Envelope Subunit Research Articles

Toward an HIV type 1 vaccine that generates potent, broadly cross-reactive neutralizing antibodies.

P ROGRESS IN T HE D EV EL OPM E NT of an effective vaccine for HIV-1 has been gauged in large part by the ability to induce measurable virus-specific CD8 1 cytotoxic T lymphocytes (CTLs) and neutralizing antibodies as lead correlates of a protective immune response. 1±3 Although there has been success in generating low-level CTLs in a subset of vaccinated volunteers in phase I clinical trials, little progress has been made in generating neutralizing antibodies. Part of the problem for neutralizing antibodies is the high degree of genetic variability in the HIV-1 surface gp120 and transm embrane gp41 glycoproteins. 7 Although conserved neutralization epitopes exist, the current repertoire of candidate HIV-1 vaccines fails to generate sufficient antibodies to them. Critical features of the structure of the oligomeric envelope glycoproteins as they exist in their native form on the virus present yet another level of complexity for immunogen design. Most candidate HIV-1 envelope subunit vaccines are based on monomeric gp120 from T cell line-adapted (TCLA) strains of virus. Those vaccines have generated neutralizing antibodies with variable and sometimes potent activity against the vaccine strain, low activity against heterologous TCLA strains, and poor activity against primary isolates. It is conceivable that this narrow specificity, particularly the inability to neutralize primary isolates, will greatly limit the effectiveness of the antibody component of a vaccine. Scientists have been divided on the issue of whether the weak neutralizing antibody response generated by candidate HIV-1 vaccines is a property of the vaccine or the assay. A major argument has been that current assays underestimate neutralization potency, an argument that stems mostly from the fact that primary isolates are generally less sensitive to neutralization than TCLA strains. A concentrated effort has been made to resolve the vaccine-versus-assay issue by focusing mainly on the assay. We now recognize that primary isolates are relatively insensitive to neutralization under a variety of conditions, 27±30 including those that take into account the differential use of CXCR4 and CCR5 as coreceptors. 31±33 These developments, combined with features revealed by the crystal structure of gp120 and gp41, are making it increasingly clear that the dichotomy in neutralization sensitivity between TCLA strains and primary isolates is unrelated to the assay and actually reflects fundamental differences in the structure of the oligomeric envelope glycoproteins of these two categories of virus. Thus, repeated passage of HIV-1 in T cell lines has the consequence of selecting virus variants on which the neutralization epitopes are readily accessible for efficient antibody binding. Some of these epitopes reside in the V3 cysteine±cysteine loop of gp120, which is a major target for neutralizing antibodies on TCLA strains but not primary isolates. The Division of AIDS, Preventive and Vaccine Branch of the National Institutes of Health (Bethesda, MD), convened a workshop in October of 1998 to discuss new strategies to improve the neutralizing antibody responses generated by candidate HIV-1 vaccines. The goals of the workshop were four-fold: (1) consider the advantages and disadvantages of using genetically engineered cell lines to assess a neutralizing antibody response, (2) examine evidence from passive immunization experiments showing that potent neutralization is required for protection, (3) review the structure of gp120 and gp41 revealed by X-ray crystallography as it relates to neutralization and (4) discuss the merits of novel immunization strategies in early phases of development. A summary of this workshop is presented here.

