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J L Hurwitz

Publications and source records attributed to J L Hurwitz.

At least 37 records · Page 2Linked to original sources
PubMed1997

Effect of natural HIV-1 envelope V1-V2 sequence diversity on the binding of V3-specific and non-V3-specific antibodies.

In past years, much attention has been paid to the HIV-1 envelope (env) protein variable region 3 (V3), termed the principal neutralizing determinant. HIV-1 vaccines were often designed to target V3, and vaccine efficacy was often measured with V3-based assays. Thus, some disappointment resulted when volunteers in first clinical vaccine trials generated V3-specific antibodies that could not protect against V3-similar viruses. We describe an analysis of V1 and V2 sequence effects on antibody binding to V3 and non-V3 determinants. This study involved the preparation of seven full-length (gp160), chimeric env proteins in a vaccinia virus (VV) expression system. Chimeric proteins displayed different V1-V2 sequences but were otherwise identical. A panel of 12 monoclonal antibodies was then tested for binding activities toward the seven chimeras. Results showed that V1-V2 sequences affected antibody binding to env, both in V3 and non-V3 positions. These data demonstrate the enormous complexity of HIV-1 env protein conformation and antigenic determinants. Respect for the complexity of antibody-antigen interactions encourages the design of sophisticated immunoglobulin and protein cocktails for use in HIV-1 therapies and vaccines, respectively.

Amino Acid Sequence↗
PubMed1997

Screening of HIV-1 Env glycoproteins for the ability to raise neutralizing antibody using DNA immunization and recombinant vaccinia virus boosting.

HIV-1 envelopes from two series of primary isolates (from Swedish patients 5 and 6), from JR-FL and BaL (prototypic monocyte/macrophage tropic viruses) and from HXB-2 (a prototypic T-cell-line-adapted virus), have been screened for their ability to elicit neutralizing antibody to HIV-1. Rabbits were primed by gene gun inoculation with plasmids expressing secreted monomeric (gp120) and oligomeric (gp140) forms of each Env. After four to six DNA immunizations administered over a 1-year period, rabbits were boosted with 10(8) plaque-forming units of a mixture of seven recombinant vaccinia viruses which express chimeric gp140 Envs (primary clade B sequences in a IIIb-related BH10 backbone). Neutralizing antibodies were assayed against two T-cell-line-adapted viruses (MN and IIIb), two non-syncytium-inducing (NSI) and two syncytium-inducing (SI) primary isolates, and two HIV-1-NL4-3-recombinants with patients 5 or 6 Envs (NL4-3/5A, NL4-3/6C). The DNA priming and recombinant vaccinia virus boosting raised low titers of neutralizing antibody in 10 of 19 rabbits. The highest titers of neutralizing activity (approximately 1:150 for MN) were raised in rabbits DNA primed with Envs from Swedish patients 5. These sera cross neutralized IIIb and MN but did not neutralize the primary isolates or the NL4-3 recombinant with the homologous 5A Env. Sera from rabbits primed with the HXB-2 Env DNA were, for the most part, type-specific for neutralization of IIIb. In one of three assays, sera from rabbits primed with plasmids expressing the JR-FL and BaL had possible low titer neutralizing activity for two NSI, but not two SI, primary isolates. Our results highlight the low immunogenic potential of the HIV-1 Env and demonstrate that different Envs have different potentials to raise low titer neutralizing antibody.

AIDS Vaccines↗
PubMed1997

Eliciting HIV-1 envelope-specific antibodies with mixed vaccinia virus recombinants.

Recombinant vaccinia virus (VV) vectors that express the envelope (Env) protein of the human immunodeficiency virus-type 1 (HIV-1) have been previously shown to elicit HIV-specific cytotoxic T-lymphocyte (CTL) and weak antibody responses in non-human primate studies and clinical trials. In first clinical trials, single Env proteins were presented to the immune system by VV recombinants and other vectors, but individuals were not protected against later exposures to heterologous HIV. It is likely that the generation of protective immune responses against diverse HIV will require that vaccines encompass proteins from not just one, but multiple distinct HIV isolates. Here is described the simple construction of numerous new VV, each expressing a unique, truncated, Env protein (VVenv). Mouse experiments were performed to evaluate the ability of these VVenv to elicit immune responses. HIV-1-specific antibodies appeared within one month following one intraperitoneal inoculation of mice with single or mixed VVenv, reaching plateau levels by 4 months. The magnitude of antibody production was poor at the dose of 10(5) p.f.u. VVenv per animal, but improved with increasing doses of VVenv up to 10(7) p.f.u. per animal. The subcutaneous route of VV immunization, previously proven safe in human trials, was also effective for administering VVenv. These results highlight the strengths of recombinant VV constructs as vehicles for the presentation of multiple HIV-1-Env proteins to the naive immune system.

