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S L Hoffman

Publications and source records attributed to S L Hoffman.

At least 91 records · Page 5Linked to original sources
PubMed1998

Simultaneous induction of multiple antigen-specific cytotoxic T lymphocytes in nonhuman primates by immunization with a mixture of four Plasmodium falciparum DNA plasmids.

CD8(+) T cells have been implicated as critical effector cells in protective immunity against malaria parasites developing within hepatocytes. A vaccine that protects against malaria by inducing CD8(+) T cells will probably have to include multiple epitopes on the same protein or different proteins, because of parasite polymorphism and genetic restriction of T-cell responses. To determine if CD8(+) T-cell responses against multiple P. falciparum proteins can be induced in primates by immunization with plasmid DNA, rhesus monkeys were immunized intramuscularly with a mixture of DNA plasmids encoding four P. falciparum proteins or with individual plasmids. All six monkeys immunized with PfCSP DNA, seven of nine immunized with PfSSP2 DNA, and five of six immunized with PfExp-1 or PfLSA-1 DNA had detectable antigen-specific cytotoxic T lymphocytes (CTL) after in vitro restimulation of peripheral blood mononuclear cells. CTL activity was genetically restricted and dependent on CD8(+) T cells. By providing the first evidence for primates that immunization with a mixture of DNA plasmids induces CD8(+) T-cell responses against all the components of the mixture, these studies provide the foundation for multigene immunization of humans.

Amino Acid Sequence↗
PubMed1998

In vitro expression and in vivo immunogenicity of Plasmodium falciparum pre-erythrocytic stage DNA vaccines.

DNA vaccine plasmids were constructed that encoded four pre-erythrocytic antigens from the human malaria parasite Plasmodium falciparum: circumsporozoite protein (PfCSP); sporozoite surface protein 2 (PfSSP2); carboxyl terminus of liver stage antigen 1 (PfLSA-1 c-term); and, exported protein 1 (PfExp-1). Antigen expression was evaluated in vitro by immunoblot analysis of tissue culture cells following transient transfection with each plasmid. Clearly detectable levels of expression depended upon, or were markedly enhanced by, fusion of the antigen encoding sequences in-frame with the initiation complex and peptide leader sequence of human tissue plasminogen activator protein. Mice injected with these plasmids produced antigen specific antibody and cytotoxic T lymphocyte responses. However, the magnitudes of the responses were not always predicted by the in vitro expression assay. The results of this study provided the basis for further testing of these plasmids in primates and the formulation of multi-component pre-erythrocytic DNA vaccines for efficacy testing in human volunteers.

Amino Acid Sequence↗
PubMed1998

Humoral immune responses against Plasmodium vivax MSP1 in humans living in a malaria endemic area in Flores, Indonesia.

The aim of this study was to evaluate the relationship among age, parasitemia status, spleen size, hematocrit, and antibody levels to Plasmodium vivax merozoite surface protein 1 (MSP1) in individuals chronically exposed to P. vivax. Subjects were recruited from the population of three adjacent villages on the Island of Flores in Indonesia where malaria transmission is hyperendemic and tropical splenomegaly syndrome is highly prevalent. Subjects were evaluated for spleen size, hematocrit, presence of parasitemia, and presence of antibodies to a recombinant peptide consisting of 90 amino acids from the carboxy terminus of MSP1. Fifty-seven percent of 2-4 year olds, 45% of 5-9 years old, and 7% of > or = 15 years old were parasitemic; 99% of the > or = 15 years old had splenomegaly, and 31% of them had Hackett 4 or 5 spleens. The frequency of antibody positivity to MSP1 antigen in ELISA increased with age reaching a maximum of 89% in > or = 20 years old. The frequency of antibody positivity to MSPI also increased with spleen size, and with a decline in the prevalence of parasitemia.

Adolescent↗
PubMed1998

Immune response to a hepatitis B DNA vaccine in Aotus monkeys: a comparison of vaccine formulation, route, and method of administration.

