [31], so dermal inoculation may not be a sensitive indicator of vaccinia virulence. Whether HIV AC-l e is less likely than control vaccinia virus to cause vaccinia-related complications in humans is therefore still unknown. Alternative strategies for controlling the virulence of poxvirus vectors are being developed and will provide an additional degree of safety. There was no apparent difference in viral replication or local reactions between doses. We concluded that inocula-
60 0
50
use of vaccine in areas with a high prevalence of HIV infection. A recent trial in which HIV-seropositive subjects were immunized with paraformaldchyde-fixcd, autologous Epstein-Barr virus-transformed B cells infected with recombinant vaccinia virus expressing the env and gag gene products [29] may have been complicated by vaccinia necrosum [30]. Widespread use of an unattenuated vaccinia vector in the field might therefore require prescreening for HIV status. Containment was an aspect of safety emphasized in this trial. Persons who had contact with those at special risk for vaccinia complications were excluded from participation. Limited experience with transparent dressings in an earlier inpatient trial of a recombinant vaccinia vaccine suggested that this approach provided effective containment [20]. We found that when the dressing was handled under optimal conditions by skilled personnel, virus containment was secure. Leakage of exudate, requirement for a person other than the vaccinee to apply the dressing, and the problem of used dressing disposal, however, are concerns that need to be addressed before this dressing technique can be recom-
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DA YS AFTER PRIMARY VACCINATION Figure 5. Lymphoproliferative responses to I J.Lg of recombinant envelope glycoprotein (rgp 160) for 5 control vaccinia recipients and 10 HIVAC-I e recipients are depicted as stimulation index of cryopreserved peripheral blood mononuclear cells as function of time after vaccination. Note y axis scale is interrupted. Differences between groups were not significant at any individual time point by Wilcoxon rank sum test. Two individuals with high background responses to baculovirus-derived protein prior to vaccination are not depicted or included in analysis.
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lID 1992; 166 (August)
JID 1992; 166 (August)
gp 160 Recombinant Vaccinia Vaccine
it induces a stronger immune response to the foreign gene product in vaccinia-naive adults than in vaccinia-immune adults [20]. We have also demonstrated a long duration of gp 160-specific antibody response in vaccine recipients. Ultimately, the perceived risks and benefits will determine whether there are sufficient immunologic advantages to justify large-scale vaccination with live recombinant vaccinia. The long duration of gp 160-specific antibody induced by HIVAC-1e is encouraging in this regard, as is the documentation of CTL responses specific to the foreign gene product in primates [11, 12]. Previous studies in mice have also shown that recombinant vaccinia may be an exceptionally good way to prime the immune system for boosting with a subunit vaccine [34], and recent studies have indicated that protective immunity could be elicited by this combination immunization regimen in the SIV macaque model [35]. This regimen is currently under investigation in humans [36]. Additional investigation is therefore justified to explore potentially unique advantages ofimmunization with live recombinant vaccinia vectors against HIV and other pathogens. Acknowledgments
We acknowledge the substantial contributions of the following individuals: David T. Karzon, Lois Wagner, B. J. Hensman, Lori Crane, Lori Bunton, Donna Hummell, Chris Parham, Ji Ying Zhou, and Mentoria Jennings (Vanderbilt University School of Medicine); T. Ulf Westblom, Donald J. Kennedy, Sharon E. Frey, Carol Berry, William LaRock, Sharon Nugent, Frances Newman, Gira Patel, Mahendra Mandava, Kathy Feurer, and Judy Hayes (S1. Louis University School of Medicine); Carol Hilton, LeslieGarrison, Sally Slome, Edward Ellerbeck, Charles Flexner, William Adler, Robert Cormier, Michael Prenger, Karen Christina, and Bhavin Thumar (Johns Hopkins University School of Medicine); William Bonnez, Richard C. Reichman, Nizar Tejani, ShirleyErb, Carol Greisberger, Joanne Strussenberg, Suzan Cole, and Steven Pomeroy (University of Rochester School of Medicine and Dentistry); Elizabeth Cooney, Phillip Greenberg, and David Berger (University of Washington); Patricia E. Fast, Sue L. Wescott, and Mary Clare Walker (National Institute of Allergy and Infectious Diseases); Carol M. Smith, Donna M. Brown, and Joel W. Novack (EMMES Corporation); Marcia Hanson and Tracy Dykers (Bristol-Meyers Squibb); and Ronald Engle, Gail O. Dapolito, Allyson F. Chambers, Karen E. Myers, Edmund Leung, and Paul J. Cote (Georgetown University School of Medicine).