Therapeutic vaccines for pre venting AIDS: their use with HAART

The possibility of preventing or slowing progression to AIDS by therapeutic immunisation with HIV-1 vaccines continues to provoke interest. 1 Salk J Bretscher PA Salk PL Clerici M Shearer GM A strategy for prophylactic vaccination against HIV. Science. 1993; 260: 1270-1272 Crossref PubMed Scopus (183) Google Scholar The well-designed and well-executed clinical trial by Eric Sandstrom and colleagues in this issue of The Lancet and other recently published articles 2 Ratto-Kim S Sitz KV Garner RP et al. Repeated immunization with recombinant gp160 human immunodeficiency virus (HIV) envelope protein in early HIV-1 infection: evaluation of the cell proliferative response. J Infect Dis. 1999; 179: 337-344 Crossref PubMed Scopus (24) Google Scholar , 3 Tsoukas CM Raboud J Bernard NF et al. Active immunization of patients with HIV infection: a study of the effect of VaxSyn, a recombinant HIV envelope subunit vaccine, on progression of immune deficiency. AIDS Res Human Retrovir. 1998; 14: 483-490 Crossref PubMed Scopus (36) Google Scholar add to the growing body of knowledge about the prospects and limitations of this form of immune-based therapy for HIV-1 infection. These studies have confirmed that HIV-1 gp160 envelope-based vaccines are generally well tolerated and induce new humoral and T-cell responses to HIV-1 antigens that were not present at baseline. However, despite evidence of immune responses to the vaccine, the vaccination did not show benefit in terms of suppression of disease or progression to AIDS. By contrast with previously reported studies with this product, Sandstrom's team did find that significantly fewer patients receiving the gp160 recombinant HIV-1 vaccine showed a drop in their CD4-cell counts, and that a higher proportion of vaccine recipients had CD4-cell counts above their baseline value. Therapeutic immunisation with recombinant gp160 in HIV-1 infection: a randomised double-blind placebo-controlled trialTherapeutic immunisations with rgp160 have a modest effect on CD4-cell counts, but this treatment alone did not lead to clinical benefit when given in addition to best clinical practice at the time of the trial. Immunisation in conjunction with antiretroviral therapy was also effective, which strongly suggests that a combination with highly active therapy would improve the total effect. Full-Text PDF

Formation of mature egg envelope subunit proteins from their precursors (choriogenins) in the fish, Oryzias latipes: loss of partial C-terminal sequences of the choriogenins.

The inner layer of egg envelope of the medaka, Oryzias latipes, comprises two major groups of glycoprotein subunits, ZI-1,2 and ZI-3. Their precursor proteins, choriogenin H (Chg H) and choriogenin L (Chg L), respectively, are synthesized in spawning female liver. In the present study, the primary structures of the precursors and the corresponding mature subunits were compared by peptide mapping and amino acid sequencing to find what difference in their molecular structures is relevant to the assembly of the soluble precursors into the insoluble inner layer. The primary structures of the solubilized subunits were mostly identical to those of the respective precursors, but they lacked C-terminal partial sequences that their precursors possessed, namely, ZI-1,2 subunit was shorter than Chg H by 34 amino acid residues and ZI-3 was shorter than Chg L by 27 residues. In addition, a consensus amino acid sequence, Arg-Lys-X-Arg, was found at the putative cleavage sites in the C-terminal region of the precursors. It is conjectured that the truncation of the precursor proteins is prerequisite for formation of mature chorion subunit proteins and their assembly into chorion.

At least three epitopes are recognized by the human repertoire in the hepatitis B virus group a antigen inducing protection; possible consequences for seroprevention and serodiagnosis

Many recent investigations have shown that both HBV and anti-HBs antibodies coexist in the same patient, and HBV can be found in individuals with anti-HBc antibodies in the absence of immunologically detectable HBsAg. In most cases, mutant forms of HBV affecting the region of the envelope gene coding for the group a determinant recognized by human antibodies have been found. The nature of the group a determinant was revisited with an ELISA involving dissociated, but not alkylated, envelope subunits. No antibody recognizing a continuous epitope of the major S envelope protein could be found in humans; the full activity of human anti-HBs antibodies appeared to be focused on the discontinuous group a determinant. The immunological human repertoire against the HBsAg group a determinant was analysed by competitive inhibition of three mouse monoclonal antibodies (mAbs) selected as recognizing three distinct specificities on the group a determinant. Antibodies to specificities #1 and 3 were found in 52/70 anti-HBs human sera and generally predominated over specificity #2, which was lacking in some sera. The heterogeneity of the group a determinant suggested by these data argue for the use of more than one type of anti-HBs mAb for seroprevention of recurrence after liver transplantation and HBs serodiagnosis. Provided all three types of mAbs characterized here recognize HBV variants with mutations in the a determinant and are virus-neutralizing, it may be helpful, after association of such mAbs, to use them for diagnosis and to devise new immunotherapeutic strategies to prevent emergence of HBsAg escape mutants.

The third egg envelope subunit in fish: cDNA cloning and analysis, and gene expression.