BACKGROUND: Attempts to optimize DNA vaccines in mice include using different routes of administration and different formulations. It may be more relevant to human use to carry such studies out in nonhuman primates. Here we compare different approaches to delivery of a DNA vaccine against the hepatitis B virus (HBV) in Aotus monkeys. MATERIALS AND METHODS: Thirty-two adult Aotus l. lemurinus monkeys divided into 8 groups of four were immunized with 400 microg of a DNA vaccine which encoded hepatitis B surface antigen (HBsAg). DNA in saline was administered by intradermal (ID) or intramuscular (IM) injection with needle and syringe, IM injection with the Biojector needleless injection system or combined ID (needle) and IM (Biojector). DNA formulated with cationic liposomes (CellFECTIN) was injected IM with needle or Biojector. DNA with added E. coli DNA (100 microg) was injected IM with the Biojector or ID. A ninth group of 4 monkeys was injected IM (needle) with Engerix-B, a commercial vaccine containing recombinant HBsAg (10 microg) adsorbed onto alum. Monkeys were boosted in an identical fashion to their prime at 8 weeks, but all received the protein vaccine (Engerix-B) at 16 weeks. Sera was assessed for antibodies against HBsAg (anti-HBs) by enzyme-linked imunosorbent assay (ELISA). RESULTS: The primary humoral response induced by IM delivery of the DNA vaccine was very poor. In most cases there was no detectable anti-HBs even after 2 DNA doses but the kinetics of the response to subsequent protein indicated that a memory B cell response had been induced. In contrast, following IM-administration of DNA using the Biojector, detectable anti-HBs were observed in 3 of 8 animals and evidence for immunological priming was apparent in an additional 4 of the 8 monkeys. ID injection of DNA vaccine in saline induced a potent antibody response which was augmented 6-fold by the addition of E. coli DNA. Combining ID and IM administration did not improve humoral immunity over ID injection alone. CONCLUSIONS: For immunization of primates with DNA vaccines, ID may be a preferable route to IM, although it is not clear whether the Aotus monkey is a relevant model for humans in this respect. Nevertheless, the use of the Biojector needleless injection system may improve responses with IM delivery of DNA vaccines. As well, the immunostimulatory action of E. coli DNA may be used to augment the humoral response induced by a DNA vaccine.

Adjuvants, Immunologic↗
PubMed1997

Induction of protective CTL responses against the Plasmodium yoelii circumsporozoite protein by immunization with peptides.

To determine the optimum combination, concentration, and formulation of synthetic peptides and adjuvants to induce protective CTL responses against the Plasmodium yoelii circumsporozoite protein (PyCSP), BALB/c mice were immunized with linear and multiple antigen peptides (MAP) including PyCSP CTL and Th epitopes in Montanide's ISA51, Lipofectin, and Lipofectamine. An H-2K(d)-restricted PyCSP CTL epitope, SYVPSAEQI (amino acids (aa) 280-288), recognized by protective CTL clones, was included in the following peptides: a 9-aa linear peptide (SYVPSAEQI; PyCSP9), a 20-aa linear peptide (aa 280-299; SYVPSAEQILEFVKQISSL; PyCSP20), a MAP containing four branches of PyCSP20 (MAP(280-299)), and a linear peptide and a MAP(MAP(280-299)p2p30) in which PyCSP20 was colinearly synthesized with two universal Th epitopes from tetanus toxin (p2p30). A MAP containing the PyCSP Th epitope (aa 57-70; KIYNRNIVNRLLGD) was included in some experiments. The highest specific lytic activity against peptide-pulsed target cells was obtained with splenocytes from mice immunized with three doses at 3-wk intervals of MAP(280-299)p2p30 in Lipofectin or Lipofectamine. Forty percent of the mice immunized with MAP(280-299)p2p30 and Lipofectin were protected against sporozoite challenge. Immunization with CTL and Th epitopes co-linearly synthesized in a MAP induced significantly better CTL than did immunization with the same sequence as a linear peptide, or immunization with a mixture of two individual MAPs, one with the CTL epitope and the second with the Th epitope.

Adjuvants, Immunologic↗
PubMed1997

Pre-erythrocytic-stage immune effector mechanisms in Plasmodium spp. infections.

