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Biomedical subjects

S L Hoffman

Publications and source records attributed to S L Hoffman.

At least 127 records · Page 7Linked to original sources
PubMed1996

DNA vaccines against malaria: immunogenicity and protection in a rodent model.

Since the first demonstration of the technology a few years ago, DNA vaccines have emerged as a promising method of vaccination. In a variety of experimental systems, DNA vaccines have been shown not only to induce potent immune responses, but also to offer many advantages in terms of ease of construction, testing, and production. In this article we summarize the progress achieved in development of DNA vaccines that can protect mice from infection by the rodent malaria parasite Plasmodium yoelii, describe initial studies of immunogenicity of a malaria DNA vaccine in a primate model, and outline the strategies being employed to design the next generation of malaria DNA vaccines.

Animals↗
PubMed1996

Lymphocyte proliferative response and subset profiles during extended periods of chloroquine or primaquine prophylaxis.

Immune suppression and disturbances of normal leukocyte populations are side effects attributed to many antimalarial drugs and were concerns during a recent year-long placebo-controlled trial that compared daily primaquine (0.5 mg of base per kg of body weight per day) with weekly chloroquine (300 mg of base one time per week) for malaria prophylaxis. The study took place in Irian Jaya, Indonesia, from July 1994 to August 1995 and enrolled 129 Javanese men with normal glucose-6-phosphate dehydrogenase function. Tests for lymphocyte function and subset composition were conducted blindly on a cross-section of subjects during weeks 10 (n = 42) and 48 (n = 72) of supervised prophylaxis. Lymphocyte function, measured as the proliferative response of peripheral blood mononuclear cells to a panel of mitogens (pokeweed mitogen, phytohemagglutinin, and concanavalin A) and antigens (purified protein derivative of Mycobacterium tuberculosis and Clostridium tetani toxoid) and expressed as a stimulation index, allowed for statistical comparison between groups and sampling times. The lymphocyte subset composition for each group and time point was based on flow cytometry profiling, and the results were expressed as the mean percentages of CD3 (total T cells), CD19 (total B cells), CD4+ (T-helper and inducer cells), and CD8+ (T suppressor and cytotoxic cells), CD3/CD16+ CD56 (natural killer cells), CD3/anti-HLA-DR (activated T cells) cells and the CD4+/CD8+ ratios. Lymphocyte stimulation indices were statistically comparable among the placebo, primaquine, and chloroquine groups at both time points, although the primaquine group was distinguished by having repeatedly greater proportions of subjects with high ( > 3.0) stimulation indices. The lymphocyte subset profiles of these groups at both time points were also similar and undistorted relative to those of healthy reference populations matched for age, sex, and ethnicity. The results provide quantitative support for the safety of daily primaquine prophylaxis.

Adult↗
PubMed1995

Irradiated sporozoite vaccine induces HLA-B8-restricted cytotoxic T lymphocyte responses against two overlapping epitopes of the Plasmodium falciparum sporozoite surface protein 2.

Vaccines designed to protect against malaria by inducing CD8+ cytotoxic T lymphocytes (CTL) in individuals of diverse HLA backgrounds must contain multiple conserved epitopes from various preerythrocytic-stage antigens. Plasmodium falciparum sporozoite surface protein 2 (PfSSP2) is considered an important antigen for inclusion in such vaccines, because CD8+ CTL against the P. yoelii SSP2 protect mice against malaria by eliminating infected hepatocytes. To develop PfSSP2 as a component of malaria vaccines, we investigated the presence of anti-PfSSP2 CTL in two HLA-B8+ volunteers immunized with irradiated P. falciparum sporozoites and characterized their CTL responses using PfSSP2-derived 15-amino acid peptides bearing the HLA-B8-binding motif. Peripheral blood mononuclear cells from both volunteers stimulated with recombinant vaccinia expressing PfSSP2 displayed antigen-specific, genetically restricted, CD8+ T cell-dependent CTL activity against autologous target cells expressing PfSSP2. Of the five HLA-B8 motif-bearing 15-mers identified in the PfSSP2 sequence, two peptides sharing a 10-amino acid overlap sensitized HLA-B8-matched target cells from both volunteers for lysis by peptide-stimulated effectors. The CTL activity was HLA-B8 restricted and dependent on CD8+ T cells. Analysis of the three shorter peptides representing HLA-B8 motif-bearing sequences within the two positive peptides for their ability to bind to HLA-B8 in vitro, and to sensitize target cells for lysis by effectors stimulated with the 15-mers, identified two overlapping HLA-B8-restricted CTL epitopes. Available data indicate that the sequence of one CTL epitope is conserved and the other is variant among P. falciparum isolates. Circulating activated CTL against the conserved epitope could be directly identified in one of the two volunteers. The identification of two HLA-B8-restricted CTL epitopes on PfSSP2 provides data critical to developing an epitope-based anti-liver stage malaria vaccine.