The inner layer of the egg envelope of a teleost fish, the medaka, Oryzias latipes, consists of two major subunit groups, ZI-1,2 and ZI-3. On SDS-PAGE, the ZI-1,2 group presents three glycoprotein bands that were considered to be composed of a common polypeptide moiety derived from their precursor, choriogenin H (Chg H). ZI-3 is a single glycoprotein derived from the precursor, choriogenin L (Chg L). In the present study, a fraction of a novel subunit protein was found in the V8 protease digest of ZI-1.2 that was partially purified from oocyte envelopes. This protein fraction was not present in the purified precursor, Chg H. By RT-PCR employing the primers based on the amino acid sequence of this fraction, a cDNA for the novel subunit was amplified, and a full-length clone of the cDNA was obtained by screening a cDNA library constructed from the spawning female liver. The clone consisted of 2025 b.p. and contained an open reading frame encoding the novel protein of 634 amino acids. This protein included Pro-X-Y repeat sequences in two-fifths of the whole length from its N-terminus. Northern blot analysis revealed that the gene expression for this protein occurred in the liver but not in the ovary of spawning female fish. This protein is considered as the third major subunit of the inner layer of the egg envelope of medaka.

A recombinant prime, peptide boost vaccination strategy can focus the immune response on to more than one epitope even though these may not be immunodominant in the complex immunogen

Rhesus monkeys were successfully vaccinated using a strategy of priming with a candidate envelope subunit vaccine and boosting with synthetic peptides. Priming was carried out with recombinant HIV-1 SF2 envelope glycoprotein incorporated into ISCOMs, following the attachment of a lipid tail. Peptides, covalently linked to ISCOMs, representing linear sequences within the V2 and V3 regions, were used to boost functional antibodies to neutralizing epitopes in both of these regions. Injections with these peptide formulations substantially increased the titre of serum neutralizing antibodies from low or undetectable levels. In addition to completely neutralizing the homologous HIV-1 SF2 strain, these sera also neutralized the escape variant, HIV-1 SF13. However, no antibodies were boosted which could compete with human, neutralizing monoclonal antibodies recognising conformational epitopes. The peptides also boosted antibodies to a peptide whose sequence lies close to the V2 region neutralizing epitope but does not overlap with it. Importantly, the level of antibodies to an unrelated epitope associated with enhancement of HIV-1 SF13 continued to fall after the peptide boost. Successful protection against challenge with chimeric simian immunodeficiency virus expressing HIV-1 SF13 envelope glycoproteins (SHIV SF13) may be due to an increase in the ratio of neutralizing to enhancing antibodies by selectively boosting with peptides to critical neutralizing epitopes.

The critical N-linked glycan of murine leukemia virus envelope protein promotes both folding of the C-terminal domains of the precursor polyprotein and stability of the postcleavage envelope complex.

The infectivity of Friend ecotropic murine leukemia virus was previously shown to be highly sensitive to modification in its envelope protein (Env) at only one of the eight signals for N-linked glycan attachment, the fourth from the N terminus (gs4). In the present study, a set of six single-amino-acid substitutions in or near gs4 was used to determine the function of this region of Env and the role played by the glycan itself. One mutant that lacked the gs4 glycan was fully infectious, while one that retained this glycan was completely noninfectious, indicating that the gs4 glycan per se is not required for Env function. Infectivity correlated with the level of mature Env complex incorporated into virus particles, which was determined by the severity of defects in transport of the envelope precursor protein (gPrEnv) from the endoplasmic reticulum into the Golgi apparatus, in cleavage of gPrEnv into the two envelope subunits (the surface protein [SU] and the transmembrane protein [TM]), and in the association of SU with cellular membranes. All of the mutants induced the wild-type level of superinfection interference, indicating that the gs4 region mutations did not interfere with proper folding of the N-terminal domain of SU. These results suggest that the gs4 region mediates folding of the C-terminal domains of gPrEnv and stability of the interaction between SU and TM. Although the gs4 glycan was not essential for infectivity, processing of all mutant Envs lacking this glycan was significantly impaired, suggesting that efficient folding of gPrEnv requires a glycan at this position. The conservation of a glycosylation site homologous to gs4 across a broad range of retroviruses suggests that this sequence may play a similar role in many retroviral Envs.