The potent protective immunity against malaria induced by immunization of mice and humans with radiation-attenuated Plasmodium spp. sporozoites is thought to be mediated primarily by T-cell responses directed against infected hepatocytes. This has led to considerable efforts to develop subunit vaccines that duplicate this protective immunity, but a universally effective vaccine is still not available and in vitro correlates of protective immunity have not been established. Contributing to this delay has been a lack of understanding of the mechanisms responsible for the protection. There are now data indicating that CD8+ T cells, CD4+ T cells, cytokines, and nitric oxide can all mediate the elimination of infected hepatocytes in vitro and in vivo. By dissecting the protection induced by immunization with irradiated sporozoite, DNA and synthetic peptide-adjuvant vaccines, we have demonstrated that different T-cell-dependent immune responses mediate protective immunity in the same inbred strain of mouse, depending on the method of immunization. Furthermore, the mechanism of protection induced by a single method of immunization may vary among different strains of mice. These data have important implications for the development of pre-erythrocytic-stage vaccines designed to protect a heterogeneous human population, and of assays that predict protective immunity.

Animals↗
PubMed1997

Dose- and time-dependent relations between infective Anopheles inoculation and outcomes of Plasmodium falciparum parasitemia among children in western Kenya.

Blood-stage level Plasmodium falciparum infection (parasitemia density) is generally elevated prior to, or at the time of, clinical presentation of severe pediatric malaria episodes. Intensity of exposure to infective Anopheles mosquito bites is a suspected determinant of higher density parasitemia. Analyses of entomologic and parasitologic data collected in 1986-1987 were conducted to investigate whether the dose of infective bites predicted the incidence or degree of P. falciparum parasitemia in Kenyan children < 6 years old. At 21 consecutive 30-day intervals, a new cohort (n approximately 50 each) was enrolled, cured of malaria parasites, and monitored over 84 days for recurrent parasitemia. Outcomes included time to parasitemia, time to parasitemia > or = 5,000/microliter, and parasitemia density. Ecologic and individual-level analyses were conducted. The mean infective bite exposure experienced by each cohort was significantly associated with the incidence of parasitemia (age-adjusted r2 = 0.38, p = 0.022) and more strongly associated with the incidence of parasitemia > or = 5,000/microliter (age-adjusted r2 = 0.72, p < 0.001). The infective bite dose, analyzed as a time-dependent covariate, was associated with a 2.8 times higher rate of parasitemia > or = 5,000/microliter among children exposed to > or = 1 infective bite per day as compared with the referent (rate ratio (RR) = 2.82, 95% confidence interval (CI) 2.24-3.56). Cumulative infective bite exposure, exposure duration, and age were significant predictors of recurrent parasitemia density in multiple linear regression analyses. The results support the contention that reductions in P. falciparum transmission intensity, in the absence of complete elimination, will reduce higher level parasitemia among African children.

Animals↗
PubMed1997

Cytotoxic T lymphocyte (CTL) adherence assay (CAA): a non-radioactive assay for murine CTL recognition of peptide-MHC class I complexes.

Cytotoxic T lymphocytes (CTL) form an important immune surveillance system against intracellular pathogens. Here we describe a simple, visual assay for identifying peptides specifically recognized by CTL, based on the discovery that CTL develop increased adhesive properties upon TCR triggering. Several CTL lines were shown to pellet to the bottom of a round bottom 96-well plate in the absence of peptide. In contrast, these same CTL lines incubated with their cognate peptide, allowing them to present peptide to each other, adhered to the sides of the well and were readily distinguished by macroscopic visual examination of the plate after 4-5 h or overnight incubation. This CTL adherence assay (CAA) demonstrated peptide specificity and MHC restriction, and was titratable with peptide concentration. With this technique, a minimal-sized, malaria CTL epitope was correctly identified from a panel of overlapping nonamers, although the adherence pattern of two mono-substituted, variant peptides was less predictive of lytic activity. Also, substitutions in an HIV-1 envelope CTL epitope that reduced lytic activity were correctly predicted. Inhibitors of RNA and protein synthesis, upon preincubation, abrogated the adherence, indicating, at minimum, a need for live cells. Wortmannin, a PI-3 kinase inhibitor, inhibited the peptide specific adherence, consistent with a role for TCR or integrin signal transduction in CAA. Other cytoskeletal and metabolic inhibitors had no effect. Adherence of the T cells may involve low affinity, nonspecific interactions since wells coated with FCS, BSA or milk powder all produced an effective CAA in the presence of peptide under serum free conditions. Consequently, CAA may represent a rapid, simple method for screening large numbers of peptides to find cytolytic epitopes for a given CTL line and may identify additional epitopes causing T cell activation and adherence but not cytolytic activity.