References I. Mackett M, Smith Gl., Moss B. Vaccinia virus: a selectable eukaryotic cloning and expression vector. Proc Natl Acad Sci USA 1982;
79:7415-9. 2. Panicali D. Paoletti E. Construction of poxviruses as cloning vectors: insertion ofthe thymidine kinase gene from herpes simplex virus into the DNA of infectious vaccinia virus. Proc Natl Acad Sci USA
1982;79:4927-31. 3. Smith G. Murphy BR. Moss B. Construction and characterization of an
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tion leading to lesion formation resulted in the same set of clinical and immunologic responses regardless ofdose. Of22 vaccinia-naive HIVAC-I e recipients, 16 developed HIV-l envelope-specific antibodies. Although low in titer, these gp 160-specific antibodies persisted for> 1 year in >50% of responders. This duration of humoral response has not been seen in subjects immunized with low dosages of gp 160 subunit vaccine products until after the fourth dose, given 18 months after the primary immunization [33]. The mechanism underlying the long duration of antibody induced by recombinant vaccinia vaccination is not known. Persistence of antigen in follicular dendritic cells or other antigen-presenting cells, however, is an attractive possibility and would also be expected to expand populations ofmemory cells. The low frequency of serum neutralizing activity in volunteers was not surprising given the low magnitude ofgp 160-specific antibody response. Revaccination with HIV AC-1e or Dryvax did not result in lesion formation if a lesion occurred after primary vaccination and there was no consequent immune response to revaccination in the absence of a lesion. This implies that although primary vaccination in vaccinia-naive adults is likely to result in lesion formation and expression of the foreign gene product, proximate revaccination with recombinant vaccinia will not result in boosting of the response to the foreign gene product. Persons vaccinated with vaccinia>20 years ago reportedly have a different response to revaccination, with most developing lesions after primary vaccination and r - 50%experiencing another lesion after second vaccination [20]. Whereas there is no evidence that proximate revaccination will boost responses in vaccinia-naive persons, the difference in rates of lesion formation after revaccination in vaccinia-immune persons provides a rationale for repeated vaccination in that group. It may ultimately be possible to demonstrate evidence ofa booster response in vaccinia-naive persons by modifying the route of revaccination or by lengthening the interval between vaccinations. Lymphoproliferative responses specific for gp 160 were detected in 5 of 10 vaccinees, indicating that HIV AC-1e can induce cell-mediated immune responses in addition to humoral responses as previously shown [20]. The expression of native gp 160 on the host cell membrane is one potential advantage of recombinant vaccinia over non replicating vaccine products. Another immunologic advantage of expressing gp 160 in a live vector is that antigen will be presented in the context of both class I and class II MHC antigens. Indeed, recombinant vaccinia viruses expressing retroviral glycoproteins have been shown to induce CD8+ MHC class 1restricted CTL in monkeys [II, 12] and CD4+ CTL in chimpanzees [15]. These responses have not yet been found in volunteers in this study. This was the first large trial evaluating recombinant vaccinia in vaccinia-naive individuals. We have extended the observation that gp 160 recombinant vaccinia is safe and that
251
Graham et al,
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infectious vaccinia virus recombinant that expresses the influenza hemagglutinin gene and induces resistance to influenza virus infection in hamsters. Proc Natl Acad Sci USA 1983:80:7155-9.
20.
4. Wiltor TJ. Macfarlan RL Reagan KJ. et al. Protection from rabies by vaccinia virus recombinant containing the rabies glycoprotein gene. Proc Natl Acad Sci USA 1984;81:7194-8.
21.
5. Paoletti E, Lipinskas BR. Sarnsonoff C, Mercer S. Panicali D. Construction oflive vaccines using genetically engineered pox viruses: biological activity of vaccinia virus recombinants expressing the hepatitis B virus surface antigen and the herpes simplex virus glycoprotein D. Proc Natl Acad Sci USA 1984;81: 193-7.
7. Yilma T. Hsu D. Jones L. et al. Protection of cattle against rinderpest with vaccinia virus recombinants expressing the HA or F gene. Science 1988;242: 1058-61. 8. Moss B. Smith GL, Gerin JL. Purcell RH. Live recombinant vaccinia virus protects chimpanzees against hepatitis B. Nature 1984;311:679.
9. Hu SL. Zarling IM, Chin J, et al. Protection of macaques against simian AIDS by immunization with a recombinant vaccinia virus expressing the envelope glycoprotein of simian type 0 retrovirus. Proc Natl Acad Sci USA 1989;86:7213-7. 10. Bennick JR, Yewdell JW. Smith GL. Moller C, Moss B. Recombinant vaccinia virus primes and stimulates influenza haemagglutinin-specifie cytotoxic T cells. Nature 1984;311 :578-9. 1 L Shen L. Chen ZW, Miller MD, et al. Recombinant virus vaccine-induced Sl V-specific CD8+ cytotoxic T lymphocytes. Science 1991;252:440-3. 12. Gotch FM, Hovell R. Delchambre M, Silvera P, McMichael AJ. Cytotoxic T-cell response to simian immunodeficiency virus by cynomolgus macaque monkeys immunized with recombinant vaccinia virus. AIDS 1991;5:317-20. 13. Chakrabarti S. Robert-Guroff M, Wong-Staal F. Gallo RC, Moss B. Expression of the HTLV-III envelope gene by a recombinant vaccinia virus. Nature 1986;320: 535-7. 14. Hu SL. Kosowski SG, Dalrymple JM. Expression of AIDS virus envelope gene in recombinant vaccinia viruses. Nature 1986;320:537-9. 15. Zarling JM. Eichberg JW, Moran PA, McClure J. Sridhar P, Hu SL. Proliferative and cytotoxic T cells to AIDS virus glycoproteins in chimpanzees immunized with a recombinant vaccinia virus expressing AIDS virus envelope glycoproteins. J Immunol 1987; 139:98890. 16. Jones L, Ristow S, Yilma T. Moss B. Accidental human vaccination with vaccinia virus expressing nucleoprotein gene. Nature 1986;319:543. 17. Openshaw PJM, Alwan WH, Cherrie AH, Record FM. Accidental infection oflaboratory worker with recombinant vaccinia virus. Lancet 1991;338:459. 18. Zagury 0, Leonard R, Fouchard M, et al. Immunization against AIDS in humans. Nature 1987;326:249-50. 19. Zagury D. Bernard J, Cheynier R. et al. A group specific anamnestic
23.
