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Cellulose acetate 1,2-benzenedicarboxylate (CAP), a pharmaceutical excipient used for enteric film coating of capsules and tablets, was previously shown to have potent inhibitory activity against infection by human immunodeficiency virus type 1 (HIV-1) T cell line-adapted (TCLA) strains. In the present study, we determined the inhibitory activity of CAP against infection by cell-free and cell-associated primary HIV-1 isolates with distinct genotypes and biotypes in cervical explants, peripheral blood mononuclear cells (PBMCs), monocytederived macrophages (MDMs), and CEMx174 5.25M7 cells. CAP blocked infection by cell-free and cell-associated HIV-1 in cervical explants. It inhibited infection by cell-free primary HIV-1 isolates (clades A to G and group O) in PBMCs, MDMs, and CEMx174 5.25M7 cells and blocked transmissions of the cell-associated primary HIV-1 isolates from dendritic cells (DCs) to PBMCs, from MDMs to PBMCs, and from PBMCs to CEMx174 5.25M7 cells. The inhibitory activity of CAP on infection by the cell-free and cell-associated primary HIV-1 isolates is independent of viral subtypes and coreceptor usage. These data suggest that CAP is a good microbicide candidate that can be further developed for preventing sexual transmission of HIV-1.

A human monoclonal antibody (MAb) (2F5) neutralizing a variety of laboratory strains and clinical isolates of the human immunodeficiency virus type 1 (HIV-1) and binding to an epitope of the envelope glycoprotein gp41 encompassing the amino acid sequence ELDKWA has been described (Muster T et al., J Virol 1993;67:6642-6647). It was suggested that an immunogen eliciting virus-neutralizing antibodies having a specificity similar to that of MAb 2F5 should be considered as a component of HIV-1 vaccines. Efforts in this direction would benefit from understanding the mechanism whereby MAb 2F5 neutralizes the infectivity of HIV-1. The segment of gp41 encompassing residues ELDKWA has so far not been directly implicated in initiation of infection by HIV-1, suggesting that MAb 2F5 might affect other sites on HIV-1 envelope glycoproteins playing a role in virus entry into target cells. We provide here evidence that MAb 2F5 binding to HIV-1 virus particles decreases the accessibility or conformation of the gp41 fusion domain and of gp120 domains, including the binding site for the CD4 cell receptor. These apparently indirect consequences of MAb 2F5 binding to HIV-1 are likely to account for or contribute to the virus-neutralizing activity of this MAb.

Autoimmunity mediated by cross-reactive antibodies, elicited by HIV-1 envelope glycoproteins gp120/gp160, has been postulated to contribute to the pathogenesis of AIDS. Partial amino acid sequence homology between gp120/gp160 and several human host proteins, including MHC antigens and immunoglobulins, has been perceived as the basis for immunological cross-reactivity. Binding of antibodies from sera of HIV-1-infected individuals to selected host proteins and/or to synthetic peptides derived from them and the inhibitory activity of such sera in assays measuring the functional activity of T cells provided apparent support for the autoimmunity hypothesis, which is also relevant to the issue of safety of anti-HIV-1 vaccines. Considering the possibility that the detected autoantibodies may arise for reasons other than antibody responses to gp120/gp160, the immunological cross-reactivity between gp120/gp160 and the relevant host proteins was investigated using hyperimmune rabbit anti-gp120/gp160 and monoclonal antibodies. As determined from dilution end-point comparisons for polyclonal anti-gp120, the cross-reactivity of anti-gp120 with CD4 was undetectable (< 10(-5)%). The cross-reactivity of anti-gp120/gp160 with HLA-I and HLA-II antigens was also undetectable (< 4 x 10(-4)%) and that with other human proteins reported to have partial sequence homology with gp120/gp41 was < or = 0.013%. Anti-gp120/gp160 did not have detectable inhibitory effects in functional assays measuring proliferative T cell responses. Therefore, immunization with gp120/gp160 is unlikely to elicit harmful autoimmune responses.

Immunization of primates or humans with human immunodeficiency virus type 1 (HIV-1) glycoproteins usually elicited moderate immune responses to the principal neutralizing determinant (PND) located within the V3 hypervariable loop of gp120. Since an antibody response to the PND appears to be protective, experiments were carried out to determine the responsiveness of chimpanzees to immunization with synthetic peptides corresponding to the full-length V3 loop. Seven chimpanzees (4 preimmunized with gp160, 2 preimmunized with HIV-1 antigens unrelated to gp160, and 1 unimmunized) were vaccinated with a mixture of full-length V3 loop peptides from 21 distinct HIV-1 isolates (clones) either in unconjugated form or linked to carrier proteins from HIV-1 nef and gag P18, respectively. Six chimpanzees developed high levels of antibodies to the peptides (dilution endpoints 1: greater than 25,000), and 5 had high levels of antibodies to gp120 from HIV-1IIIB (endpoint titers 1: greater than 500,000). Chimpanzees immunized with peptide-carrier conjugates (4) had antibodies to the carrier proteins nef and gag P18, respectively (endpoint titers 1: greater than or equal to 35,000). Virus-neutralizing (VN) antibodies were detected in sera of 5 of 7 chimpanzees, but were present at titers of 1: greater than or equal to 400 only in sera of 2 chimpanzees. One of these was challenged with HIV-1 and was protected against infection, as reported elsewhere. The antibodies were primarily specific for the HIV-1 isolate used for primary immunization before boosting with peptides. The relatively low dilution endpoints of VN antibodies as compared with endpoints determined by site-specific immunoassays probably can be ascribed to imperfect mimicry of conformational epitopes by synthetic peptides. Nevertheless, sequential or simultaneous immunization with recombinant envelope glycoproteins of HIV-1 and selected synthetic peptides offers an approach for eliciting protective immunity against HIV-1.

