Biomedical subjects

S Zolla-Pazner

Publications and source records attributed to S Zolla-Pazner.

At least 37 records · Page 2Linked to original sources

PubMed2000

A global neutralization resistance phenotype of human immunodeficiency virus type 1 is determined by distinct mechanisms mediating enhanced infectivity and conformational change of the envelope complex.

We have described previously genetic characterization of neutralization-resistant, high-infectivity, and neutralization-sensitive, low-infectivity mutants of human immunodeficiency virus type 1 (HIV-1) MN envelope. The distinct phenotypes of these clones are attributable to six mutations affecting functional interactions between the gp120 C4-V5 regions and the gp41 leucine zipper. In the present study we examined mechanisms responsible for the phenotypic differences between these envelopes using neutralization and immunofluorescence assays (IFA). Most monoclonal antibodies (MAbs) tested against gp120 epitopes (V3, CD4 binding site, and CD4-induced) were 20 to 100 times more efficient at neutralizing pseudovirus expressing sensitive rather than resistant envelope. By IFA cells expressing neutralization sensitive envelope bound MAbs to gp120 epitopes more, but gp41 epitopes less, than neutralization-resistant envelope. This binding difference appeared to reflect conformational change, since it did not correlate with the level of protein expression or gp120-gp41 dissociation. This conformational change was mostly attributable to one mutation, L544P, which contributes to neutralization resistance but not to infectivity enhancement. The V420I mutation, which contributes a major effect to both high infectivity and neutralization resistance, had no apparent effect on conformation. Notably, a conformation-dependent V3 neutralization epitope remained sensitive to neutralization and accessible to binding by MAbs on neutralization-resistant HIV-1 envelope. Sensitivity to sCD4 did not distinguish the clones, suggesting that the phenotypes may be related to post-CD4-binding effects. The results demonstrate that neutralization resistance can be determined by distinguishable effects of mutations, which cause changes in envelope conformation and/or function(s) related to infectivity. A conformation-dependent V3 epitope may be an important target for neutralization of resistant strains of HIV-1.

Antibodies, Monoclonal↗

PubMed1999

Exclusion of HIV coreceptors CXCR4, CCR5, and CCR3 from the HIV envelope.

Both HIV-1 primary isolates and laboratory strains incorporate cell-derived molecules into their envelopes depending on the host cell in which they are grown. This incorporation is not random and, specifically, HIV-1 has been shown to select against the incorporation into its surface of CD4, its main receptor. In this study, we have looked at the incorporation of HIV coreceptors CXCR4, CCR5, and CCR3 into the HIV envelope. For this purpose, we grew HIV-1 primary isolate BZ167 in several cell lines and PBMCs, and the envelope profiles of the resulting viruses were determined with a virus-binding ELISA. While the virus particle gained several molecules when passed through the different cell lines (e.g., ICAM-3, LFA-1, ICAM-1, or MHC class II), BZ167 never incorporated significant levels of CXCR4, CCR5, or CCR3 into its envelope even though some or all of the cell lines in which it was grown expressed them. These results show that HIV-1 selects against the incorporation of these chemokine receptors into its envelope molecule, as it does against the incorporation of CD4.

Cell Line↗

PubMed1999

Enhanced HIV type 1 neutralization by human anti-glycoprotein 120 monoclonal antibodies in the presence of monoclonal antibodies to lymphocyte function-associated molecule 1.

Cellular adhesion receptor LFA-1 and its ICAM ligands are known to play a role in HIV infection. The presence of these molecules on virions and target cells promotes virus infectivity and has previously been shown to hinder virus neutralization by anti-HIV antibodies. To delineate the effect of these molecules on neutralization of HIV-1, human monoclonal antibodies (MAbs) to V3 and the CD4-binding domain (CD4bd) of gp120 were examined in the presence of anti-LFA-1 MAbs. When either of two anti-LFA-1 MAbs was present, higher levels of virus neutralization were achieved by both anti-V3 and anti-CD4bd MAbs. This effect was observed with primary HIV-1 isolates as well as with a laboratory-adapted strain. However, this activity was seen only when an anti-LFA-1 MAb was combined with anti-gp120 MAbs that exhibited virus-specific neutralizing activities, demonstrating the specificity of both the anti-LFA-1 and anti-gp120 MAbs. Enhanced neutralization by anti-gp120 MAbs was observed if the anti-LFA-1 MAb was present during the initial 24 hr only, if added 24 hr after infection, or if present throughout the culture period. These data suggest that the anti-LFA-1 MAbs could act at different stages of HIV-1 infection, including the initial virus-cell interaction as well as during the amplification and spread of virus from cell to cell. These findings demonstrate the significant role of LFA-1 in HIV-1 infection and have important implications for evaluating the neutralizing activity of anti-HIV antibodies.