AIDS Vaccines↗
PubMed1997

Intranasal Sendai virus vaccine protects African green monkeys from infection with human parainfluenza virus-type one.

Human parainfluenza virus-type I (hPIV-1) infections are a common cause of "group" and hospitalizations among young children. Here we address the possibility of using the xenotropic Sendai virus [a mouse parainfluenza virus (PIV)] as a vaccine for hPIV-1. Sendai virus was administered to six African green monkeys (Cercopithecus aethiops) by the intranasal (i.n.) route. A long lasting virus-specific antibody response was elicited, both in the serum and nasal cavity. Sendai virus caused no apparent clinical symptoms in the primates, but live virus was detected in the nasal cavity for several days after inoculation. No virus was detected after a second dose of Sendai virus was administered on day 126 after the initial priming. Animals were challenged with hPIV-1 i.n. on day 154. All six vaccinated animals were fully protected from infection while six of six control animals were infected with hPIV-1. The antibody responses induced by Sendai virus immunizations proved to be greater than those induced by hPIV-1. These results demonstrate that unmanipulated Sendai virus is an effective vaccine against hPIV-1 in a primate model and may constitute a practical vaccine for human use.

Administration, Intranasal↗
PubMed1997

Inter- and intra-patient sequence diversity among parainfluenza virus-type 1 nucleoprotein genes.

Parainfluenza viruses (PIV) have been categorized into four discrete types (types 1-4), based on antigenic similarities. Here is described an evaluation of nucleoprotein (NP) sequence variability among nine patients infected with the type 1 virus. The examination of short segments of the NP sequence was sufficient to define significant variability both within and between patient samples. These data, in conjunction with previous studies of hemagglutinin-neuraminidase and fusion protein sequences from PIV-infected patient populations suggest a lack of absolute stability among isolates within each virus type. Potentially, antigenic variability exists to the extent that an immune response elicited toward one isolate may not be fully protective against another of the same type. Thus, sequence variability could contribute to natural re-infections with PIV, as well as to previous vaccine failures. Results highlight the importance of analyzing viruses that break through vaccine-induced immunity, in order to measure the influence of virus diversity on PIV vaccine outcome.

Amino Acid Sequence↗
PubMed1996

Restricted onset of T cell receptor alpha gene rearrangement in fetal and neonatal thymocytes.

The initial T cell receptor (TCR) alpha gene rearrangements were analyzed in fetal and neonatal thymocyte hybridomas by Southern blotting. Interestingly, in 30% of all thymocyte hybridomas and in all fetal day 16 thymocyte hybridomas the most proximal J alpha 50 (psi J alpha) gene was rearranged. This rearrangement was found on one chromosome only and mostly in association with a delta rearrangement on the homologous chromosome. J alpha 50 was rearranged to multiple target genes based on the variable size of the restriction fragments. In addition, delta rearrangement was found with a concomitant alpha rearrangement in the majority of hybridomas and it was not only associated with J alpha 50 but with several other rearranged J alpha genes as well. Our results clearly demonstrate that T cell precursors are not pre-committed to either delta or alpha rearrangement but a flexible progenitor responds to multiple regulatory signals during T cell differentiation and they do not support the notion that delta rec-psi J alpha rearrangement is required for cell commitment to TCR alpha gene rearrangement.

Animals↗
PubMed1996

Eliminating graft-versus-host potential from T cell immunotherapeutic populations.

It is common knowledge that donor T cells are responsible for graft-versus-host disease (GVHD) following bone marrow transplantation (BMT), yet GVHD remains a grave threat to transplant patients. The donor marrow can be purged of T cells to reduce this danger, but the risks of viral infections, tumor relapse and graft rejection are then increased. Here we describe a method that may be used to provide BMT patients with T cell immunotherapeutic populations responsive to foreign antigens, but unresponsive to host HLA. The method involves the culture of donor T cells with host-derived B lymphoblastoid cell lines (BLCL). During culture, the T cells are activated by the mismatched host HLA. Activated cells are subsequently removed by fluorescence-activated cell sorting. Criteria for removal include cell size and the expression of multiple T cell activation antigens on cell membranes. After the procedure, T cell populations retain helper and cytotoxic T cell responses against foreign antigens, but are specifically devoid of responses to host HLA. This technique offers a promising method for providing BMT patients the benefits of T cell immunity without the consequences of GVHD.