The positions of all 9 intrachain disulfide bonds within the envelope glycoprotein gp120 of the human immunodeficiency virus (HIV-1) have been established recently. Peptides expected to mimic some of the disulfide-bonded domains [(120-133)-(203-221); (133-138)-(164-203); (224-254); (391-425) and (385-392)-(425-452)] were synthesized. All peptides, except (120-133)-(203-221), elicited in immunized rabbits relatively high levels of antibodies reacting with gp120 in enzyme-linked immunosorbent assay (ELISA) and/or Western immunoblot assays. However, these antibodies failed to neutralize the infectivity of HIV-1. Combined with earlier reports concerning other gp120 loop peptides, these results confirm the uniqueness of the V3 (303-338) loop in encompassing a principal determinant(s) involved in virus neutralization.

It is essential for the development of strategies for prevention and therapy of human immunodeficiency virus (HIV-1) infections to define host factors playing a dominant role in determining the clinical outcome of infection. Antibodies directed against restricted regions of the HIV-1 glycoproteins gp120 and gp41 are likely to represent important factors involved in host defense against HIV-1. Definition of qualitative and quantitative differences in the spectrum of anti-gp120 and anti-gp41 antibodies between two vastly different groups of HIV-1-infected individuals, long-term asymptomatic carriers, and individuals with acquired immunodeficiency syndrome (AIDS) who died, might reveal the epitope specificity of antibodies contributing to prevention of clinical disease. To accomplish this goal, sera from both groups were assayed for antibodies recognizing synthetic peptides from gp120/gp41 which were shown in earlier experiments to mimic epitopes on the two HIV-1 glycoproteins. None of the sera recognized all of the distinct 27 peptides from gp120 and gp41. The spectrum of antibodies was distinct for each of the sera from both groups of HIV-1-infected individuals. Nevertheless, antibody responses distinguishing the two groups from each other were discerned. In particular, it was possible to predict the unfavorable outcome of disease by comparative measurements of levels of antibodies to a peptide (303-338), corresponding to the entire V3 hypervariable loop of gp120 and/or by providing evidence for declining levels of these antibodies during the course of infection. Antibodies recognizing additional peptides [(219-245), (280-306), (425-452), (658-682), (729-758), (808-845), and (845-862)] were significantly less prevalent in AIDS patients than in asymptomatic carriers. It appears possible that maintenance of high levels of the respective antibodies would contribute to preventing AIDS in HIV-1-infected individuals. Active immunization with antigens containing epitopes defined by the respective peptides and/or administration of the corresponding antibodies may be considered as a modality for therapy of HIV-1 infections.

A solid phase radioimmunoassay (RIA) for human fibroblast-derived beta (beta) interferon is described using dinitrophenylated (DNP) IgG from rabbit anti-interferon serum and quantitation of the antigen-bound antibodies by subsequently measuring the attachment of 125I-labeled anti-DNP. The sensitivity limit of the assay is 50 u of interferon corresponding to 0.5 ng of interferon protein. Preparations of alpha leukocyte-derived interferon show only marginal cross-reactivity (less than 0.01%) while the mouse interferons (mixture of alpha and beta)are negative in this assay. The simplicity, reproducibility and the possibility of using 125I-labeled anti-DNP as a universal reagent for different anti-interferon sera may widen the development of RIA tests for interferons both in research and clinical areas.

The region of the hepatitis B virus (HBV) genome coding for the viral envelope proteins contains three inphase ATGs that are conserved among viral subtypes. Each of these ATGs can be used as mRNA initiation codons. The three translated proteins share a carboxy-terminal region (the S protein) and extend amino-terminally to include the pre-S2 region in the middle (M) protein, and the pre-S1 and pre-S2 regions in the large (L) protein. We have inserted the HBV DNA coding for the M protein into a baculovirus expression vector. Infected insect cells transcribe a mRNA that is initiated solely within a baculovirus promoter, and that contains the initiator codons for both M and S proteins. Although these cells primarily secrete the M protein, the major translational product is the S protein, which is not secreted. This preferential translation, the result of the use of an internal initiator codon, demonstrates that the regulation of HBV envelope protein production can occur at the translational level.