Evolution of envelope-specific antibody responses in monkeys experimentally infected or immunized with simian immunodeficiency virus and its association with the development of protective immunity.

Previous studies of attenuated simian immunodeficiency virus (SIV) vaccines in rhesus macaques have demonstrated the development of broad protection against experimental challenge, indicating the potential for the production of highly effective immune responses to SIV antigens. However, the development of this protective immune status was found to be critically dependent on the length of time postvaccination with the attenuated virus strain, suggesting a necessary maturation of immune responses. In this study, the evolution of SIV envelope-specific antibodies in monkeys experimentally infected with various attenuated strains of SIV was characterized by using a comprehensive panel of serological assays to assess the progression of antibodies in longitudinal serum samples that indicate the development of protective immunity. In parallel studies, we also used the same panel of antibody assays to characterize the properties of SIV envelope-specific antibodies elicited by inactivated whole-virus and envelope subunit vaccines previously reported to be ineffective in producing protective immunity. The results of these studies demonstrate that the evolution of protective immunity in monkeys inoculated with attenuated strains of SIV is associated with a complex and lengthy maturation of antibody responses over the first 6 to 8 months postinoculation, as reflected in progressive changes in antibody conformational dependence and avidity properties. The establishment of long-term protective immunity at this time in general parallels the absence of further detectable changes in antibody responses and a maintenance of relatively constant antibody titer, avidity, conformational dependence, and the presence of neutralizing antibody for at least 2 years postinoculation. In contrast to the mature antibody responses elicited by the attenuated SIV vaccines, the whole-virus and envelope subunit vaccines in general elicited only immature antibody responses characterized by poor reactivity with native envelope proteins, low avidity, low conformational dependence, and the absence of neutralization activity against the challenge strain. Thus, these studies establish for the first time an association between the effectiveness of experimental vaccines and the capacity of the vaccine to produce a mature antibody response to SIV envelope proteins and further indicate that a combination of several antibody parameters (including titer, avidity, conformational dependence, and virus neutralization) are superior to any single antibody parameter as prognostic indicators to evaluate candidate AIDS vaccines.

Assembly of a rod-shaped chimera of a trimeric GCN4 zipper and the HIV-1 gp41 ectodomain expressed in Escherichia coli.

The HIV-1 envelope subunit gp41 plays a role in viral entry by initiating fusion of the viral and cellular membranes. A chimeric molecule was constructed centered on the ectodomain of gp41 without the fusion peptide, with a trimeric isoleucine zipper derived from GCN4 (pIIGCN4) on the N terminus and part of the trimeric coiled coil of the influenza virus hemagglutinin (HA) HA2 on the C terminus. The chimera pII-41-HA was overexpressed as inclusion bodies in bacteria and refolded to soluble aggregates that became monodisperse after treatment with protease. Either trypsin or proteinase K, used previously to define a protease-resistant core of recombinant gp41 [Lu, M., Blacklow, S. C. & Kim, P. S. (1995) Nat. Struct. Biol. 2, 1075-1082], removed about 20-30 residues from the center of gp41 and all or most of the HA2 segment. Evidence is presented that the resulting soluble chimera, retaining the pIIGCN4 coiled coil at the N terminus, is an oligomeric highly alpha-helical rod about 130 A long that crystallizes. The chimeric molecule is recognized by the Fab fragments of mAbs specific for folded gp41. A similar chimera was assembled from the two halves of the molecule expressed separately in different bacteria and refolded together. Crystals from the smallest chimera diffract x-rays to 2.6-A resolution.

Vaccine-specific antibody responses induced by HIV-1 envelope subunit vaccines.