Bone Marrow Transplantation↗
PubMed1995

Human parainfluenza virus type 1 immunization of infant mice protects from subsequent Sendai virus infection.

Human parainfluenza virus type 1 (hPIV-1) infections are a common cause of "croup" and hospitalizations among young children, yet no vaccine is yet available. Sendai virus (mouse PIV-1) is the closest known homologue of hPIV-1. Here we address the possibility of using a xenotropic, nonpathogenic PIV as a vaccine in infants, by assessing the efficacy of hPIV-1 vaccination of infant mice against a subsequent challenge with Sendai virus. hPIV-1 was administered intranasally to mice age 3-6 days and shown by serum antibody ELISA and elispot analysis to elicit virus-specific IgM and isotype-switched antibody-forming cells (AFC). The response was completely cross-reactive between hPIV-1 and Sendai virus. Mice were challenged with Sendai virus 6-8 weeks later and generated AFC and serum antibody responses composed of IgM, as well as IgG and IgA, unlike challenged, age-matched controls. The high IgM response among AFC was not seen in mice primed as adults with hPIV-1 and challenged with Sendai virus. The hPIV-1 priming of infant mice afforded protection, as the majority of these mice survived the lethal Sendai virus challenge, as did all adult primed animals. These data support the notion that the unmodified xenotropic Sendai virus might function effectively in human infants as a vaccine against hPIV-1.

Age Factors↗
PubMed1995

T cell receptor repertoire of CD4+ and CD8+ T cell subsets in the allogeneic bone marrow transplant recipient.

Allogeneic bone marrow transplantation (BMT) has become a therapy of choice for the treatment of certain malignancies and hematopoietic disorders. However, immunodeficiencies following BMT continue to cause significant morbidity and mortality. We have compared the T cell receptor (TCR) repertoire of BMT patients and healthy control individuals by staining peripheral blood mononuclear cells with fluorochrome-labeled TCR-specific antibodies. Several patients exhibited a biased pattern of TCR expression atypical of the healthy controls, yet no particular TCR bias characterized all patients. For example, we found that 2%-8% of T cells from healthy individuals expressed the V beta 19 TCR. One BMT patient exhibited V beta 19 expression on more than 60% of peripheral T cells, while additional patients expressed V beta 19 on less than 1% of T cells. The patients with the most extreme skewing of TCR types suffered from graft-versus-host disease. The causes of skewed TCR V beta expression patterns in BMT patients are not fully understood, yet some researchers have suggested that an oligoclonal expansion of CD8+ T cell populations may be largely responsible. To test this hypothesis, we examined the TCR V beta repertoire of CD4+ and CD8+ T cell populations. We found that biased V beta expression characterized both CD4+ and CD8+ T cell populations, sometimes within a single individual. Thus, therapies directed toward CD8+ T cells alone may not fully correct repertoire abnormalities following BMT.

Adolescent↗
PubMed1995

Gene rearrangement patterning and DNase-I hypersensitive sites within the T-cell receptor J alpha locus.

For several years, the relationship between alpha beta and gamma delta T-cell progenitors has been a topic of debate. Some argue that a subset of T-cell progenitors is "pre-committed" to the alpha beta lineage and is thus programmed to rearrange alpha, but not delta genes. It is further argued that the deletion of the delta locus by a unique rearrangement, delta rec-psi J alpha, may be the critical forerunner to V-J alpha joins in alpha beta committed cells and that a hypersensitive site (HS) termed 5'TEA might regulate such rearrangement. Here we present an alternative hypothesis. We first emphasize that directed J alpha gene rearrangements do not exclusively target the psi J alpha gene, but that clustered gene rearrangements occur throughout the J alpha locus during T-cell development. We describe the existence of not one, but at least two HS sites distributed along the J alpha locus which might serve as regulators for the gene rearrangement event. Finally, we suggest that progenitor T-cells are not committed to a particular delta or alpha gene rearrangement, but that a flexible progenitor responds to complex interactions between environmental signals and multiple regulatory elements interspersed among delta/alpha genes.

