A live, cold-passaged (cp) candidate vaccine virus, designated respiratory syncytial virus (RSV) B1 cp-52͞ 2B5 (cp-52), replicated efficiently in Vero cells, but was found to be overattenuated for RSV-seronegative infants and children. Sequence analysis of reverse-transcription-PCRamplified fragments of this mutant revealed a large deletion spanning most of the coding sequences for the small hydrophobic (SH) and attachment (G) proteins. Northern blot analysis of cp-52 detected multiple unique read-through mRNAs containing SH and G sequences, consistent with a deletion mutation spanning the SH:G gene junction. Immunological studies confirmed that an intact G glycoprotein was not produced by the cp-52 virus. Nonetheless, cp-52 was infectious and replicated to high titer in tissue culture despite the absence of the viral surface SH and G glycoproteins. Thus, our characterization of this negative-strand RNA virus identified a novel replication-competent deletion mutant lacking two of its three surface glycoproteins. The requirement of SH and G for efficient replication in vivo suggests that selective deletion of one or both of these RSV genes may provide an alternative or additive strategy for developing an optimally attenuated vaccine candidate.Respiratory syncytial virus (RSV), the leading cause of severe viral respiratory illness in pediatric populations throughout the world (reviewed in ref. 1), accounts for approximately 90,000 hospitalizations in infants and children in the United States each year (2). The importance of RSV as a respiratory pathogen makes development of a safe and effective RSV vaccine a public health priority (3). Although a number of approaches to RSV vaccine development have been taken, live RSV vaccines may provide the best alternative for immunizing young infants, because a live vaccine would mimic natural infection, induce a balanced cellular and humoral immune response, and be unlikely to produce enhanced disease (4).RSV exists as two antigenically distinct subgroups, A and B, and both RSV A and RSV B infections are capable of inducing severe lower respiratory tract disease (5-7). For this reason, a bivalent live RSV vaccine containing attenuated RSV A and RSV B components would be most desirable. Recently, a live attenuated RSV A candidate vaccine has been identified that appears to be safe and immunogenic in infants and children over 6 months of age (8). In addition, a cold-passaged (cp) RSV B candidate vaccine, designated RSV B1 cp-52͞2B5 (cp-52), was derived by passage of the RSV B1 wild-type (wt) virus 52 times at low temperature (21-32°C) (9). Cp-52 was shown to be restricted in replication in vivo but still able to induce RSV serum-neutralizing antibody responses in cotton rats, African green monkeys, and chimpanzees (9). Also, it was found to be phenotypically stable after prolonged replication in cotton rats (9). Here, we describe the phase I evaluation of the cp-52 candidate vaccine in adults, children, and infants. Although this virus mutant grew to high titer (Ͼ10 7....

Two live-attenuated, cold-passaged (cp), temperature-sensitive (ts) candidate vaccines, designated cpts530/1009 and cpts248/955, were attenuated, genetically stable, and immunogenic in chimpanzees and were highly attenuated for human adults. In respiratory syncytial virus (RSV)-seropositive children, cpts530/1009 was more restricted in replication than cpts248/955. In seronegative children, 10(4) pfu of cpts248/955 was insufficiently attenuated, and a high titer of vaccine virus was shed (mean peak titer, 10(4.4) pfu/mL), whereas 10(4) pfu of cpts530/1009 was relatively attenuated and restricted in replication (mean peak titer, 10(2.0) pfu/mL). At a dose of 10(5) pfu, cpts530/1009 was immunogenic in seronegative children (geometric mean titer of RSV neutralizing antibodies, 1:724). Transmission of either vaccine to seronegative placebo recipients occurred at a frequency of 20%-25%. Of importance, vaccine viruses recovered from chimpanzees and humans were ts. In contrast to previous studies, this study indicates that live attenuated RSV vaccines that are immunogenic and phenotypically stable can be developed. Additional studies are being conducted to identify a live RSV vaccine that is slightly more attenuated and less transmissible than cpts530/1009.