Animals↗
PubMed1997

Do DNA vaccines induce autoimmune disease?

This report examines whether plasmid DNA vaccines induce the production of anti-DNA or anti-muscle cell autoantibodies. A three-fold increase in the number of B cells secreting immunoglobulin G (IgG) anti-DNA autoantibodies was detected in BALB/c mice immunized and boosted with any of three DNA plasmids (p < 0.004). This correlated with a transient increase in serum anti-DNA autoantibody titers but was not associated with the development of glomerulonephritis or autoimmune disease. None of the DNA vaccines examined stimulated the production of anti-muscle cell autoantibodies or the development of myositis. The effect of DNA vaccines on the development of nascent autoimmunity in lupus-prone (NZB x NZW)F1 mice was also examined. Repeated vaccination did not alter the onset or course of disease in these animals. These findings suggest that DNA vaccines neither initiate nor accelerate the development of systemic autoimmunity.

Animals↗
PubMed1997

Cytotoxic T cell reactivity and HLA-B35 binding of the variant Plasmodium falciparum circumsporozoite protein CD8+ CTL epitope in naturally exposed Kenyan adults.

In this study, we have investigated the extent of natural polymorphism in the CD8+ cytotoxic T lymphocyte (CTL) determinant (amino acids 368-390) of circumsporozoite (CS) protein of Plasmodium falciparum field isolates from a holoendemic region of Kenya, and determined how this variation affects the CTL reactivities in clinically immune adults and binding specificities to human histocompatibility leukocyte antigen (HLA)-B35. Among the eight variant sequences that were found in this region, four were new and not seen in parasites from other geographical regions. When synthetic peptides corresponding to the eight variants were used to test the presence of CTL response in different donors, a different spectrum of CTL reactivity to these variants was noticed. While CTL from some donors recognized the P1 sequence (the most prevalent type of sequence) but not P8 (another major variant), other donors showed a reverse pattern of reactivity. Although none of the donors was able to recognize all the variants, CTL responses to all the eight variant sequences were found in this population. An octamer peptide with P1 sequence KPKDELDY in this polymorphic determinant was known to bind HLA-B35. When we tested the effect of natural variation in this octamer sequence on HLA-B35 binding, it became evident that SP13 with D --> N substitution retained its binding specificity to HLA-B35. On the other hand, the SP12 octamer sequence which had two substitutions did not bind HLA-B35. The most interesting finding was the observation that a D --> G substitution at position 374 rescued the binding ability of SP14, which otherwise could not bind to this HLA molecule due to E --> Q amino acid substitution at position 372. To our knowledge, this is the first demonstration showing that a natural polymorphism can rescue the binding specificity to an HLA-class I molecule that was lost due to another natural amino acid substitution. Altogether, these results demonstrate that natural polymorphism in the CS protein affects both the CTL reactivity and the ability to bind to HLA-B35.

Adult↗
PubMed1997

Autologous lymphoblastoid cell lines stably transfected with Plasmodium falciparum circumsporozoite protein as targets in cytotoxic T-lymphocyte assays.

To produce cell lines that can be used as a continuous source of antigen presenting cells for stimulating T-cell lines and clones and as targets in cytotoxic T-lymphocyte (CTL) assays, we used a retroviral vector with a simian virus (SV40) early promotor to transfer a Plasmodium falciparum circumporozoite (PfCSP) gene into human EBV transformed B-lymphoblastoid cell lines (B-LCL). We herein report successful, stable transfection and cell surface expression of this gene, as confirmed by PCR, Western blot analysis and immunoelectron microscopy. One of three successfully transfected autologous cell lines expressed PfCSP on the cell surface and was lysed by CD8+ T-cell dependent CTL from a donor volunteer who had been immunized with irradiated P. falciparum sporozoites. Such cell lines should provide excellent tools for characterizing human CD8+ T-cell responses against Plasmodium sp. proteins.

Animals↗
PubMed1997

Multi-gene vaccination against malaria: A multistage, multi-immune response approach.