24. 25. 26.
immune reaction against HIV -I induced by a candidate vaccine against AIDS. Nature 1988;332:728-31. Cooney EL. Collier AC, Greenberg PO, et al. Safety ofand immunological response to a recombinant vaccinia virus vaccine expressing HIV envelope glycoprotein. Lancet 1991;337:567-72. Kniker WT, Anderson CT. Rourneantzcff M. The MULTITEST system: a standardized approach to evaluation of delayed hypersensitivity and cell mediated immunity. Ann Allergy 1979;43:73-9. Fenner F. Henderson DA, Arita L Jezek Z, Ladnyi 10. Smallpox and its eradication. Geneva: World Health Organization, 1988. Montefiori DC, Robinson WE, Schuffman SS, Mitchell WM. Evaluation ofantiviral drugs and neutralizing antibodies to human immunodeficiency virus by a rapid and sensitive microtiter infection assay. J Clin Microbiol 1988;26:231-5. Graham BS, Bunton LA, Wright PF, Karzon DT. Reinfection of mice with respiratory syncytial virus. J Med Virol 1991;34:7-13. Brernan JG, Arita I. The confirmation and maintenance of smallpox eradication. N Engl J Med 1980;303: 1263-73. Baumgaertner JC, Hogan R, Borm C, Berg J, Davis JP. Contact spread of vaccinia from a National Guard vaccinee. MMWR 1985;34: 182-
3. 27. Goldstein JA, Neff JM, Lane JM, Koplan JP. Smallpox vaccination reactions, prophylaxis, and therapy of complications. Pediatrics 1975;55:342-7. 28. Redfield RR, Wright DC, James WD, Jones TS, Brown C, Burke OS. Disseminated vaccinia in a military recruit with human immunodeficiency virus (HIV) disease. N Engl J Med 1987;316:673-6. 29. Picard 0, Giral P, Defer MC, et al. AIDS vaccine therapy: phase I trial. Lancet 1990;336: 179. 30. Guillaume JC, Saiag P, Wechsler J, Lescs MC, Roujeau Jc. Vaccinia from recombinant virus expressing HIV genes. Lancet 1991;337: 1034-5. 31. Buller RML. Smith GL, Cremer K. Notkins AL, Moss B. Decreased virulence of recombinant vaccinia virus expression vectors is associated with a thymidine kinase-negative phenotype. Nature 1985;317:813-5. 32. Taylor G, Stott EJ, Wertz G, Ball A. Comparison of the virulence of wild-type thymidine kinase (tk)-deficient and tk+ phenotypes ofvaccinia virus recombinants after intranasal inoculation of mice. J Gen ViroI1991;72:125-30. 33. Dolin R, Graham B, Greenberg SB, et al. Safety and immunogenicity of an HIV-I recombinant gpl60 candidate vaccine in humans. Ann Intern Med 1991;114:119-27. 34. Hu SL, Klaniecki J, Dykers T. Sridhar P, Travis BM. Neutralizing antibodies against HIV-I BRU and SF-2 isolates generated in mice expressing HIV -I envelope glycoproteins and boosted with gp 160. AIDS Res Hum Retroviruses 1991;7:615-20. 35. Hu SL, Abrams K Barber GN, et al. Protection of macaques against SIV infection by subunit vaccines of SIV envelope glycoprotein gp 160. Science 1992;255:456-9. 36. Graham BS, Belshe R, Clements ML, et al. HIV-gpI60 recombinant vaccinia vaccination of vaccinia-naive adults followed by rgp 160 booster immunization [abstract F.A.I.] In: Program and abstracts of the VII International Conference on AIDS (Florence. Italy). Rome: Instituto Superiore di Sanita, 1991.
Downloaded from http://jid.oxfordjournals.org/ at University of Iowa Libraries/Serials Acquisitions on June 5, 2016
6. Mackett M. Yilma T. Rose J K. Moss B. Vaccinia virus recombinants: expression of VSV genes and protective immunization of mice and cattle. Science 1985;227:433-5.
22.
JID 1992;166 (August)
Vaccination of Vaccinia-Naive Adults with Human Immunodeficiency Virus Type 1 gp160 Recombinant Vaccinia Virus in a Blinded, Controlled, Randomized Clinical Trial Vanderbilt University School o] Medicine. Nashville. Tennessee: St. Louis Universitv School of Medicine. St. Louis. Missouri: Universit v o]' Rochester School of Medicine and Dentistrv. Rochester. New York: Bristol-Myers Squibb Pharmaceutical Research Institute and University of Washington School (1' Medicine. Seattle; Johns Hopkins School of Hygiene and Public Health and School of Medicine. Baltimore. Georgetown University. Rockville. EMMES Corporation. Potomac. and National Institutes of Health, Bethesda. Maryland
The safety and immunogenicity of a human immunodeficiency virus type 1 (HIV-l) gp160 recombinant vaccinia virus (HIVAC-le) vaccine was evaluated in vaccinia-naive, healthy adults at low risk for acquiring HIV-l infection. Volunteers (n = 36) were randomized to receive HIVAC-l e or control vaccinia virus at two dosages by bifurcated needle puncture at 0 and 2 months; 12 HIVAC-le and 6 control vaccinia virus recipients received either 106 or 107 pfujmL at each inoculation. There was no significant difference in lesion size, level of viral replication, or systemic symptoms after vaccination with HIVAC-le or control vaccinia virus. Of 22 HIVAC-le recipients with lesion formation, 16 developed low-titer gp 160-specific antibody responses detectable by Western blot. The peak response occurred between days 70 and 120 and was still detectable at day 365 in 9 of 18 vaccinees. gp 160-specific Iymphoproliferative responses were detected in 5 of 10 vaccinees. Vaccination with HIVAC-le was safe in vaccinia-naive, healthy adults and could induce both humoral and cell-mediated gp 160-specific immune responses.
