© 1995 Nature Publishing Group http://www.nature.com/naturemedicine .. .... ARTICLES HIV-1 recombinant poxvirus vaccine induces cross-protection against HIV-2 challenge in rhesus macaques 1 2 ALASH'LE G. ABIMIKU , GENOVEFFA FRANCHINI', }AMES TARTAGLIA , KRISTINE ALDRICH', 1 3 4 MARIA MYAGKIKH , PHILLIP D. MARKHAM , PELE CHONG , MICHEL KLEIN\ 5 2 1 1 MARIE-PAULE KIENY , ENZO PAOLETTI , ROBERT C. GALL0 & MARJORIE ROBERT-GUROFF 'Laboratory of Tumor Cell Biology, Building 37, Room 6A09, 37 Convent Drive MSC 4255. National Cancer Institute, National Institutes ofHealth, Bethesda, Maryland 20892-4255, USA ' Virogenetics Corporation, 465 Jordan Road, Rensselaer Technology Park, Troy, New York 12180, USA 'Advanced BioScience Laboratories, Inc., 5510 Nicholson Lane, Kensington, Maryland 20895, USA ' Connaught Laboratories, 1755 Steeles Avenue West, Willowdale, Ontario, M2R 3T4, Canada 5Transgene, 11 rue de Molsheim, 67082 Strasbourg, France Correspondence should be addressed to M.R.G. Rhesus macaques were immunized with attenuated vaccinia or canarypox human Im­ munodeficiency virus type 1 (HIV-1) recombinants and boosted with HIV-1 protein sub­ units formulated In alum. Following challenge with HIV-2, •.....,, three out of eight immunized macaques resisted infection for six months and another exhibited signifi­ cantly delayed Infection, whereas all three naive controls became infected. Immuniza­ tions elicited both humoral and cellular immune responses; however, no clear correlates of protection were discerned. Although more extensive studies are now called for, this first demonstration of cross-protection between HIV-1 and -2 suggests that viral variabil­ ity may not be an Insurmountable problem In the design of a global AIDS vaccine. One of the main problems hindering HIV-1 vaccine develop­ function effectively as immunization vehicles... '0 (unpublished ment is the extensive variability of the virus. HIV-1 variants results). With respect to the use of these vectors in retrovirus not only are observed within and between infected individu­ vaccine development, an ALVAC recombinant expressing the als, but worldwide, eight distinct subtypes, A through H, have feline leukaemia virus (A subtype) Env and Gag proteins was been described'. A ninth grouping, subtype 0, is composed of shown to afford protection in cats against the development of even more highly divergent members. As a result of this het­ persistent viraemia when challenged with virus simultane­ 11 erogeneity, design of an effective vaccine for even a geograph­ ously given in the mouth and sprayed on the nose • More­ ically restricted area is a daunting task. The problem is over, protocols consisting of an ALVAC-HIV-1MN gp160 compounded on a global scale, where tailoring of vaccines to recombinant followed by boosting with native envelope pro­ specific populations is a current strategy. tein (gp120) demonstrated the ability of the ALVAC-HIV-1 re­ Although HIV-2 is clearly related to HIV-1 structurally and combinant to prime HIV-1 specific humoral and cellular functionally, it nevertheless constitutes a different viral type. immune responses in Phase I human trials, even in individu­ The viruses exhibit limited homology, with only 40 to 60% als previously exposed to vaccinia 0.-L. Excler, oral presenta­ similarity in the amino acid sequences of the structural gene tion, Septieme colloque des "Cent Gardes", 1992). 2 products • Although less pathogenic and infectious than HIV- Lack of an affordable, readily available, and proven animal 1, HIV -2 also causes AIDS, albeit with a slower disease model for HIV-1 has greatly hindered vaccine research. The courseJ-'. HIV-2 is endemic in West Africa, and has become a chimpanzee is inappropriate on the first two counts, and the 12 growing public health problem. Therefore, our AIDS vaccine pig-tailed macaque (Macaca nemestrina) model , although studies have addressed not only HIV-1 but also HIV-2, in part promising, has yet to be proven reliable. For these reasons we as a model for HIV-1, but also as a legitimate candidate itself have chosen to use the rhesus macaque (M. mulatta) system. for vaccine development. This species has been shown to be susceptible to infection not Our investigations have focussed on attenuated poxvirus only by several simian immunodeficiency virus (SIV) strains recombinants as vaccine vehicles. NYV AC, derived from the but also by HIV-2 (ref. 13-16), thus allowing not only assess­ Copenhagen vaccinia virus vaccine strain, was attenuated by ment of HIV-2 vaccine efficacy, but also evaluation of HIV-1 the precise deletion of 18 open reading frames implicated in vaccine immunogenicity followed by heterologous cross-chal­ poxvirus virulence•·•. The naturally attenuated ALVAC is a ca­ lenge experiments. nary poxvirus vector system that is unable to productively In separate studies a lasting protection against HIV -2 infec­ replicate in non-avian species•·•. Despite their highly attenu­ tion of rhesus macaques (G. Franchini et al., manuscript sub­ ated phenotype, NYV AC and AL