Amino Acid Sequence↗
PubMed1995

Complexity of the cytokine and antibody response elicited by immunizing mice with Plasmodium yoelii circumsporozoite protein plasmid DNA.

The number, type, and location of cytokine- and Ab-secreting cells activated in mice immunized and boosted with plasmid DNA encoding the circumsporozoite protein of the malarial parasite Plasmodium yoelii (PyCSP) were monitored. The initial humoral response was localized to the draining lymph nodes and was characterized by production of IgG1 anti-PyCSP Abs and the Th2 cytokine IL-4. In contrast, the secondary response was dominated by IFN-gamma production (a Th1 cytokine) and the secretion of IgG2a anti-PyCSP Abs in the spleen. PyCSP DNA and mRNA were detected only in the quadriceps muscles (sites of plasmid injection), yet these sites lacked either cytokine- or Ab-secreting cells. These findings indicate that circulating lymphocytes encounter plasmid-encoded Ag in the muscle bed, initiate a humoral response in the draining lymph nodes, and then seed distal lymphoid organs. Profound differences were observed between the primary and secondary immune responses induced by plasmid immunization, which may influence vaccine efficacy.

Animals↗
PubMed1995

HLA-A2-restricted cytotoxic T lymphocyte responses to multiple Plasmodium falciparum sporozoite surface protein 2 epitopes in sporozoite-immunized volunteers.

CD8+ CTL specific for the Plasmodium yoelii sporozoite surface protein 2 (PySSP2) protect mice against malaria. For this reason, vaccines designed to induce CTL against P. falciparum SSP2 (PfSSP2) are under development. Optimal development of PfSSP2 as a component of human malaria vaccines requires characterization of HLA class I-restricted CTL against this Ag. For this purpose, PBMC from four HLA-A2+ human volunteers immunized with P. falciparum irradiated sporozoites were stimulated with a recombinant vaccinia virus expressing PfSSP2 and with 35 PfSSP2-derived 15-amino acid peptides containing sequences conforming to HLA-A2 binding motifs. Ag-specific, genetically restricted, CD8+ T cell-dependent cytotoxic activity against autologous target cells transfected with the PfSSP2 gene was demonstrated in the four volunteers. Twelve of the 35 peptides sensitized HLA-A2-matched target cells for lysis by peptide-stimulated effectors. Three volunteers had CTL against 9 of the 12 peptides, and one had no peptide-specific CTL. HLA-A*0201 restriction was confirmed by demonstrating that effectors from the three responders could be stimulated with six different peptides to lyse HLA-A*0201+ T2 cells incubated with the homologous peptides. Peptide-specific, genetically restricted, CD8+ T cell-dependent cytotoxic activity was also demonstrated against two peptides using unstimulated PBMC as effectors. Available data indicate that the motif-bearing sequences in 6 of the 12 positive peptides are conserved among P. falciparum isolates and clones. Demonstration of HLA-A2-restricted CTL responses to multiple PfSSP2-derived peptides, and of circulating activated CTL against PfSSP2 in immune volunteers provide important information for optimal design and evaluation of vaccines containing this pre-erythrocytic stage Ag.