The precedent for using vaccinia virus in a successful, worldwide immunization program makes it an attractive vehicle for presentation of novel antigens. Therefore, the capability to engineer vaccinia virus to express foreign genes [I, 2] was an important breakthrough in vaccine development. Recombinant vaccinia viruses expressing the appropriate foreign genes have since been shown to provide protective immunity against experimental challenge with influenza virus in hamsters [3], rabies virus in rabbits and mice [4],
Received 29 January 1992: revised 12 March 1992. Presented in part: 3rd annual meeting of the National Cooperative Vaccine Development Groups for AIDS. Clearwater Beach. Florida. October 1990; VII International Conference on AIDS. Florence. Italy. June 1991 (abstract F.A.I). All volunteers underwent an extensive informed consent process and were required to comprehend fully the purpose and details of the study. All protocols and consent forms were approved by local Institutional Review Boards and Institutional Biosafety Committees. The study was conducted according to the human experimentation guidelines of the US Department of Health and Human Services. Grant support: National Institutes of Health (AI-82500. -05061. -05062. -05063. -05064. -05065. -15106). Reprints or correspondence: Dr. Barney S. Graham. Department ofMedicine. Division of Infectious Diseases. A-331 0 MCN. Vanderbilt University School of Medicine. Nashville. TN 37232.
The Journal of Infectious Diseases 1992;166:244-52 © 1992 by The University of Chicago, All rights reserved. 0022-1899/92/6602-0003$01.00
herpes simplex virus in mice [5], vesicular stomatitis virus in mice and cattle [6], rinderpest virus in cattle [7], hepatitis B virus in chimpanzees [8], and simian AIDS caused by a type D retrovirus in macaques [9]. Vaccinia recombinant vectors can induce both humoral antibody responses and cellular immune responses specific to the inserted gene product(s). Induction of cytotoxic T lymphocytes (CTL) is thought to be particularly important in containing human immunodeficiency virus type I (HIYI) infection. Expression of the influenza hemagglutinin by recombinant vaccinia virus vectors has been shown to induce hemagglutinin-specific major histocompatibility complex (MHC) class I-restricted CTL in inbred mice [10]. Simian immunodeficiency virus (SIV) genes expressed in vaccinia recombinant viruses have been shown to induce Sl V-specific MHC class I-restricted CTL in monkeys [II, 12]. The envelope gene of HIV-l, expressed in vaccinia, has induced gp 160-specific antibody in mice and rhesus monkeys [13, 14]. Vaccination of chimpanzees with vaccinia recombinant virus expressinggp 160 induced HIV-specific lyrnphoproliferative responses and MHC class II-restricted CD4+ CTL [15]. Although there is extensive clinical experience with native vaccinia, there is limited experience with recombinant vaccinia viruses in humans in terms of either clinical behavior of the recombinant viruses or their ability to effectively express
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Barney S. Graham, Robert B. Belshe, Mary Lou Clements, Raphael Dolin, Lawrence Corey, Peter F. Wright, Geoffrey J. Gorse, Karen Midthun, Michael C. Keefer, Norbert J. Roberts, Jr., David H. Schwartz, Jan M. Agosti, Bruce F. Fernie, Donald M. Stablein, David C. Montefiori, John S. Lambert, Shiu-Lok Hu, Joy R. Esterlitz, Dale N. Lawrence, Wayne C. Koff, and the AIDS Vaccine Clinical Trials Network
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gpl60 Recombinant Vaccinia Vaccine
Materials and Methods Vaccine Product Preparation HIV AC-l e (Bristol-Myers Squibb, Seattle) is the product designation for the vaccinia recombinant containing the HIV-1 envelope gene [14]. The parent vaccinia virus was derived by plaque purification from a commercial preparation of smallpox vaccine (Dryvax, lot 321501 G; Wyeth Laboratories, Philadelphia) grown in BSC-40 cells (derived from BSC-l, ATCC CCL26). The master seed lot of the recombinant virus was then prepared in human diploid MRC-5 cells (ATCC CeL-171). The vaccine virus was prepared in MRC-5 cells by inoculation with master seed virus. The infected cell monolayer was washed twice with PBS; cells were scraped and pelleted by centrifugation at 1500 g. resuspended in PBS, and then ruptured by two cycles offreeze-thaw and sonication. After shipment and storage at -70°C, the resulting suspension had a titer of --2 X 108 pfu/ml, as determined at inoculation. A single lot ofcommercial native vaccinia product (Dryvax; Wyeth) prepared as lyophilized calflymph was reconstituted before use with a diluent containing 50%glycerin. The titer of the control vaccinia virus suspension was -- 2 X 108 pfu/rnl..
Subjects Inclusion criteria were that the subjects be healthy HIV-seronegative adults 18-60 years old with negligible risk for HIV-l exposure and with no evidence of prior smallpox vaccination. Volunteers were excluded if there were any indications of highrisk behavior for HIV-l infection (determined by history, questionnaire, and laboratory evaluation). Volunteers were excluded if there was any evidence of chronic illness or if they or their
Table 1. Clinical and virologic characteristics of vaccinia-naive adults vaccinated with different doses of human immunodeficiency virus type I gp 160 recombinant vaccinia virus (HIVAC-I e) or control vaccinia virus. Dosage 107 pfu/rnl.