The first generation of candidate vaccines to prevent HIV infection consisted of recombinant envelope proteins (Env, gp120, and gp160) derived from a single laboratory strain of HIV, designated IIIB/LAV, but produced with different expression systems. In this study we examined the fine specificity of the human Ab response to each vaccine and compared them to the responses of laboratory workers infected with the same strain of HIV. The best responders from each vaccine protocol were studied and compared. Detailed comparisons of the fine specificity of the Ab response were possible because all immunologic assays were performed using homologous recombinant proteins, peptides, and virus stocks. Although the total amounts of anti-Env Ab were comparable, the groups exhibited significant differences in epitope specificity, avidity, and functional capacity of the Ab response. The data demonstrate that the form of the immunogen (e.g., live virus or recombinant protein) is important in determining the quality of the Ab response. Conclusions are also drawn regarding characteristics of the anti-HIV-neutralizing Ab response. These studies represent one of the most detailed analyses of the human Ab response to any Ag and have implications for the development of vaccines for HIV as well as for other microbial pathogens.

A model for the maturation of protective antibody responses to SIV envelope proteins in experimentally immunized monkeys.

Studies using live attenuated virus vaccines in the simian immunodeficiency virus (SIV) rhesus macaque model have demonstrated broad protection against experimental challenge. Protection in these studies was found to be critically dependent on the length of time postvaccination, suggesting that protective immunity involves a necessary maturation of immune responses. The current study characterizes the evolution of protective envelope-specific antibody responses from monkeys inoculated with the highly attenuated SIV/17E-Cl virus vaccine. For comparison, the same antibody assays were used to characterize the properties of SIV envelope-specific antibodies elicited by inactivated whole virus and envelope subunit vaccines that failed to provide protection from experimental virus challenge. Results of these studies identify a continuous and complex maturation of envelope-specific antibody responses during the first six to eight months postinfection. Furthermore, the time required for maturation of SIV envelope-specific antibodies parallels the time required for the development of protective immunity against experimental challenge with heterologous strains of SIV. While no single immune correlate of protection has been identified, we suggest that a combination of antibody parameters may serve as prognostic indicators in the development of candidate AIDS vaccines.

Probing the Conformation of the Human T-Lymphotropic Virus I Envelope Protein Complex with Monoclonal Antibodies

We are investigating the binding of a series of monoclonal antibodies to native and detergent-treated human T-lymphotropic virus I (HTLV-I) envelope proteins to explore their conformation. A comparison of our data with previously published findings suggests that a central neutralization domain (aa 175-200) is folded such that only short stretches are exposed at the surface of the native envelope protein complex. However, the complete domain becomes accessible after treatment with mild non-ionic detergents, suggesting that envelope subunit interaction may partially obscure this domain. We further provide immunochemical evidence that a region containing a heptad repeat in the extracellular part of the transmembrane protein is folded towards the interior of the HTLV-I envelope complex.

Evaluation of solubilizing methods of the egg envelope of the fish, Oryzias latipes, and partial determination of amino acid sequence of its subunit protein, ZI-3

The inner layer of most teleostean egg envelopes, especially those after hardening, is almost insoluble in ordinary solvents, and therefore the inner layer of only the unhardened egg envelope has been subjected to solubilization with some potent solvents. We comparatively evaluated the methods of solubilization of the inner layer of egg envelope of medaka, Oryzias latipes, with SDS, urea and guanidium chloride (GuHCl). Analysis of the solubilized samples by SDS-polyacrylamide gel electrophoresis, comparison of their amino acid compositions or peptide maps using high-performance liquid chromatography and partial determination of their amino acid sequences showed that SDS and GuHCI were appropriate for solubilization and characterization of the envelope. Urea solubilization resulted in some artificial modifications of lysine and/or cysteine residues of envelope proteins. Partial determination of amino acid sequence of a subunit, ZI-3, isolated from the SDS-or GuHCl-solubilized envelope strongly suggested the identity of the envelope subunit, ZI-3, and its precursor, L-SF.

Immunization with Envelope Subunit Vaccine Products Elicits Neutralizing Antibodies against Laboratory-Adapted but Not Primary Isolates of Human Immunodeficiency Virus Type 1

Phase I studies of volunteers not infected with human immunodeficiency virus type 1 (HIV-1) have shown that immunization with envelope subunit vaccine products elicits antibodies that neutralize laboratory-adapted (prototype) HIV-1 strains in vitro. Prototype strains are adapted to grow in continuous (neoplastic) cell lines and are more susceptible to neutralization than are primary isolates cultured in human peripheral blood mononuclear cells. In this study, 50 sera from nine phase I vaccine trials and 16 from HIV-1-infected persons were evaluated for neutralizing antibody activity against 3 laboratory-adapted and 5 primary HIV-1 isolates. Of 50 sera, 49 neutralized at least 1 of the prototype strains; however, none displayed neutralizing activity against primary isolates of HIV-1. Serum from most HIV-1-infected persons neutralized both laboratory-adapted and primary HIV-1 isolates. These data demonstrate a qualitative, or large quantitative, difference in the neutralizing antibody response induced by envelope subunit vaccination and natural HIV-1 infection.