Antibodies, Monoclonal↗

PubMed1999

The implications of antigenic diversity for vaccine development.

The reactivity of human monoclonal and polyclonal anti-HIV-1 antibodies demonstrates that shared epitopes, including those that induce neutralizing antibodies, exist and are recognized by the human immune system. A priori, there is no reason why cross-clade neutralizing antibodies could not be induced by an appropriately constructed HIV vaccine. But to construct such a vaccine, it is critical to understand, as completely as possible, the antigenic structure of HIV, to establish and identify immunologic classifications for HIV, and to choose rationally the minimum number and types of viruses from these immunologic groupings that will induce the broadest protective responses.

AIDS Vaccines↗

PubMed1999

A longitudinal study of neutralizing antibodies and disease progression in HIV-1-infected subjects.

Sera from human immunodeficiency virus-1-infected participants in the Multicenter AIDS Cohort Study were tested to assess the association between serum neutralizing antibodies (NAbs) and disease progression. Each of 14 pairs, retrospectively matched for age, sex, race, and CD4+ lymphocyte numbers early in the study, consisted of a rapid progressor (RP) who developed AIDS and a long-term nonprogressor (LTNP) who remained asymptomatic. Serum samples were drawn early, when all participants were asymptomatic, and late, when the RPs had developed clinical AIDS. The LTNPs and RPs had similar levels of NAbs against primary isolates at the early time point, indicating that NAb levels are not predictive of disease progression; at the late time point, the LTNPs had significantly higher titers because of an increase in the level of serum NAbs in the LTNPs and/or a decrease in the NAbs in the RPs. The patterns of neutralizing activity over time suggest that changes in effective NAbs against different viruses do not occur in parallel.

Adult↗

PubMed1999

Generation of neutralizing human monoclonal antibodies against parvovirus B19 proteins.

Infections caused by human parvovirus B19 are known to be controlled mainly by neutralizing antibodies. To analyze the immune reaction against parvovirus B19 proteins, four cell lines secreting human immunoglobulin G monoclonal antibodies (MAbs) were generated from two healthy donors and one human immunodeficiency virus type 1-seropositive individual with high serum titers against parvovirus. One MAb is specific for nonstructural protein NS1 (MAb 1424), two MAbs are specific for the unique region of minor capsid protein VP1 (MAbs 1418-1 and 1418-16), and one MAb is directed to major capsid protein VP2 (MAb 860-55D). Two MAbs, 1418-1 and 1418-16, which were generated from the same individual have identity in the cDNA sequences encoding the variable domains, with the exception of four base pairs resulting in only one amino acid change in the light chain. The NS1- and VP1-specific MAbs interact with linear epitopes, whereas the recognized epitope in VP2 is conformational. The MAbs specific for the structural proteins display strong virus-neutralizing activity. The VP1- and VP2-specific MAbs have the capacity to neutralize 50% of infectious parvovirus B19 in vitro at 0.08 and 0.73 microgram/ml, respectively, demonstrating the importance of such antibodies in the clearance of B19 viremia. The NS1-specific MAb mediated weak neutralizing activity and required 47.7 micrograms/ml for 50% neutralization. The human MAbs with potent neutralizing activity could be used for immunotherapy of chronically B19 virus-infected individuals and acutely infected pregnant women. Furthermore, the knowledge gained regarding epitopes which induce strongly neutralizing antibodies may be important for vaccine development.

Amino Acid Sequence↗

PubMed1999

Immunotyping of human immunodeficiency virus type 1 (HIV): an approach to immunologic classification of HIV.