Amino Acid Sequence↗
PubMed1995

Induction of protective polyclonal antibodies by immunization with a Plasmodium yoelii circumsporozoite protein multiple antigen peptide vaccine.

Monoclonal Abs against the repeat region of the circumsporozoite protein (CSP) completely protect mice against Plasmodium yoelii (Py), but synthetic peptide and recombinant protein vaccines designed to produce only Abs to the PyCSP repeat region have never been reported to consistently provide protection. This lack of protection in the rodent model system has predicted the poor protection achieved in humans after immunization with synthetic peptide and recombinant protein P. falciparum CSP vaccines and has raised serious questions regarding the capacity for vaccine-induced polyclonal Abs against the CSP to consistently protect humans. We now report immunization studies with a multiple Ag peptide vaccine designed to rely on "universal" T epitopes from tetanus toxin to produce T cell help for induction of protective Abs against the repeat region of the PyCSP. When delivered with a nonionic block co-polymer adjuvant, the vaccine protected 78 to 100% of three inbred strains of mice, and 100% of outbred mice against P. yoelii sporozoite challenge. Protection was associated with Ab titer, and passive transfer of purified IgG from immune mice protected naive recipients. Similar protection was achieved when the peptide was encapsulated in liposomes with lipid A and mixed with aluminum hydroxide. By demonstrating for the first time solid protection against P. yoelii by polyclonal Abs against the CSP, these data provide the rationale for assessment of a similarly constructed and formulated P. falciparum CSP multiple Ag peptide vaccine in humans.

Amino Acid Sequence↗
PubMed1995

Inhibition of extracellular ATP-mediated lysis of human macrophages by calmodulin antagonists.

Lysis of human culture-derived macrophages by extracellular ATP has recently been described, and treatment of macrophages with interferon-gamma rendered those cells significantly more sensitive to lysis. In addition, cell death occurred more rapidly in interferon (IFN)-treated cells than in untreated macrophages. In an attempt to identify the mechanism by which extracellular ATP affects macrophages, as well as to explore the differences between interferon-gamma-treated and untreated macrophages, selected metabolic inhibitors were included in the lytic assays. Of the compounds tested, three antagonists of calmodulin-linked pathways (trifluoperazine, KN-62, and calmidazolium) blocked the ATP-mediated lysis of both interferon-gamma-treated and colony-stimulating factor-treated macrophages in a dose-dependent manner. Early signals of the ATP ligation of the P2Z purinoceptors of human macrophages included increases in cytosolic [Ca2+] and depolarization of the plasma membrane. However, the inclusion of calmodulin antagonists in these assays did not abrogate either effect. These results suggest that the mechanism which mediates the efflux of 51Cr-labeled proteins from ATP-lysed macrophages is distinct from calcium mobilization and membrane depolarization, and may involve the generation of secondary pores/channels in the plasma membrane via a calmodulin-linked pathway.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1995

Plasmodium yoelii: 17-kDa hepatic and erythrocytic stage protein is the target of an inhibitory monoclonal antibody.

Infected hepatocytes are important targets for malaria vaccines. To identify Plasmodium yoelii proteins expressed in infected hepatocytes, we immunized BALB/c ByJ mice with P. yoelii liver stage schizonts and produced a panel of monoclonal antibodies (Mabs). An IgG1 Mab, navy yoelii liver stage 3 (NYLS3), had the strongest reactivity against liver stage parasites and was selected for further characterization. The Mab does not recognize P. yoelii sporozoites, but recognizes liver stage parasites within 6 hr of invasion of mouse hepatocytes and throughout the hepatic and asexual erythrocytic stages of the parasite life cycle as determined by the immunofluorescent antibody test. This Mab is species-specific, and it reacts with liver stages of P. yoelii but does not react with liver stages of other Plasmodium species. The protein recognized by this Mab is present on the parasitophorous vacuole membrane of infected hepatocytes and erythrocytes as demonstrated by immunoelectron microscopy and has a relative molecular weight of 17 kDa as demonstrated by immunoblot of an extract of infected erythrocytes. It is therefore designated P. yoelii hepatic and erythrocytic stage protein, 17 kDa or PyHEP17. When added to primary cultures of mouse hepatocytes 24 hr after inoculation with P. yoelii sporozoites, when all sporozoites have invaded hepatocytes, NYLS3 eliminates up to 98% of liver-stage parasites. Intravenous injection of NYLS3 into mice delays the onset and reduces the density of blood-stage parasitemia after sporozoite or blood-stage challenge. The P. falciparum and P. vivax homologs of PyHEP17 may therefore be important targets for vaccines designed to attack the hepatic and erythrocytic stages of the parasite life cycle.