106 pfu/ml.
Subjects Men Women Total Heterosex ual Homosexual Age (mean years) No. with lesion formation after Primary vaccination Secondary vaccination Total with lesion formation No lesion formation Maximum skin titer Day of maximum titer Last day of shedding
HIVAC-Ie
Control
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9 3 12 10 2 20.0
5 I 6 4 2 20.5
9 3 12 8 4 23.5
4 2 6 4 2 21.5
5
4
11*
6
5
2
10 2t 5.6 ± 0.4
6 0 6.1 ± 0.6
12 0* 5.0 ± 0.4
6 0 5.5±0.7
II (6-18)
10 (5-14)
8 (4-20)
14 (5-21)
21(14-22)
19 (9-21)
18 (6-26)
18 (9-21)
P
NOTE. Maximum skin titers are given as mean loglo pfu/rnl, ± SE. Days of maximum titer and shedding are given as median (range). * Two volunteers were vaccinated with stocks stored at - 35°C for> 12 months: virus titers in inocula were 1000-fold in titer from inside to outside (mean 10gIO pfu/ml. inside was 7.0 vs. 3.4 outside). Genetic stability of HIVAC-le. The last samples from which virus could be isolated in 5 HIV AC-I e recipients (days 8, 15, 15, 18, and 20) and one Dryvax recipient were evaluated for gp 160 expression. None of the Dryvax plaques (>500) had evidence ofgpl60 expression. Of> 1000 individual plaques from HIVAC-I e recipients, all showed gp 160 expression. Vaccinia-specific antibody responses to vaccination. ELISA (figure I) and plaque-reduction neutralization tests showed that vaccinia-specific antibody could not be detected until after an inoculation that resulted in lesion formation. The level of antibody was not augmented further in persons who had lesion formation after initial vaccination and then received a booster vaccination. HI V-specific antibody responses to vaccination. The antibody responses to vaccination were recorded from the time of the inoculation that resulted in lesion formation. Western blot assays were used to detect antibody responses to the HIV-I envelope glycoproteins gp 160, gp 120, or gp41. In 8 of 10 HIVAC-Ie 106-dose recipients and 8 of 12 HIVAC-Ie 107-dose recipients, antibody was detected. In 4 106-dose HIVAC-I e recipients, antibody to HIV-I envelope bands did
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not develop until day 120 after lesion formation, and in 3 of those 4 recipients, antibody persisted past day 365. Overall, 9 of 18 HIV AC-l e recipients had gp l60-specific antibody detectable by Western blot beyond day 365. The earliest development of antibody to HIV-I envelope bands was day 28. The responses were generally weak, with serum from only 1 volunteer showing a positive intensity against envelope bands on Western blot at one time point (figure 4). The delayed development ofantibody made interpreting the HIV antibody response to the second vaccination difficult. However, antibody did not appear to be augmented after revaccination. Evaluation of antibody responses by ELISA identified only 1 volunteer at one time point as positive, with a ratio of test sample 00 to cutoffof2.35. This serum, from day 210 after vaccination, was the same sample that tested strongly
positive by Western blot. Neutralizing antibody activity against the homologous HIV -1 IIIB strain was detected only in the day 180 serum from the individual with the strong positive Western blot and positive ELISA. Cell-mediated immune responses to vaccination. Lymphoproliferative responses specific for HIV-I gp 160 and responses induced by mitogen and another recall antigen were measured. Lymphoproliferative responses to phytohemagglutinin and Candida antigen remained stable beyond day 365. No control vaccinia virus recipients developed a significant Iymphoproliferative response (SI >5.0) to rgp 160, whereas 5 of 10 HIVAC-Ie recipients did (figure 5). The peak number of responders (5/1 0) occurred on day 120 after vaccination. Only I of 6 still had significant rgp 160 responses on day 365.
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DAYS AFTER PRIMARY VACCINATION Figure 3. Absolute CD4 lymphocyte counts throughout followup after vaccination. There was no deleterious effect on CD4 cell numbers as a result of gp 160 expression.
The induction of immunity through presentation of antigen by live recombinant vectors is an important vaccination strategy. Vaccinia is among a number of viral and bacterial recombinant vaccine vectors in development; it offers many advantages as a vaccine vehicle including low cost, heat stability, and documentation of vaccination via a scar. Its ease of administration in developing nations is attested by its use in the global eradication of smallpox [22, 25]. However, there are drawbacks to consider. Spread of vaccinia to contacts is well recognized [26]. Transmission to healthy individuals is a concern, especially with regard to ocular infection and unanticipated acquisition of seropositivity to the foreign gene product. The greater concern, however, is accidental transmission to people with eczema or immunodeficiency [27]. Vaccinia virus has been reported to have caused a severe local reaction and dissemination in an asymptomatic but severely immunodeficient HIV-I-infected person [28]. This has implications for the
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DA YS AFTER PRIMARY VACCINATION Figure 4. Human immunodeficiency virus type I (HIV-I) envelope-specific Western blot responses in volunteers after vaccinations that led to lesion formation. Results are aligned from time of inoculation that resulted in lesion formation. Band intensity was graded according to kit directions as absent (-); present, but less intense than anti-p24 band on weakly reactive control strip (± or visible); at least as intense as anti-p24 band on weakly reactive control strip, but less intense than anti-p24 on strongly reactive control strip (+ or positive); or greater than or equal to intensity of anti-p24 on strongly reactive control strip (++ or strong positive). Percentage of volunteers with responses represented by bars with absolute number of positive responses per number tested recorded above bar.