Because immunologic classification of human immunodeficiency virus type 1 (HIV) might be more relevant than genotypic classification for designing polyvalent vaccines, studies were undertaken to determine whether immunologically defined groups of HIV ("immunotypes") could be identified. For these experiments, the V3 region of the 120-kDa envelope glycoprotein (gp120) was chosen for study. Although antibodies (Abs) to V3 may not play a major protective role in preventing HIV infection, identification of a limited number of immunologically defined structures in this extremely variable region would set a precedent supporting the hypothesis that, despite its diversity, the HIV family, like the V3 region, might be divisible into immunotypes. Consequently, the immunochemical reactivities of 1,176 combinations of human anti-V3 monoclonal Abs (MAbs) and V3 peptides, derived from viruses of several clades, were studied. Extensive cross-clade reactivity was observed. The patterns of reactivities of 21 MAbs with 50 peptides from clades A through H were then analyzed by a multivariate statistical technique. To test the validity of the mathematical approach, a cluster analysis of the 21 MAbs was performed. Five groups were identified, and these MAb clusters corresponded to classifications of these same MAbs based on the epitopes which they recognize. The concordance between the MAb clusters identified by mathematical analysis and by their specificities supports the validity of the mathematical approach. Therefore, the same mathematical technique was used to identify clusters within the 50 peptides. Seven groups of peptides, each containing peptides from more than one clade, were defined. Inspection of the amino acid sequences of the peptides in each of the mathematically defined peptide clusters revealed unique "signature sequences" that suggest structural motifs characteristic of each V3-based immunotype. The results suggest that cluster analysis of immunologic data can define immunotypes of HIV. These immunotypes are distinct from genotypic classifications. The methods described pave the way for identification of immunotypes defined by immunochemical and neutralization data generated with anti-HIV Env MAbs and intact, viable HIV virions.

Antibodies, Monoclonal↗

PubMed1998

Neutralizing antibodies directed against the V3 loop select for different escape variants in a virus with mutated reverse transcriptase (M184V) than in wild-type human immunodeficiency virus type 1.

The M184V substitution in human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) encodes high-level resistance to the (-)-enantiomer of 2',3'-dideoxy-3'-thiacytidine (3TC) and low-level resistance to each of 2',3'-dideoxycytidine (ddC) and 2',3'-dideoxyinosine (ddI). This mutation also results in decreased HIV replication fitness in primary cells, diminished RT processivity, and increased RT fidelity. To assess the effect of this substitution on genetic variation in the HIV env region, we cultured both M184V-containing and wild-type BH10 in MT-4 cells in the presence of the neutralizing monoclonal antibody 447-52D, targeted to the GPGR epitope within the V3 loop of gp120. Outgrowth of viruses resistant to neutralization was followed by sequence analysis of the V3 loop by standard methodology. Wild-type HIV first showed escape after 15-22 days in culture. Sequence analysis revealed an arginine-to-lysine change within the GPGR epitope in the V3 loop (R20K, AGA --> AAA) in six of six clones sequenced after day 36. In contrast, M184V-containing HIV first showed escape between days 25 and 32 and sequence analysis revealed an aspartate-to-tyrosine change at amino acid 5 in V3 (N5Y; AAC --> TAC) in two of six clones at day 36 and in five of five clones at day 55. Similar results were obtained in two independent antibody selection protocols. The escape mutation in the wild type is consistent with the G --> A hypermutation observed in wild-type HIV-1, recently shown to cause an initial M184I change (before M184V) in 3TC-treated patients. In contrast, the N5Y substitution seen with M184V-containing HIV-1 is an A --> T transversion in V3.

Amino Acid Sequence↗

PubMed1998

A human monoclonal antibody specific for the V3 loop of HIV type 1 clade E cross-reacts with other HIV type 1 clades.

To ascertain the antigenic relationship between HIV-1 viruses belonging to various genetically defined subgroups (clades), shared epitopes need to be defined. Human monoclonal antibodies (MAbs) are particularly useful for this purpose because they can detect complex regions of viral proteins that may be missed by sequence analysis and because, by definition, they react with epitopes that stimulate the human immune system. Monoclonal antibodies derived from the cells of HIV-1 clade B-infected subjects have been used extensively for this purpose. Here we describe the first human MAb derived from a clade E-infected individual; the MAb is specific for the V3 loop, recognizing a core epitope represented by the amino acids TRTSVR on the N-terminal side of the crown of the V3 loop. The IgG1(kappa) MAb, designated 1324E, binds to the clade E consensus V3 loop, to rgp120 proteins from clade E and to peripheral blood mononuclear cells infected in vitro with the virus that infected the subject from whose cells the MAb-producing heterohybridoma was derived. Strong cross-reactivity of the MAb to the V3 peptides, rgp120 proteins, and native monomeric gp120s representing clades A and C, as well as to cells infected with a clade C primary isolate, revealed a shared V3 epitope between these clades. When tested for its neutralizing ability, MAb 1324E neutralized a clade E isolate that had been adapted for growth in H9 cells but failed to neutralize five clade E primary isolates.

Amino Acid Sequence↗

PubMed1998

Early-stage HIV infection and hepatitis C virus infection are associated with elevated serum porphyrin levels.

