Identification of<i>Plasmodium falciparum</i>antigens by antigenic analysis of genomic and proteomic data

Doolan, Denise L.; Southwood, Scott; Freilich, Daniel; Sidney, John; Graber, Norma L.; Shatney, Lori; Bebris, Lolita; Florens, Laurence; Dobaño, Carlota; Witney, Adam A.; Appella, Ettore; Hoffman, Stephen L.; Yates, John R.; Carucci, Daniel J.; Sette, Alessandro

doi:10.1073/pnas.1633254100

Cited by 228 publications

(257 citation statements)

References 40 publications

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“…Each predicted ORF of the VACV Western Reserve strain (VACV-WR) was analyzed by using previously described algorithms (14). Peptides predicted to bind with an IC 50 Ͻ 100 nM were selected for study.…”

Section: Methodsmentioning

confidence: 99%

“…A more extensive definition of the antigens and the epitopes contained within VACV is necessary to (i) enable adequate immunogenicity testing of new vaccine candidates, (ii) help elucidate the correlates of protection offered by the current vaccine, and (iii) understand human cellular immune responses to complex viruses. To identify viral epitopes recognized by human CTL we employ a multistep process that begins with computational prediction of peptides that have the capacity of binding various HLA molecules (14,15).…”

mentioning

confidence: 99%

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HLA class I-restricted responses to vaccinia recognize a broad array of proteins mainly involved in virulence and viral gene regulation

Oseroff

Kos

Bui

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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We have analyzed by ex vivo ELISPOT the anti-vaccinia cytotoxic T lymphocyte responses of peripheral blood mononuclear cells from humans vaccinated with Dryvax vaccine. More than 6,000 peptides from 258 putative vaccinia ORFs predicted to bind the common molecules of the HLA A1, A2, A3, A24, B7, and B44 supertypes were screened with peripheral blood mononuclear cells of 31 vaccinees. A total of 48 epitopes derived from 35 different vaccinia antigens were identified, some of which (B8R, D1R, D5R, C10L, C19L, C7L, F12, and O1L) were recognized by multiple donors and contain multiple epitopes recognized in the context of different HLA types. The antigens recognized tend to be >100 residues in length and are expressed predominantly in the early phases of infection, although some late antigens were also recognized. Viral genome regulation and virulence factor were recognized most frequently, whereas few structural proteins were immunogenic. Finally, most epitopes were highly conserved among vaccinia virus Western Reserve, variola major and modified vaccinia Ankara, supporting their potential use in vaccine and diagnostic applications.immunodominance ͉ smallpox vaccination ͉ virulence factors

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

HLA class I-restricted responses to vaccinia recognize a broad array of proteins mainly involved in virulence and viral gene regulation

Oseroff

Kos

Bui

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

139

178

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“…The lack of targets may reflect technical limitations in antigen discovery, antigenic bias due to homologous sporozoite vaccination, lack of late-stage antigen expression, and/or CSP immunodominance. Traditionally, pooled synthetic peptides are used in IFN-γ (IFNγ) enzyme-linked immunosorbent spot (ELISPOT) assays to identify T-cell targets (29), but even with pooling strategies (30), these approaches require complex, expensive peptide libraries. In lieu of peptides, plasmidencoded parasite genes can be transfected into antigen-presenting cells (APCs) (31), but this approach is low throughput and difficult because of the A/T-richness of Plasmodial genes.…”

mentioning

confidence: 99%

A T-cell response to a liver-stagePlasmodiumantigen is not boosted by repeated sporozoite immunizations