Animals↗
PubMed1994

Evidence for recombinatorial hot spots at the T cell receptor J alpha locus.

The complex genomic organization of the murine T cell receptor (TcR) delta-alpha region has hindered detailed studies of alpha gene rearrangement and J alpha gene usage in individual differentiating T cell precursors. We have isolated a novel set of J alpha probes which, in combination with a few restriction enzyme digests, enable a reliable, simple and nearly complete analysis and location of any rearrangement at the J alpha locus by conventional Southern blotting. The probes were used to analyze TcR alpha gene rearrangements in T cell hybridomas derived from an in vitro culture system that supports T cell differentiation of bone marrow cells. Our results indicate that J alpha genes are unequally accessible for rearrangement and two hot spots for rearrangement could be demonstrated. In addition, only a restricted set of J alpha genes was rearranged in each culture indicating that the slightly variable composition of factors can influence the recombinatorial accessibility of J alpha genes. The hot spots for rearrangement were not only limited to T cells differentiating in vitro but could also be demonstrated among functional T cell clones based on the published sequence information from isolated TcR alpha gene rearrangements. The demonstration and the location of the hot spots for rearrangement in the T cell differentiation culture system opens up the possibility to study factors and mechanisms that regulate recombinatorial accessibility of TcR alpha genes.

Animals↗
PubMed1994

Viral cross-reactivity and antigenic determinants recognized by human parainfluenza virus type 1-specific cytotoxic T-cells.

To obtain information relevant to vaccination against human parainfluenza virus type 1 (hPIV-1), cytotoxic T-lymphocyte (CTL) responses to individual viral components were tested. The CD8-positive T-cell fraction was first enriched from human, adult PBL and grown for several passages in the presence of hPIV-1-infected stimulator cells. T-cell lines were then tested for CTL activity toward hPIV-1 and toward the related viruses hPIV-3 and Sendai virus (the murine parainfluenza type 1 virus). All tested cultures which responded to hPIV-1 also responded to hPIV-3 and Sendai virus, demonstrating sequence conservation between all three viruses among major antigenic determinants for CTL. Specificity for particular viral components was defined using recombinant vaccinia viruses expressing individual proteins from either mouse or human parainfluenza type 1 viruses. Strong CTL responses toward hemagglutinin-neuraminidase, phosphoprotein, and nucleoprotein (NP) were demonstrated. The testing of vaccinia constructs expressing truncated proteins then showed that there were multiple CTL determinants within NP. Several T-cell lines from one donor recognized an NP peptide (amino acids 321-336) conserved between the hPIV-1 and Sendai virus. In total, the results demonstrated that the human CTL response is directed to multiple determinants within several distinct hPIV-1 proteins.

Adult↗
PubMed1994

Age-related development of human memory T-helper and B-cell responses toward parainfluenza virus type-1.

Human parainfluenza-1 virus (hPIV-1) infections are a major cause of respiratory illness in young children. While children and adults are each susceptible to hPIV-1 infection, the clinical symptoms in adults are mild and hospitalizations are rare. One explanation for the differences in disease severity is that immune memory responses are simply inferior in children as compared to adults and cannot counter virus growth. Alternatively, it has been suggested that immune (particularly T-helper (TH) cell) responses toward respiratory viruses are superior in children versus older individuals, and that these responses contribute to, rather than protect from, disease symptoms. As a test of these possibilities, we analyzed hPIV-1-specific T-helper (TH) and B-cell memory responses among individuals of various ages, including children hospitalized with hPIV-1-induced croup. Experiments revealed: (1) hPIV-1-specific B-cell and class-II restricted TH-cell proliferative responses were present in all tested adults. (2) TH-cells responded to internal viral proteins as well as to the external glycoprotein, hemagglutinin-neuraminidase. (3) Immune responses were highly cross-reactive with Sendai virus. (4) Memory B-cell and TH-cell responses were extremely poor in young children, inclusive of children tested upon hospital entry for hPIV-1-induced croup. In total, results did not support the theory that naturally induced hPIV-specific memory responses cause respiratory illness. Rather, results showed a correlation between memory and a good clinical outcome and highlighted Sendai virus as a strong candidate for an hPIV-1 vaccine.

Adult↗