BACKGROUND: Porphyria cutanea tarda is known to be associated with HIV infection and hepatitis C virus (HCV). OBJECTIVE: Our purpose was to evaluate whether early infection with HIV, with or without HCV infection, is associated with elevated serum porphyrin levels. METHODS: Serum porphyrin levels were measured in samples obtained from 103 patients with early HIV infection. The results were compared with those of 89 late-stage HIV-positive patients and 78 HIV-negative patients. RESULTS: The highest median porphyrin level was in early-stage HIV-positive/HCV-positive samples, followed in decreasing order by those in early-stage HIV-positive/HCV-negative, late-stage HIV-positive/HCV-positive, late-stage HIV-positive/HCV-negative, HIV-negative/HCV-positive, and HIV-negative/HCV-negative groups. Elevated porphyrin levels were independently associated with early-stage HIV infection (P < .0001) and HCV infection (P = .03). CONCLUSION: This finding suggests abnormal porphyrin metabolism is most noticeable in early-stage HIV infection; it becomes less severe with the progression of HIV disease.

Adult↗

PubMed1998

Delineation of human antibody responses to culture filtrate antigens of Mycobacterium tuberculosis.

This study was undertaken to define the antigens in culture filtrates of actively replicating Mycobacterium tuberculosis that are recognized by antibodies from tuberculosis (TB) patients. Two-dimensional Western blots were probed with sera from healthy controls and TB patients that were preabsorbed with Escherichia coli lysates to deplete cross-reactive antibodies. Antibodies from TB patients recognized 26 of the >100 culture filtrate proteins, and the repertoire changed with disease progression. Only 12 of 26 antigens, including 3 proteins implicated in colonization and invasion by mycobacteria (MPT51, MPT32, and 85C), and 9 (as yet undefined proteins) were reactive with sera from TB patients with early noncavitary or cavitary disease. Eight additional antigens, including 4 undefined proteins, were recognized only by sera from a subset of patients with advanced cavitary disease. Studies suggest that 3 of the antigens recognized by sera from patients with early TB (85C, MPT32, and a 88-kDa protein) have strong serodiagnostic potential.

Antibodies, Bacterial↗

PubMed1998

Neutralization of syncytium-inducing primary isolates by sera from human immunodeficiency virus (HIV)-uninfected recipients of candidate HIV vaccines.

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.

AIDS Vaccines↗

PubMed1998

Passive immunization with a human immunodeficiency virus type 1-neutralizing monoclonal antibody in Hu-PBL-SCID mice: isolation of a neutralization escape variant.

A monoclonal antibody (694/98-D) directed toward the V3 loop of human immunodeficiency virus type 1 (HIV-1) was evaluated for pre- and postexposure prophylaxis in SCID mice reconstituted with human peripheral blood lymphocytes (Hu-PBL-SCID). Fifty percent protection against the HIV-1LAI strain was obtained by preexposure administration of 1.32 mg/kg antibody. However, virus isolated from 1 mouse 3 weeks after passive immunization with 13.2 mg/kg antibody proved resistant to subsequent in vitro neutralization by 694/98-D. V3 loop sequence analysis of cloned virus revealed amino acid changes within the linear core epitope recognized by 694/98-D and in one flanking amino acid. Further evaluation of 694/98-D for postexposure prophylaxis in mice revealed that 694/ 98-D was effective when given 15 min after virus. However, efficacy declined to 50% if treatment was delayed to 1 h after virus inoculation. These studies point out some potential drawbacks of passive immunization with monoclonal antibodies.

Amino Acid Sequence↗

PubMed1998

Immune responses to human immunodeficiency virus (HIV) type 1 induced by canarypox expressing HIV-1MN gp120, HIV-1SF2 recombinant gp120, or both vaccines in seronegative adults. NIAID AIDS Vaccine Evaluation Group.

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) TCID50 of canarypox (ALVAC) vector expressing HIV-1MN gp160 or 10(5.5) TCID50 of ALVAC-rabies virus glycoprotein control at 0 and 1 or 2 months and ALVAC-gp160 or 50 microg of HIV-1SF2 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-1MN and HIV-1SF2 neutralizing antibodies were detected more often (100%) in ALVAC-gp160/rgp120 recipients than in recipients of ALVAC-gp160 (<65%) 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 Vaccines↗

PubMed1998

Mapping of epitopes exposed on intact human immunodeficiency virus type 1 (HIV-1) virions: a new strategy for studying the immunologic relatedness of HIV-1.