An ideal malaria vaccine will induce immune responses against each stage of the Plasmodium spp life cycle. During its complicated life cycle, the parasite exists extracellularly in the host's bloodstream, within cells that express major histocompatibility complex (MHC) molecules (hepatocytes), within cells that do not express MHC molecules (erythrocytes) and within the mosquito vector. Different arms of the immune system are required to attack the parasite at the different stages. Therefore, a multistage vaccine must be a multi-immune response vaccine. In addition, given the unique antigenicities of the different stages of the life cycle, implicit in this definition is that the vaccine be multivalent. Here, Denise Doolan and Stephen Hoffman present the rationale for developing a multistage, multivalent, multi-immune response malaria vaccine and explain why, among currently available technologies, DNA vaccines may offer the best prospect for success.

Journal Article↗
PubMed1997

Strategy for development of a pre-erythrocytic Plasmodium falciparum DNA vaccine for human use.

Data generated in the Plasmodium yoelii rodent model indicated that plasmid DNA vaccines encoding the P.yoelii circumsporozoite protein (PyCSP) or 17 kDa hepatocyte erythrocyte protein (PyHEP17) were potent inducers of protective CD8+ T cell responses directed against infected hepatocytes. Immunization with a mixture of these plasmids circumvented the genetic restriction of protective immunity and induced additive protection. A third DNA vaccine encoding the P. yoelii sporozoite surface protein 2 (PySSP2) also induced protection. The P. falciparum genes encoding the homologues of these three protective P. yoelii antigens as well as another P. falciparum gene encoding a protein that is expressed in infected hepatocytes have been chosen for the development of a human vaccine. The optimal plasmid constructs for human use will be selected on the basis of immunogenicity data generated in mice and nonhuman primates. We anticipate that optimization of multi-gene P. falciparum DNA vaccines designed to protect against malaria by inducing CD8+ T cells that target infected hepatocytes will require extensive clinical trials during the coming years.

Animals↗
PubMed1997

Potent immunogenic short linear peptide constructs composed of B cell epitopes and Pan DR T helper epitopes (PADRE) for antibody responses in vivo.

Induction of humoral immune responses against protein antigen requires that two independent signals be delivered to B cells. It is currently assumed that simple monovalent synthetic peptides would not be effective immunogens for antibody responses because they would not be anticipated to effectively generate the necessary signals unless conjugated to a complex carrier system. In this study, the immunogenicity of short linear peptide constructs comprising Plasmodium vivax B cell epitopes (PVB) and non-natural Pan-DR T helper cell epitopes (PADRE) was assessed in mice and compared to other types of antigen constructs. The 33-residue PADRE-PVB linear constructs were highly immunogenic and induced responses comparable to those obtained with the multiple antigen peptides (MAP) constructs, both in terms of absolute titers and quality of antibody responses. The anti-PVB antibody responses were of long duration, composed mostly of IgG and reactive with intact sporozoites. The PADRE-PVB constructs were immunogenic when formulated in adjuvants such as Alum and Montanide ISA 51 underlining the relevance of these findings for vaccine development.

Adjuvants, Immunologic↗
PubMed1997

DNA inoculation with a plasmid vector carrying the hepatitis E virus structural protein gene induces immune response in mice.

The plasmid construct, pJHEV, containing the full-length open reading frame 2 (ORF-2) of hepatitis E virus (HEV) gene, expresses the HEV structural protein ORF-2 in Cos-7 cells under the control of a hCMV promoter. No ORF-2 protein could be detected in Cos-7 cells transfected with either vector alone or with a vector containing the ORF-2 of HEV in the incorrect orientation. The successful construct was further tested in BALB/c (H-2d) mice for the induction of an ORF-2 specific immune response. Intramuscular (i.m.) immunization of mice pretreated 24 h earlier with bupivacaine with the naked DNA construct elicited a humoral immune response in 80% and 100% of two separate groups of mice, respectively. No anti-ORF-2 responses were observed in mice immunized with the vector only. Sera from mice injected with pJHEV specifically recognized HEV ORF-2 structural protein expressed in recombinant baculovirus in an enzyme-linked-immunosorbent assay (ELISA) and Western blot. Anti-ORF-2 serum titers peaked at ca 1:5000 in the ELISA and 1:1000 in the Western blot. These titers have remained constant for over 12 months after the last boost of pJHEV. To our knowledge, this is the first report of the use of DNA-based immunization for the generation of an immune response to a HEV structural protein.

Animals↗