Singular Fermi Liquids

An introductory survey of the theoretical ideas and calculations and the experimental results which depart from Landau Fermi-liquids is presented. Common themes and possible routes to the singularities leading to the breakdown of Landau Fermi liquids are categorized following an elementary discussion of the theory. Soluble examples of Singular Fermi liquids (often called Non-Fermi liquids) include models of impurities in metals with special symmetries and one-dimensional interacting fermions. A review of these is followed by a discussion of Singular Fermi liquids in a wide variety of experimental situations and theoretical models. These include the effects of low-energy collective fluctuations, gauge fields due either to symmetries in the hamiltonian or possible dynamically generated symmetries, fluctuations around quantum critical points, the normal state of high temperature superconductors and the two-dimensional metallic state. For the last three systems, the principal experimental results are summarized and the outstanding theoretical issues highlighted.Comment: 170 pages; submitted to Physics Reports; a single pdf file with high quality figures is available from http://www.lorentz.leidenuniv.nl/~saarloo

Production, quality control, stability, and potency of cGMP-produced Plasmodium falciparum RH5.1 protein vaccine expressed in Drosophila S2 cells

Plasmodium falciparum reticulocyte-binding protein homolog 5 (PfRH5) is a leading asexual blood-stage vaccine candidate for malaria. In preparation for clinical trials, a full-length PfRH5 protein vaccine called “RH5.1” was produced as a soluble product under cGMP using the ExpreS2 platform (based on a Drosophila melanogaster S2 stable cell line system). Following development of a highproducing monoclonal S2 cell line, a master cell bank was produced prior to the cGMP campaign. Culture supernatants were processed using C-tag affinity chromatography followed by size exclusion chromatography and virus-reduction filtration. The overall process yielded >400 mg highly pure RH5.1 protein. QC testing showed the MCB and the RH5.1 product met all specified acceptance criteria including those for sterility, purity, and identity. The RH5.1 vaccine product was stored at −80 °C and is stable for over 18 months. Characterization of the protein following formulation in the adjuvant system AS01B showed that RH5.1 is stable in the timeframe needed for clinical vaccine administration, and that there was no discernible impact on the liposomal formulation of AS01B following addition of RH5.1. Subsequent immunization of mice confirmed the RH5.1/AS01B vaccine was immunogenic and could induce functional growth inhibitory antibodies against blood-stage P. falciparum in vitro. The RH5.1/AS01B was judged suitable for use in humans and has since progressed to phase I/IIa clinical trial. Our data support the future use of the Drosophila S2 cell and C-tag platform technologies to enable cGMP-compliant biomanufacture of other novel and “difficult-to-express” recombinant protein-based vaccines

Development of adjuvanted multi-antigen liver-stage malaria vaccines

Despite promising progress in malaria vaccine development in recent years, an efficacious subunit vaccine against P. falciparum remains to be licensed and deployed. The most advanced candidates in clinical development focus on two pre-erythrocytic antigens (CSP and ME-TRAP), but many more immunogenic antigens have been recently identified. The work described in this thesis aimed to improve on the immunogenicity and efficacy of the leading liver-stage vaccine candidate (ChAd63-MVA ME-TRAP), which is known to confer protection by eliciting high levels of antigen-specific CD8+ T cells. To achieve this, two different strategies were pursued. First, several prime-boost regimens with vectors encoding combinations of two liver-stage antigens were investigated. When mixed, vectors expressing LSA1 and LSAP2 conferred highest levels of protective efficacy in mice and were therefore considered for inclusion in the final second generation viral vectored liver-stage malaria vaccine. Second, the MHC class II invariant chain (Ii) was developed as a molecular adjuvant. Immunogenicity analyses of ChAd63 encoding ME-TRAP fused to several different versions of Ii showed that the transmembrane domain of Ii has the ability to strongly increase antigen-specific CD8+ T-cell responses, even in the absence of the rest of the Ii protein. This finding may lead to the discovery of numerous similar adjuvants (such as the transmembrane domain of the Newcastle disease virus fusion protein). Furthermore, experiments showed that the Ii chain sequence can also be xenogenised without losing adjuvanticity. Both strategies have been combined in numerous novel adjuvanted multi-antigen vaccines, which were ranked for immunogenicity and efficacy in inbred and outbred mice. The viral vectors of the best combination (ChAdOX1-sharkTM/Ii-LSA1-LSAP2 and MVA-tPA-LSA1-LSAP2) are currently being produced to GMP grade and will progress to initial clinical trials in early 2017.</p