mended for virus containment in large-scale trials or for routine vaccination. Nevertheless, for small phase I or II trials in which study personnel can change the dressing, the transparent dressing is an effective barrier. The thymidine kinase (TK) gene, which is interrupted in the HIV AC-l e product, is considered to be a determinant of virulence. TK+ vaccinia is more virulent than TK- vaccinia in mice when the inoculation is given parenterally [31] or into the lung [32]. It was therefore anticipated that HIV ACl e would be less virulent than control vaccinia virus. Analysis of viral replication, size and character of skin lesions, and serologic responses after vaccination, however, showed no differences between the HIVAC-Ie and control vaccinia virus products in this trial. Although there was no evidence of HIV AC-l e attenuation by these measures of virulence, HIV AC-I e was derived from a plaque-purified vaccinia stock that may not accurately reflect the composite effects of minor strain variants in the control vaccinia virus product. Also, in mice inoculated by scarification, there is no apparent attenuation of TK- vaccinia as measured by mortality, viral dissemination, or serologic responses [31], so dermal inoculation may not be a sensitive indicator of vaccinia virulence. Whether HIV AC-l e is less likely than control vaccinia virus to cause vaccinia-related complications in humans is therefore still unknown. Alternative strategies for controlling the virulence of poxvirus vectors are being developed and will provide an additional degree of safety. There was no apparent difference in viral replication or local reactions between doses. We concluded that inocula-
60 0
50
use of vaccine in areas with a high prevalence of HIV infection. A recent trial in which HIV-seropositive subjects were immunized with paraformaldchyde-fixcd, autologous Epstein-Barr virus-transformed B cells infected with recombinant vaccinia virus expressing the env and gag gene products [29] may have been complicated by vaccinia necrosum [30]. Widespread use of an unattenuated vaccinia vector in the field might therefore require prescreening for HIV status. Containment was an aspect of safety emphasized in this trial. Persons who had contact with those at special risk for vaccinia complications were excluded from participation. Limited experience with transparent dressings in an earlier inpatient trial of a recombinant vaccinia vaccine suggested that this approach provided effective containment [20]. We found that when the dressing was handled under optimal conditions by skilled personnel, virus containment was secure. Leakage of exudate, requirement for a person other than the vaccinee to apply the dressing, and the problem of used dressing disposal, however, are concerns that need to be addressed before this dressing technique can be recom-
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DA YS AFTER PRIMARY VACCINATION Figure 5. Lymphoproliferative responses to I J.Lg of recombinant envelope glycoprotein (rgp 160) for 5 control vaccinia recipients and 10 HIVAC-I e recipients are depicted as stimulation index of cryopreserved peripheral blood mononuclear cells as function of time after vaccination. Note y axis scale is interrupted. Differences between groups were not significant at any individual time point by Wilcoxon rank sum test. Two individuals with high background responses to baculovirus-derived protein prior to vaccination are not depicted or included in analysis.
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lID 1992; 166 (August)
JID 1992; 166 (August)
gp 160 Recombinant Vaccinia Vaccine
it induces a stronger immune response to the foreign gene product in vaccinia-naive adults than in vaccinia-immune adults [20]. We have also demonstrated a long duration of gp 160-specific antibody response in vaccine recipients. Ultimately, the perceived risks and benefits will determine whether there are sufficient immunologic advantages to justify large-scale vaccination with live recombinant vaccinia. The long duration of gp 160-specific antibody induced by HIVAC-1e is encouraging in this regard, as is the documentation of CTL responses specific to the foreign gene product in primates [11, 12]. Previous studies in mice have also shown that recombinant vaccinia may be an exceptionally good way to prime the immune system for boosting with a subunit vaccine [34], and recent studies have indicated that protective immunity could be elicited by this combination immunization regimen in the SIV macaque model [35]. This regimen is currently under investigation in humans [36]. Additional investigation is therefore justified to explore potentially unique advantages ofimmunization with live recombinant vaccinia vectors against HIV and other pathogens. Acknowledgments
We acknowledge the substantial contributions of the following individuals: David T. Karzon, Lois Wagner, B. J. Hensman, Lori Crane, Lori Bunton, Donna Hummell, Chris Parham, Ji Ying Zhou, and Mentoria Jennings (Vanderbilt University School of Medicine); T. Ulf Westblom, Donald J. Kennedy, Sharon E. Frey, Carol Berry, William LaRock, Sharon Nugent, Frances Newman, Gira Patel, Mahendra Mandava, Kathy Feurer, and Judy Hayes (S1. Louis University School of Medicine); Carol Hilton, LeslieGarrison, Sally Slome, Edward Ellerbeck, Charles Flexner, William Adler, Robert Cormier, Michael Prenger, Karen Christina, and Bhavin Thumar (Johns Hopkins University School of Medicine); William Bonnez, Richard C. Reichman, Nizar Tejani, ShirleyErb, Carol Greisberger, Joanne Strussenberg, Suzan Cole, and Steven Pomeroy (University of Rochester School of Medicine and Dentistry); Elizabeth Cooney, Phillip Greenberg, and David Berger (University of Washington); Patricia E. Fast, Sue L. Wescott, and Mary Clare Walker (National Institute of Allergy and Infectious Diseases); Carol M. Smith, Donna M. Brown, and Joel W. Novack (EMMES Corporation); Marcia Hanson and Tracy Dykers (Bristol-Meyers Squibb); and Ronald Engle, Gail O. Dapolito, Allyson F. Chambers, Karen E. Myers, Edmund Leung, and Paul J. Cote (Georgetown University School of Medicine).