Animals↗
PubMed1995

Synthesis and immunological characterization of 104-mer and 102-mer peptides corresponding to the N- and C-terminal regions of the Plasmodium falciparum CS protein.

We investigated the immunogenicity and the conformational properties of the non-repetitive sequences of the Plasmodium falciparum circumsporozoite (CS) protein. Two polypeptides of 104 and 102 amino acids long, covering, respectively, the N- and C-terminal regions of the CS protein, were synthesized using solid phase Fmoc chemistry. The crude polypeptides were purified by a combination of size exclusion chromatography and RP-HPLC. Sera of mice immunized with the free polypeptides emulsified in incomplete Freund's adjuvant strongly reacted with the synthetic polypeptides as well as with native CS protein as judged by ELISA and IFAT assays. Most importantly, these antisera inhibited the sporozoite invasion of hepatoma cells. In addition, sera derived from donors living in a malaria endemic area recognized the CS 104- and 102-mers. Conformational studies of the CS polypeptides were also performed by circular dichroism spectroscopy showing the presence of a weakly ordered structure that can be increased by addition of trifluoroethanol. The obtained results indicate that the synthetic CS polypeptides and the natural CS protein share some common antigenic determinants and probably have similar conformation. The approach used in this study might be useful for the development of a synthetic malaria vaccine.

Amino Acid Sequence↗
PubMed1995

Partial protection against malaria by immunization with Leishmania enriettii expressing the Plasmodium yoelii circumsporozoite protein.

Since infection with Leishmania species induces CD4+ and CD8+ anti-leishmania T cells, we assessed protection against malaria by immunization with Leishmania enriettii transfected with the gene encoding the Plasmodium yoelii circumsporozoite protein (PyCSP). The recombinant plasmid appeared to be a circular episome in the host cells. Reverse transcription PCR showed that the PyCSP was trans-spliced by the addition of the 39-bp spliced leader of L. enriettii at its 5' end. The transfectant expressed a protein in a pattern similar to that found in the sporozoite itself. Immunofluorescence and immunoelectron microscopy indicated that PyCSP was abundantly expressed on the surface of the parasite. Mice immunized with the transfectant produced antibodies to sporozoites, had a delay in onset of parasitemia after challenge, and 4 of 22 (18%) were completely protected. The protected mice had cytotoxic T lymphocytes against the PyCSP. Immunization with recombinant vaccinia, Salmonella typhimurium, and pseudorabies virus expressing the PyCSP induces excellent immune responses, but has not been shown to protect against challenge. Thus, the modest protection found in these initial studies represents a step forward. After further work Leishmania may prove to be an important live vector vaccine system for induction of protective immune responses.

Amino Acid Sequence↗
PubMed1995

Malarial epitopes expressed on the surface of recombinant tobacco mosaic virus.