Lack of induction of antibodies specific for conserved, discontinuous epitopes of HIV-1 envelope glycoprotein by candidate AIDS vaccines.

We examined the humoral immune response in both HIV-1 infected and uninfected volunteers immunized with candidate HIV-1 recombinant envelope subunit vaccines (Genentech gp120IIIB, MicroGeneSys gp160IIIB, or ImmunoAG gp160IIIB). Immunization of both HIV-1 infected and uninfected volunteers with these immunogens resulted in the induction of Abs preferentially reactive with epitopes accessible on a denatured form of gp120. While sera from HIV-1 uninfected gp120/gp160IIIB vaccinees bound gp120/gp41, which was expressed on the surface of H9 cells infected with HIV-1IIIB, minimal binding to HIV-1MN or HIV-1RF infected cells was obtained. Induction of qualitatively similar immune responses by these immunogens would not have been predicted based on their different tertiary structures. These data indicate a restriction of the immune response to linear, conserved epitopes poorly accessible on both monomeric gp120 and cell-surface expressed oligomeric gp120/gp41 and a lack of Abs specific for conformational epitopes conserved across divergent HIV-1 strains. Poor recognition of HIV-1 envelope tertiary and quaternary structure may explain the restricted neutralization profiles of vaccinee sera against laboratory-adapted strains of HIV-1 and their inability to neutralize primary HIV-1 isolates. Alternate immunogens or reformulations with the capacity to elicit Abs that preferentially bind to natively folded gp120 should be investigated and correlated with their ability to neutralize more diverse laboratory-adapted and primary HIV-1 isolates.

Potential impact of low efficacy HIV-1 vaccines in populations with high rates of infection.

A safe and effective HIV vaccine to prevent infection and/or to moderate disease is urgently needed. Research progress has been slower than anticipated for a variety of reasons including uncertainty over which immunogen to use (i.e. recombinant subunit envelope proteins or whole HIV-1 products), confusion on which immunological markers best correlate with protection, the relevance of the HIV-1 chimpanzee model to infection in humans and the significance of the rapid evolution of HIV-1, with different clades of the virus emerging in different parts of the world. However, what some would interpret as encouraging results, from Phase I and II trials of recombinant envelope glycoprotein vaccines, have raised the question of whether the time is right to start Phase III trials in humans with immunogens that may have low to moderate efficacy. By using mathematical models and data from epidemiological studies, we examine the potential impact of such vaccines within heterosexual communities with high rates of infection. Analyses suggest that it will be difficult to block HIV-1 transmission even with very high levels of mass vaccination. The cost of sustaining high levels of herd immunity with a vaccine of short protection duration is likely to be high. However, assessments of impact over the long duration of an HIV-1 epidemic indicate that many cases of HIV infection and associated mortality can be prevented by immunogens with efficacy of 50% or less and a five year protection duration.(ABSTRACT TRUNCATED AT 250 WORDS)

Influence of transmembrane domains on the fusogenic abilities of human and murine leukemia retrovirus envelopes.