Murphy

Kas²,

Stone³

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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Development of an antimalarial subunit vaccine inducing protective cytotoxic T lymphocyte (CTL)-mediated immunity could pave the way for malaria eradication. Experimental immunization with sporozoites induces this type of protective response, but the extremely large number of proteins expressed by Plasmodium parasites has so far prohibited the identification of sufficient discrete T-cell antigens to develop subunit vaccines that produce sterile immunity. Here, using mice singly immunized with Plasmodium yoelii sporozoites and high-throughput screening, we identified a unique CTL response against the parasite ribosomal L3 protein. Unlike CTL responses to the circumsporozoite protein (CSP), the population of L3-specific CTLs was not expanded by multiple sporozoite immunizations. CSP is abundant in the sporozoite itself, whereas L3 expression does not increase until the liver stage. The response induced by a single immunization with sporozoites reduces the parasite load in the liver so greatly during subsequent immunizations that L3-specific responses are only generated during the primary exposure. Functional L3-specific CTLs can, however, be expanded by heterologous prime-boost regimens. Thus, although repeat sporozoite immunization expands responses to preformed antigens like CSP that are present in the sporozoite itself, this immunization strategy may not expand CTLs targeting parasite proteins that are synthesized later. Heterologous strategies may be needed to increase CTL responses across the entire spectrum of Plasmodium liver-stage proteins.

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“…During assay development, a fluorescein-labeled anti-HA mouse mAb (Boehringer Mannheim) was used to probe HA-Epi-TAP transfected, fixed and permeabilized cells to verify the effectiveness of the transfection procedure. Six well-characterized P. falciparum antigens (PfCSP, PfSSP2, PfExp1, PfMSP1-42kD, PfAMA1 or PfEBA175-RII) were used to develop the assay, and a panel of six novel proteins (PFC0450w, PFC0700c, PFC0210c, PF14_0074, PF14_0751, PFL0800c) identified from the P. falciparum genomic sequence database [12,28] were screened using the refined assay.…”

Section: In Vitro Humoral Immunoscreening Assaymentioning

confidence: 99%

“…A panel of 9 genes (PFL0800c, PFI0165c, PFA0515w, PFD0425w, PFC0210c, PFC0450w, PF14_0751, PF14_0074, PFC0700c) was selected from the genomic sequence database for TAP amplification. These genes represented a subset of the 27 putative proteins assayed for recognition by human peripheral blood mononuclear cells derived from individuals experimentally immunized with radiation attenuated P. falciparum sporozoites [28]. The initial selection of these antigens was based on data derived from MudPIT of P. falciparum sporozoite preparations [34].…”

Section: Adaptation Of Standard Parameters For Epi-tap To Genome-widementioning

confidence: 99%

Transcriptionally active PCR for antigen identification and vaccine development: In vitro genome-wide screening and in vivo immunogenicity

Regis

Dobaño

Quiñones-Olson

et al. 2008

Molecular and Biochemical Parasitology

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We have evaluated a technology called Transcriptionally Active PCR (TAP) for high throughput identification and prioritization of novel target antigens from genomic sequence data using the Plasmodium parasite, the causative agent of malaria, as a model. First, we adapted the TAP technology for the highly AT-rich Plasmodium genome, using well-characterized P. falciparum and P. yoelii antigens and a small panel of uncharacterized open reading frames from the P. falciparum genome sequence database. We demonstrated that TAP fragments encoding six wellcharacterized P. falciparum antigens and five well-characterized P. yoelii antigens could be amplified in an equivalent manner from both plasmid DNA and genomic DNA templates, and that uncharacterized open reading frames could also be amplified from genomic DNA template. Second, we showed that the in vitro expression of the TAP fragments was equivalent or superior to that of supercoiled plasmid DNA encoding the same antigen. Third, we evaluated the in vivo immunogenicity of TAP fragments encoding a subset of the model P. falciparum and P. yoelii antigens. We found that antigen-specific antibody and cellular immune responses induced by the TAP fragments in mice were equivalent or superior to those induced by the corresponding plasmid DNA vaccines. Finally, we developed and demonstrated proof-of-principle for an in vitro humoral immunoscreening assay for down-selection of novel target antigens. These data support the potential of a TAP approach for rapid high throughput functional screening and identification of potential candidate vaccine antigens from genomic sequence data.

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Identification ofPlasmodium falciparumantigens by antigenic analysis of genomic and proteomic data

Cited by 228 publications

References 40 publications

HLA class I-restricted responses to vaccinia recognize a broad array of proteins mainly involved in virulence and viral gene regulation

HLA class I-restricted responses to vaccinia recognize a broad array of proteins mainly involved in virulence and viral gene regulation

A T-cell response to a liver-stagePlasmodiumantigen is not boosted by repeated sporozoite immunizations

Transcriptionally active PCR for antigen identification and vaccine development: In vitro genome-wide screening and in vivo immunogenicity

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