Using malaria as a model disease, we engineered the surface of tobacco mosaic tobamovirus (TMV) for presentation of selected epitopes to the mammalian immune system. The TMV coat protein is a well-characterized and abundant self-assembling polymer previously shown to be a highly immunogenic carrier. Selected B-cell epitopes were either inserted into the surface loop region of the TMV coat protein or fused to the C terminus using the leaky stop signal derived from the replicase protein reading frame. Tobacco plants systemically infected with each of these constructs contained high titers of genetically stable recombinant virus, enabling purification of the chimeric particles in high yield. Symptoms induced in tobacco ranged from a normal mosaic pattern similar to that induced by the parental U1 strain to a unique bright yellow mosaic. As measured by quantitative ELISA against synthetic peptide standards, wild type TMV coat protein and fusion protein synthesized by the leaky stop mechanism coassembled into virus particles at the predicted ratio of approximately 20:1. Recombinant plant viruses have the potential to meet the need for scalable and cost effective production of subunit vaccines that can be easily stored and administered.

Amino Acid Sequence↗
PubMed1995

Daily primaquine is effective for prophylaxis against falciparum malaria in Kenya: comparison with mefloquine, doxycycline, and chloroquine plus proguanil.

Primaquine was tested as a prophylactic drug against Plasmodium falciparum in a region in western Kenya in which malaria is holoendemic. Children 9-14 years old were randomized to receive regimens of daily primaquine, daily doxycycline, daily proguanil plus weekly chloroquine, daily vitamin plus weekly mefloquine, or daily vitamin alone. Primaquine, doxycycline, and mefloquine were equally effective in preventing both symptomatic and asymptomatic malarial infections. Chloroquine plus proguanil was the least effective regimen. There was no toxicity from daily primaquine during the 11 weeks of the study. Findings show that primaquine can be successfully used as a causal prophylactic regimen against falciparum malaria in western Kenya; chloroquine plus proguanil was not as efficacious as the three other preventive regimens; most Kenyan children receiving standard doses of mefloquine and doxycycline had lower than expected serum trough drug levels; and some volunteers with adequate mefloquine or doxycycline levels at trough developed asymptomatic parasitemias and clinical malaria.

Adolescent↗
PubMed1995

Treatment of chloroquine-resistant Plasmodium vivax with chloroquine and primaquine or halofantrine.

Optimal therapy for infection by chloroquine-resistant Plasmodium vivax has not been established. From 1992 to 1994 during three separate studies, 147 Javanese residents of Irian Jaya infected by P. vivax were treated with either chloroquine (25 mg of base/kg during 3 days or 10 mg of base/kg in one dose) plus primaquine (10 mg/kg during 28 days or 2.5 mg/kg during 3 days) (n = 78), chloroquine plus placebo (n = 50), or halofantrine (24 mg base/kg in 12 h; n = 19). There was no difference in tolerance to or side effects of any of the regimens. Within 14 days of starting therapy, therapeutic failure among these patients was 44% for chloroquine, 5% for chloroquine plus primaquine (P < .001), and 0 for halofantrine (P < .001). After 28 days, therapeutic failure was 78%, 15%, and 6%, respectively. Thus, chloroquine plus primaquine in combination and halofantrine alone are effective therapies for chloroquine-resistant P. vivax.

Adolescent↗
PubMed1995

Impact of transmission intensity and age on Plasmodium falciparum density and associated fever: implications for malaria vaccine trial design.

To facilitate design of vaccine trials, malaria was studied in 6-month- to 6-year-old Kenyans during high (HI) and low intensity transmission seasons. During 84 days after cure, exposure to infected mosquitoes was 9-fold greater in the HI group, yet incidence of P. falciparum infection was increased only 2-fold, with no age effect. The density of recurrent P. falciparum was 14-fold greater in the HI group, and there was a striking association between age and parasitemia > or = 5000/microL. Fever was the only clinical manifestation attributable to parasitemia and only when the parasite density was > or = 5000/microL. Sixty-four percent of children with > or = 20,000 parasites/microL versus 10% with 1-4999/microL were febrile when parasitemic. Recurrent P. falciparum infection as a vaccine trial end point can be studied year-round among children < or = 6 years [corrected] in western Kenya. However, high-grade parasitemia (> or = 5000 or 20,000/microL) with or without elevated temperature will be optimally studied in the high transmission season among children < 2 years.

Age Factors↗