The envelopes of two highly divergent oncoviruses, human T-cell leukemia virus type 1 (HTLV-1) and Friend murine leukemia virus (F-MuLV), have distinct patterns of cellular receptor recognition, fusion, and syncytium formation. To analyze the influence of the transmembrane envelope subunit (TM) on fusogenic properties, we substituted either the entire TM or distinct domains from F-MuLV for the corresponding domains in the HTLV-1 envelope. Parental, chimeric, and truncated envelopes cloned into a eukaryotic expression vector were monitored for fusogenic potential in human, rat, and murine indicator cell lines by using a quantitative assay. This highly sensitive assay allowed us to assess the fusogenic properties and syncytium-forming abilities of the HTLV-1 envelope in murine NIH 3T3 cells. All chimeric envelopes containing extracellular sequences of the F-MuLV TM were blocked in their maturation process. Although deletions of the HTLV-1 cytoplasmic domain, alone and in combination with the membrane-spanning domain, did not prevent envelope cell surface expression, they impaired and suppressed fusogenic properties, respectively. In contrast, envelopes carrying substitutions of membrane-spanning and cytoplasmic domains were highly fusogenic. Our results indicate that these two domains in F-MuLV and HTLV-1 constitute structural entities with similar fusogenic properties. However, in the absence of a cytoplasmic domain, the F-MuLV membrane-spanning domain appeared to confer weaker fusogenic properties than the HTLV-1 membrane-spanning domain.

Synchronous Production of the Low- and High-Molecular-Weight Precursors of the Egg Envelope Subunits, in Response to Estrogen Administration in the Teleost Fish Oryzias latipes

Estrogen stimulation of production and accumulation of L-SF and H-SF, the low- and high-molecular-weight precursors of the egg envelope subunits in Oryzias latipes , was analyzed by immunohistochemical and immunoblotting methods. Both SF substances were detected in the liver of the male fish as quickly as 16-24 hr after estrogen treatment, and their production and accumulation in the liver and blood plasma increased as estrogen treatment continued. Immunohistochemical analysis revealed that the number of cells producing both SP substances was increased in the liver as estrogen treatment continued. These results strongly suggest the occurrence of synchronous production of L-SF and H-SF in response to estrogen. In liver extracts, but not in blood plasma, an additional L-SF-like immunoreactive protein was detected on immunoblots. Preliminary data indicate parallel appearance of vitellogenin and SF substances in the liver of estrogenized fish.

HIV vaccine development and clinical trials

The magnitude of HIV pandemic has made the development of HIV vaccines an urgent biomedical research priority. Although the biologic problems in designing a vaccine for a chronic viral infection like HIV are formidable, there has been encouraging progress. More than a dozen first generation prophylactic HIV vaccine candidates have completed phase I human trials that have established the safety and immunogenicity of these products in adults. A phase II trial of two HIV subunit envelope vaccines in adults at high risk of HIV infection is underway in the United States, and preparations for phase III efficacy trials have begun. Preliminary studies are under way to evaluate the potential application of active and passive immunization for preventing vertical transmission of HIV. Because of the higher rate of HIV transmission and a more abbreviated time course to disease, it may be more efficient to evaluate the efficacy of HIV vaccines in HIV infected pregnant women and their offspring than in adults who are exposed sexually to HIV.

Prevention of HIV/AIDS with vaccines

HIV vaccines are urgently needed to help control the relentlessly expanding epidemic. However, the development of a safe and effective vaccine for this rapidly mutating virus which produces life-long infection is elusive. Feasibility of HIV vaccination has been demonstrated by challenge studies in primates and by the presence of neutralizing antibodies which confer passive protection and can be detected in vitro. Given safety considerations, the first vaccines developed have been recombinant subunit vaccines consisting of viral envelope antigens formulated with adjuvants or incorporated into live vectors, such as vaccinia and avipox. Subunit envelope vaccines have proven to be safe and immunogenic in human phase I trials. Two gp120 subunit products are being considered for phase III trials based on ongoing evaluation of data from a phase II trial, primate-challenge studies, and laboratory studies evaluating the breadth, magnitude, and duration of immunogenic responses, including neutralization against recent isolates. Efficacy trials will be large and complex, involving people at high behavioral risk for infection. Baseline studies with recruitment goals of over 10000 people are being sponsored in the USA, with many additional vaccine preparedness studies being sponsored in developing nations by international and national authorities. Because high-risk populations will be recruited for trials of many year's duration, new approaches to trial design and community participation will be needed in order for these trials to succeed. Systems must be in place to ensure that trial participants receive preventive counseling and services to minimize discrimination from vaccine-related seroconversions. Here we document the urgent need for HIV vaccines, provide a brief preclinical and clinical update, and present the plans and considerations for testing HIV vaccine efficacy.