Induction of CD8+ cytotoxic T cells is considered one of the important correlates for the protective efficacy of candidate human immunodeficiency virus type 1 (HIV-1) vaccines. To induce CD8+ cytotoxic T lymphocytes (CTLs) along with neutralizing antibody and CD4+ T cell help, a live canarypox virus construct expressing gp120, transmembrane gp41, the gag and protease genes, and sequences containing CTL epitopes in nef and pol was given simultaneously with, or followed by, rgp120 SF2. CD8+ CTLs were detected in 61% of volunteers at some time during the trial. Three to 6 months after the last immunization, the gene-specific responses were gag, 26/81; env, 17/77; nef, 12/77; and pol, 3/16. Simultaneous immunization with the canarypox vector and the subunit, beginning with the initial immunization, resulted in earlier antibody responses. In summary, a strategy of immunization with a canarypox vector expressing multiple genes of HIV-1 given with gp120 results in durable CD8+ CTL responses to a broad range of epitopes.

A safety and immunogenicity trial was conducted in vaccinia-immune and vaccinia-naive human immunodeficiency virus (HIV) -uninfected adults who were randomized to receive 10 6 or 10 7 TCID 50 of canarypox (ALVAC) vector expressing HIV-1 MN gp160 or 10 5.5 TCID 50 of ALVAC -rabies virus glycoprotein control at 0 and 1 or 2 months and ALVAC-gp160 or 50 mg of HIV-1 SF2 recombinant (r) gp120 in microfluidized emulsion at 9 and 12 months; others received rgp120 at 0, 1, 6, and 12 months. All vaccines were well-tolerated. Neither vaccinia-immune status before vaccination nor ALVAC dose affected HIV immune responses. HIV-1 MN and HIV-1 SF2 neutralizing antibodies were detected more often (100%) in ALVAC-gp160/rgp120 recipients than in recipients of ALVAC-gp160 (õ65%) or rgp120 (89%) alone. ALVAC-gp160/rgp120 also elicited more frequent HIV V3 -specific and fusion-inhibition antibodies, antibody-dependent cellular cytotoxicity, lymphoproliferation, and cytotoxic CD8 / T cell activity than did either vaccine alone. Trials with ALVAC expressing additional HIV components and rgp120 are underway. AIDS, caused by infection with the human immunodefi-duce the transmission of HIV-1 infection and to prevent or ameliorate disease are needed, especially for developing counciency virus type 1 (HIV-1), is associated with enormous mortries, where ú90% of infections occur [1] and where antiretbidity and mortality worldwide. Safe, effective vaccines to reroviral drugs are not affordable.To prevent chronic HIV-1 infection, it is thought that a vaccine To circumvent the safety issues associated with vaccinia virusThe study protocol was reviewed and approved by the institutional review boards at each site. Informed consent was obtained from each volunteer in recombinants, an avipox vector, canarypox virus (referred to as accordance with guidelines of the US Department of Health and Human Ser-ALVAC), has been used to express genes encoding human vacvices and those of the authors' institutions.cine antigens, such as rabies virus glycoprotein, measles virus

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Most candidate human immunodeficiency virus (HIV)-1 vaccines induce antibodies that neutralize T cell line-adapted HIV-1 strains. Until recently, however, no neutralizing activity against primary HIV-1 isolates had been demonstrated in sera from human vaccinees. Since most candidate HIV-1 vaccines have been constructed from T cell line-adapted syncytium-inducing (SI) strains, experiments were done to test whether sera from recipients of SI-based vaccines could preferentially neutralize SI primary HIV-1 isolates. Various neutralization assays were performed with sera from volunteers receiving ALVACgp160MN and/or rgp120SF2. Neutralizing activity was detected against 4 of 8 SI primary isolates but against none of 5 non-SI primary isolates. The data suggest that, for the induction of neutralizing antibodies to a broad array of HIV-1 primary isolates, a polyvalent vaccine will be needed containing representatives of more than a single category of viruses.