To study the antigenic conservation of epitopes of human immunodeficiency virus type 1 (HIV-1) isolates of different clades, the abilities of human anti-HIV-1 gp120 and gp41 monoclonal antibodies (MAbs) to bind to intact HIV-1 virions were determined by a newly developed virus-binding assay. Eighteen human anti-HIV MAbs, which were directed at the V2, V3 loop, CD4-binding domain (CD4bd), C5, or gp41 regions, were used. Nine HIV-1 isolates from clades A, B, D, F, G, and H were used. Microtiter wells were coated with the MAbs, after which virus was added. Bound virus was detected after lysis by testing for p24 antigen with a noncommercial p24 enzyme-linked immunosorbent assay. The anti-V3 MAbs strongly bound the four clade B viruses and viruses from the non-B clades, although binding was weaker and more sporadic with the latter. The degrees of binding by the anti-V3 MAbs to CXCR4- and CCR5-tropic viruses were similar, suggesting that the V3 loops of these two categories of viruses are similarly exposed. The anti-C5 MAbs bound isolates of clades A, B, and D. Only weak and sporadic binding of all the viruses tested with anti-CD4bd, anti-V2, and anti-gp41 MAbs was detected. These results suggest that V3 and C5 structures are shared and well exposed on intact virions of different clades compared to the CD4bd, V2, and gp41 regions.

Antibodies, Monoclonal↗

PubMed1998

Vaccine protection against a heterologous, non-syncytium-inducing, primary human immunodeficiency virus.

Vaccine-induced protection of chimpanzees against laboratory-adapted and syncytium-inducing, multiply passaged primary human immunodeficiency virus type 1 (HIV-1) isolates, but not against non-syncytium-inducing, minimally passaged ones, has been demonstrated. Following challenge with such an isolate, HIV-15016, we obtained complete protection in one of three chimpanzees previously protected against low- and high-dose HIV-1SF2 exposures after immunization with an adenovirus-HIV-1MN gp160 priming-HIV-1SF2 gp120 boosting regimen. At challenge, the protected chimpanzee exhibited broad humoral immunity, including neutralizing antibody activity. These results demonstrate the potential of this combination vaccine strategy and suggest that vaccine protection against an HIV isolate relevant to infection of people is feasible.

AIDS Vaccines↗

PubMed1998

Induction of neutralizing antibodies to T-cell line-adapted and primary human immunodeficiency virus type 1 isolates with a prime-boost vaccine regimen in chimpanzees.

Five chimpanzees were immunized by administration of one or more intranasal priming doses of one to three recombinant adenoviruses containing a gp160 insert from human immunodeficiency virus type 1 (HIV-1) MN (HIV-1MN) followed by one or more boosts of recombinant HIV-1SF2 gp120 delivered intramuscularly with MF59 adjuvant. This regimen resulted in humoral immune responses in three of five animals. Humoral responses included immunochemically active anti-H1V-1 antibodies (Abs) directed to recombinant gp120 and neutralizing Abs reactive with T-cell-line-adapted HIV-1MN and HIV-1SF2. In addition, neutralizing activity was detected to the two homologous primary isolates and to two of three heterologous primary isolates which, like the immunizing strains, can use CXCR4 as a coreceptor for infection. The three animals with detectable neutralizing Abs and a fourth exhibiting the best cytotoxic T-lymphocyte response were protected from a low-dose intravenous challenge with a cell-free HIV-1SF2 primary isolate administered 4 weeks after the last boost. Animals were rested for 46 weeks and then rechallenged, without a boost, with an eightfold-higher challenge dose of HIV-1SF2. The three animals with persistent neutralizing Abs were again protected. These data show that a strong, long-lived protective Ab response can be induced with a prime-boost regimen in chimpanzees. The data suggest that in chimpanzees, the presence of neutralizing Abs correlates with protection for animals challenged intravenously with a high dose of a homologous strain of HIV-1, and they demonstrate for the first time the induction of neutralizing Abs to homologous and heterologous primary isolates.

Animals↗

PubMed1998

Coreceptor utilization by human immunodeficiency virus type 1 is not a primary determinant of neutralization sensitivity.

We have examined the relationship between coreceptor utilization and sensitivity to neutralization in a primary isolate of human immunodeficiency virus type 1 and its T-cell line-adapted (TCLA) derivative. We determined that adaptation of the primary-isolate (PI) virus 168P results in the loss of the unique capacity of PI viruses to utilize the CCR5 coreceptor and in the acquisition by the TCLA 168C virus of sensitivity to neutralization by V3-directed monoclonal antibodies (MAbs). In experiments wherein infection by 168P is directed via either the CCR5 or the CXCR4 pathway, we demonstrate that the virus, as well as pseudotyped virions bearing a molecularly cloned 168P envelope protein, remains refractory to neutralization by MAbs 257-D, 268-D, and 50.1 regardless of the coreceptor utilized. This study suggests that coreceptor utilization is not a primary determinant of differential neutralization sensitivity in PI and TCLA viruses.

Animals↗