References I. Mackett M, Smith Gl., Moss B. Vaccinia virus: a selectable eukaryotic cloning and expression vector. Proc Natl Acad Sci USA 1982;
79:7415-9. 2. Panicali D. Paoletti E. Construction of poxviruses as cloning vectors: insertion ofthe thymidine kinase gene from herpes simplex virus into the DNA of infectious vaccinia virus. Proc Natl Acad Sci USA
1982;79:4927-31. 3. Smith G. Murphy BR. Moss B. Construction and characterization of an
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tion leading to lesion formation resulted in the same set of clinical and immunologic responses regardless ofdose. Of22 vaccinia-naive HIVAC-I e recipients, 16 developed HIV-l envelope-specific antibodies. Although low in titer, these gp 160-specific antibodies persisted for> 1 year in >50% of responders. This duration of humoral response has not been seen in subjects immunized with low dosages of gp 160 subunit vaccine products until after the fourth dose, given 18 months after the primary immunization [33]. The mechanism underlying the long duration of antibody induced by recombinant vaccinia vaccination is not known. Persistence of antigen in follicular dendritic cells or other antigen-presenting cells, however, is an attractive possibility and would also be expected to expand populations ofmemory cells. The low frequency of serum neutralizing activity in volunteers was not surprising given the low magnitude ofgp 160-specific antibody response. Revaccination with HIV AC-1e or Dryvax did not result in lesion formation if a lesion occurred after primary vaccination and there was no consequent immune response to revaccination in the absence of a lesion. This implies that although primary vaccination in vaccinia-naive adults is likely to result in lesion formation and expression of the foreign gene product, proximate revaccination with recombinant vaccinia will not result in boosting of the response to the foreign gene product. Persons vaccinated with vaccinia>20 years ago reportedly have a different response to revaccination, with most developing lesions after primary vaccination and r - 50%experiencing another lesion after second vaccination [20]. Whereas there is no evidence that proximate revaccination will boost responses in vaccinia-naive persons, the difference in rates of lesion formation after revaccination in vaccinia-immune persons provides a rationale for repeated vaccination in that group. It may ultimately be possible to demonstrate evidence ofa booster response in vaccinia-naive persons by modifying the route of revaccination or by lengthening the interval between vaccinations. Lymphoproliferative responses specific for gp 160 were detected in 5 of 10 vaccinees, indicating that HIV AC-1e can induce cell-mediated immune responses in addition to humoral responses as previously shown [20]. The expression of native gp 160 on the host cell membrane is one potential advantage of recombinant vaccinia over non replicating vaccine products. Another immunologic advantage of expressing gp 160 in a live vector is that antigen will be presented in the context of both class I and class II MHC antigens. Indeed, recombinant vaccinia viruses expressing retroviral glycoproteins have been shown to induce CD8+ MHC class 1restricted CTL in monkeys [II, 12] and CD4+ CTL in chimpanzees [15]. These responses have not yet been found in volunteers in this study. This was the first large trial evaluating recombinant vaccinia in vaccinia-naive individuals. We have extended the observation that gp 160 recombinant vaccinia is safe and that
251
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infectious vaccinia virus recombinant that expresses the influenza hemagglutinin gene and induces resistance to influenza virus infection in hamsters. Proc Natl Acad Sci USA 1983:80:7155-9.
20.
4. Wiltor TJ. Macfarlan RL Reagan KJ. et al. Protection from rabies by vaccinia virus recombinant containing the rabies glycoprotein gene. Proc Natl Acad Sci USA 1984;81:7194-8.
21.
5. Paoletti E, Lipinskas BR. Sarnsonoff C, Mercer S. Panicali D. Construction oflive vaccines using genetically engineered pox viruses: biological activity of vaccinia virus recombinants expressing the hepatitis B virus surface antigen and the herpes simplex virus glycoprotein D. Proc Natl Acad Sci USA 1984;81: 193-7.
7. Yilma T. Hsu D. Jones L. et al. Protection of cattle against rinderpest with vaccinia virus recombinants expressing the HA or F gene. Science 1988;242: 1058-61. 8. Moss B. Smith GL, Gerin JL. Purcell RH. Live recombinant vaccinia virus protects chimpanzees against hepatitis B. Nature 1984;311:679.