Development of adjuvanted multi-antigen liver-stage malaria vaccines

Despite promising progress in malaria vaccine development in recent years, an efficacious subunit vaccine against P. falciparum remains to be licensed and deployed. The most advanced candidates in clinical development focus on two pre-erythrocytic antigens (CSP and ME-TRAP), but many more immunogenic antigens have been recently identified. The work described in this thesis aimed to improve on the immunogenicity and efficacy of the leading liver-stage vaccine candidate (ChAd63-MVA ME-TRAP), which is known to confer protection by eliciting high levels of antigen-specific CD8+ T cells. To achieve this, two different strategies were pursued. First, several prime-boost regimens with vectors encoding combinations of two liver-stage antigens were investigated. When mixed, vectors expressing LSA1 and LSAP2 conferred highest levels of protective efficacy in mice and were therefore considered for inclusion in the final second generation viral vectored liver-stage malaria vaccine. Second, the MHC class II invariant chain (Ii) was developed as a molecular adjuvant. Immunogenicity analyses of ChAd63 encoding ME-TRAP fused to several different versions of Ii showed that the transmembrane domain of Ii has the ability to strongly increase antigen-specific CD8+ T-cell responses, even in the absence of the rest of the Ii protein. This finding may lead to the discovery of numerous similar adjuvants (such as the transmembrane domain of the Newcastle disease virus fusion protein). Furthermore, experiments showed that the Ii chain sequence can also be xenogenised without losing adjuvanticity. Both strategies have been combined in numerous novel adjuvanted multi-antigen vaccines, which were ranked for immunogenicity and efficacy in inbred and outbred mice. The viral vectors of the best combination (ChAdOX1-sharkTM/Ii-LSA1-LSAP2 and MVA-tPA-LSA1-LSAP2) are currently being produced to GMP grade and will progress to initial clinical trials in early 2017.</p

Identification of immunodominant responses to the Plasmodium falciparum antigens PfUIS3, PfLSA1 and PfLSAP2 in multiple strains of mice

Malaria, caused by the Plasmodium parasite, remains a serious global public health concern. A vaccine could have a substantial impact on eliminating this disease, alongside other preventative measures. We recently described the development of three novel, viral vectored vaccines expressing either of the antigens PfUIS3, PfLSA1 and PfLSAP2. Each vaccination regimen provided high levels of protection against chimeric parasite challenge in a mouse model, largely dependent on CD8+ T cells. In this study we aimed to further characterize the induced cellular immune response to these vaccines. We utilized both the IFNγ enzyme-linked immunosorbent spot assay and intracellular cytokine staining to achieve this aim. We identified immunodominant peptide responses for CD4+ and CD8+ T cells for each of the antigens in BALB/c, C57BL/6 and HLA-A2 transgenic mice, creating a useful tool for researchers for subsequent study of these antigens. We also compared these immunodominant peptides with those generated from epitope prediction software, and found that only a small proportion of the large number of epitopes predicted by the software were identifiable experimentally. Furthermore, we characterized the polyfunctionality of the induced CD8+ T cell responses. These findings contribute to our understanding of the immunological mechanisms underlying these protective vaccines, and provide a useful basis for the assessment of these and related vaccines as clinical constructs

Identification of immunodominant responses to the Plasmodium falciparum antigens PfUIS3, PfLSA1 and PfLSAP2 in multiple strains of mice

Malaria, caused by the Plasmodium parasite, remains a serious global public health concern. A vaccine could have a substantial impact on eliminating this disease, alongside other preventative measures. We recently described the development of three novel, viral vectored vaccines expressing either of the antigens PfUIS3, PfLSA1 and PfLSAP2. Each vaccination regimen provided high levels of protection against chimeric parasite challenge in a mouse model, largely dependent on CD8+ T cells. In this study we aimed to further characterize the induced cellular immune response to these vaccines. We utilized both the IFNγ enzyme-linked immunosorbent spot assay and intracellular cytokine staining to achieve this aim. We identified immunodominant peptide responses for CD4+ and CD8+ T cells for each of the antigens in BALB/c, C57BL/6 and HLA-A2 transgenic mice, creating a useful tool for researchers for subsequent study of these antigens. We also compared these immunodominant peptides with those generated from epitope prediction software, and found that only a small proportion of the large number of epitopes predicted by the software were identifiable experimentally. Furthermore, we characterized the polyfunctionality of the induced CD8+ T cell responses. These findings contribute to our understanding of the immunological mechanisms underlying these protective vaccines, and provide a useful basis for the assessment of these and related vaccines as clinical constructs