VAC have been shown to mitted) and human T-cellleukaemia/lymphoma virus type I NATIJRE MEDICINE, VOLUME I, NUMBER 4 APRIL 1995 321 © 1995 Nature Publishing Group http://www.nature.com/naturemedicine • ········ ARTICLES .... ...... ..... Weeks Post Inoculation D II il il il i I I I I I , I I I I I 10 20 30 40 50 60 70 80 90 100 110 120 130 20 ~ Rhesus 214L (NYVACgp160 + octamerlcV3) & '& ;!!. (") Q)N 1.5 t 15 <g2: ,...,~~ ~.p_ 1.0 10 -'< w.,o-'o~ 0 ~ · .o~ ~ <(- 0.5 - t 5 :> . I ~ (3300) 2000 <..:><0 w~ 1500 OJO ~~ -s· "'m """'"' 40 1000 8l ~ c · ~ i= 2: =~ = >-J: ::i- ~g<i 20 500 ~§1 Q) ·..: ~ zc:> t <~ & Rhesus 218L (NYVACgp120 + octamerlcV3) 20 ~ ;!!. Q)N 1.5 c. 15 cnO ~g~ =-~ ~.e~- 1.0 IT~ uJgo ~(f) .o~ <t- 0.5 t 5 2: i J: (3000) ~~ 1ooo . i 2000 C>j!! <)' 100 10 1500 · ~F2: = >-J: ~g <:i 40 1000 20 ~'§;: 500 <( ~ t - Recombinant Vector c::::> Octameric V3 ~Challenge : HIV-2 ssL6669 Fig. 1 Immune responses in rhesus macaques inoculated with NYVAC-HIV-1,,, gp160 or gp120, and boosted with octameric V3 peptide. Macaque 214L (top two panels) was immunized (closed arrows) with the NYVAC-HIV-1 "" gp160 recombinant, vP911, and macaque 218L (lower two panels) with the NYVAC-HIV-1 "" gp120 recombinant vP921. Both macaques were boosted (open arrows) with a mixed octameric V3 loop peptide preparation . The final boost with NYVAC recombinants consisted of 108 pfu for macaque 214L and 1 o• pfu for macaque 218L. cell-mediated immune responses marked with an asterisk (•) were obtained with HIV-2 antigens and targets. The time of challenge , with HIV-2 58 6669 is indicated by the large hatched arrow. (HTLV-1) infection of rabbits" has been demonstrated follow­ combinants and proteins for vaccinating rhesus macaques ing immunization with HIV-2 or HTLV-1 recombinant vaccine against HIV-2 (G. Franchini et al., manuscript submitted). At candidates constructed in either NYV AC or ALVAC vectors. To the conclusion of the immunogenicity studies, we evaluated pursue these promising vectors as HIV-1 candidate vaccines, the protection afforded by the immunization regimens against we first evaluated the immunogenicity of a number of NYV AC a heterologous cross challenge with HIV-2. and ALVAC recombinants carrying HIV-1 genes (gag, pol, env) As expected our results showed that the NYV AC and AL VAC followed by various viral proteins or peptides presented with recombinants primed both humoral and cellular immune re­ adjuvants in eight rhesus macaques. Our objectives were (1) to sponses to HIV-1. But remarkably, following live virus chal­ compare the priming ability of recombinants expressing both lenge SO% of the immunized animals resisted infection by HIV-1 Env proteins (gp120, gp41) versus simply the gpl20 HIV -2, while all three naive controls became infected. This moiety alone; (2) to evaluate recombinants carrying the full finding, although preliminary, suggests that viral variability complement of HIV-1 genes for viral structural proteins gag, may not necessarily preclude effective AIDS vaccine develop­ pol, and env, as well as those carrying just the env gene; (3) to ment. It should encourage further investigation of cross-pro­ compare directly NYVAC- and ALVAC-HIV-1 recombinants in tection in other heterologous systems as well as stimulate similar prime/ boost protocols; and (4) to compare these re­ research on immunologically conserved epitopes and poten­ sults with those obtained using NYV AC- and ALVAC-HIV-2 re- tially novel protective mechanisms. 322 NATURE MEDICINE, VOLUME 1, NUMBER 4 APRIL 1995 © 1995 Nature Publishing Group http://www.nature.com/naturemedicine • ··············ARTICLES Results tion (Fig. 2), two subunit booster inoculations were necessary lmmunogenicity of HIV-1 recombinants and boosts to elicit neutralizing antibody activity and also led to more Eight rhesus macaques were inoculated with NYV AC- or regularly detectable Env-specific T-cell proliferative responses ALVAC-HIV-1 recombinants and subsequently boosted with and cytolytic activities. Additional booster inoculations, ei­ peptides or HIV-1 proteins formulated in alum or with addi­ ther with subunit preparations or recombinants, had little ef­ tional ALVAC/NYVAC-HIV-1 recombinants (see Methods). fect on subsequent immune responses. Animals boosted with Macaques were bled at intervals and sera were assayed for anti­ native gp120 (Fig. 2) developed higher neutralizing antibody bodies to the entire HIV-1 envelope protein and the V3loop, a titres compared to those boosted with the octameric V3 pep­ 21 principal neutralizing determinant of the virus'.._ , and for tide (Fig. 1), although the latter macaques exhibited higher neutralizing antibodies. Peripheral blood mononuclear cells V3 loop binding titres, suggesting that native gp120 elicited (PBMC) were monitored for HIV-1-specific lymphocyte prolif­ neutralizing antibodies to epitopes in addition to the V3 loop. erative responses as well as cytolytic activities. The HIV-spe­ This is further indicated by the independent fluctuation of cific immune responses observed are in Figs 1-4.