9. Hu SL. Zarling IM, Chin J, et al. Protection of macaques against simian AIDS by immunization with a recombinant vaccinia virus expressing the envelope glycoprotein of simian type 0 retrovirus. Proc Natl Acad Sci USA 1989;86:7213-7. 10. Bennick JR, Yewdell JW. Smith GL. Moller C, Moss B. Recombinant vaccinia virus primes and stimulates influenza haemagglutinin-specifie cytotoxic T cells. Nature 1984;311 :578-9. 1 L Shen L. Chen ZW, Miller MD, et al. Recombinant virus vaccine-induced Sl V-specific CD8+ cytotoxic T lymphocytes. Science 1991;252:440-3. 12. Gotch FM, Hovell R. Delchambre M, Silvera P, McMichael AJ. Cytotoxic T-cell response to simian immunodeficiency virus by cynomolgus macaque monkeys immunized with recombinant vaccinia virus. AIDS 1991;5:317-20. 13. Chakrabarti S. Robert-Guroff M, Wong-Staal F. Gallo RC, Moss B. Expression of the HTLV-III envelope gene by a recombinant vaccinia virus. Nature 1986;320: 535-7. 14. Hu SL. Kosowski SG, Dalrymple JM. Expression of AIDS virus envelope gene in recombinant vaccinia viruses. Nature 1986;320:537-9. 15. Zarling JM. Eichberg JW, Moran PA, McClure J. Sridhar P, Hu SL. Proliferative and cytotoxic T cells to AIDS virus glycoproteins in chimpanzees immunized with a recombinant vaccinia virus expressing AIDS virus envelope glycoproteins. J Immunol 1987; 139:98890. 16. Jones L, Ristow S, Yilma T. Moss B. Accidental human vaccination with vaccinia virus expressing nucleoprotein gene. Nature 1986;319:543. 17. Openshaw PJM, Alwan WH, Cherrie AH, Record FM. Accidental infection oflaboratory worker with recombinant vaccinia virus. Lancet 1991;338:459. 18. Zagury 0, Leonard R, Fouchard M, et al. Immunization against AIDS in humans. Nature 1987;326:249-50. 19. Zagury D. Bernard J, Cheynier R. et al. A group specific anamnestic
23.
24. 25. 26.
immune reaction against HIV -I induced by a candidate vaccine against AIDS. Nature 1988;332:728-31. Cooney EL. Collier AC, Greenberg PO, et al. Safety ofand immunological response to a recombinant vaccinia virus vaccine expressing HIV envelope glycoprotein. Lancet 1991;337:567-72. Kniker WT, Anderson CT. Rourneantzcff M. The MULTITEST system: a standardized approach to evaluation of delayed hypersensitivity and cell mediated immunity. Ann Allergy 1979;43:73-9. Fenner F. Henderson DA, Arita L Jezek Z, Ladnyi 10. Smallpox and its eradication. Geneva: World Health Organization, 1988. Montefiori DC, Robinson WE, Schuffman SS, Mitchell WM. Evaluation ofantiviral drugs and neutralizing antibodies to human immunodeficiency virus by a rapid and sensitive microtiter infection assay. J Clin Microbiol 1988;26:231-5. Graham BS, Bunton LA, Wright PF, Karzon DT. Reinfection of mice with respiratory syncytial virus. J Med Virol 1991;34:7-13. Brernan JG, Arita I. The confirmation and maintenance of smallpox eradication. N Engl J Med 1980;303: 1263-73. Baumgaertner JC, Hogan R, Borm C, Berg J, Davis JP. Contact spread of vaccinia from a National Guard vaccinee. MMWR 1985;34: 182-
3. 27. Goldstein JA, Neff JM, Lane JM, Koplan JP. Smallpox vaccination reactions, prophylaxis, and therapy of complications. Pediatrics 1975;55:342-7. 28. Redfield RR, Wright DC, James WD, Jones TS, Brown C, Burke OS. Disseminated vaccinia in a military recruit with human immunodeficiency virus (HIV) disease. N Engl J Med 1987;316:673-6. 29. Picard 0, Giral P, Defer MC, et al. AIDS vaccine therapy: phase I trial. Lancet 1990;336: 179. 30. Guillaume JC, Saiag P, Wechsler J, Lescs MC, Roujeau Jc. Vaccinia from recombinant virus expressing HIV genes. Lancet 1991;337: 1034-5. 31. Buller RML. Smith GL, Cremer K. Notkins AL, Moss B. Decreased virulence of recombinant vaccinia virus expression vectors is associated with a thymidine kinase-negative phenotype. Nature 1985;317:813-5. 32. Taylor G, Stott EJ, Wertz G, Ball A. Comparison of the virulence of wild-type thymidine kinase (tk)-deficient and tk+ phenotypes ofvaccinia virus recombinants after intranasal inoculation of mice. J Gen ViroI1991;72:125-30. 33. Dolin R, Graham B, Greenberg SB, et al. Safety and immunogenicity of an HIV-I recombinant gpl60 candidate vaccine in humans. Ann Intern Med 1991;114:119-27. 34. Hu SL, Klaniecki J, Dykers T. Sridhar P, Travis BM. Neutralizing antibodies against HIV-I BRU and SF-2 isolates generated in mice expressing HIV -I envelope glycoproteins and boosted with gp 160. AIDS Res Hum Retroviruses 1991;7:615-20. 35. Hu SL, Abrams K Barber GN, et al. Protection of macaques against SIV infection by subunit vaccines of SIV envelope glycoprotein gp 160. Science 1992;255:456-9. 36. Graham BS, Belshe R, Clements ML, et al. HIV-gpI60 recombinant vaccinia vaccination of vaccinia-naive adults followed by rgp 160 booster immunization [abstract F.A.I.] In: Program and abstracts of the VII International Conference on AIDS (Florence. Italy). Rome: Instituto Superiore di Sanita, 1991.
Downloaded from http://jid.oxfordjournals.org/ at University of Iowa Libraries/Serials Acquisitions on June 5, 2016
6. Mackett M. Yilma T. Rose J K. Moss B. Vaccinia virus recombinants: expression of VSV genes and protective immunization of mice and cattle. Science 1985;227:433-5.
22.
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