Dr Christine S Rollier

Research Area: Immunology
Technology Exchange: Cell sorting, Cellular immunology, Flow cytometry and Vaccine production and evaluation
Scientific Themes: Immunology & Infectious Disease and Clinical Trials & Epidemiology
Keywords: Vaccines, Viral vectors, Bacterial infections, Humoral immunity, cellular immunity, Meningitis
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Bacterial infections are a leading cause of death and disease worldwide, dramatically affecting children, and vaccines represent the best hope for preventing their severe consequences. In particular, Neisseria meningitidis is the cause of around 500,000 cases of meningitis and septicaemia every year, disproportionately affecting children under 2 years. The case-fatality rate is 10% in resource-rich settings, and has not decreased significantly since the 1950s. 30% of survivors suffer severe long-term disability including deafness, amputation and cognitive impairment. Vaccination is the optimal way to reduce the mortality and morbidity from this disease. Our objective is to develop a new vaccine against capsular group-B meningococcal disease (MenB), in order to address the need for an improved and more cost-effective vaccine which has lower manufacturing costs, higher immunogenicity, longer duration of protection and requires a single injection to protect infants, as compared to the currently available vaccines. Two vaccine candidates have recently been licensed, but they do not provide complete protection against MenB, especially in infants, who are at most risk of this devastating disease. Furthermore, these vaccines require multiple doses, which increases the cost of vaccine implementation. Development of alternative vaccines is therefore required.

The uptake of existing vaccines as well as their safe administration could be greatly enhanced by needle-free immunizations, and by combining vaccines in order to decrease the number of injections. Therefore we need to create more effective and multivalent vaccines, and to understand the reasons why some vaccines are more efficient than others, especially in children.

Our projects aims to:

  • Develop new vaccines against bacterial and pediatric diseases such as capsular group B Neisseria meningitidis and Bordetella pertussis. This includes characterization of their immunogenicity, identification of the optimal vaccine regimen, and evaluation of their protective efficacy. We perform pre-clinical vaccine development up to early phase clinical trials.
  • Evaluate alternative vaccine delivery systems (such as needle-free).
  • Inform vaccine development by investigating immune mechanisms underlying infection and immunity, improvements and combination of vaccines.

Name Department Institution Country
Professor Adrian VS Hill Jenner Institute Oxford University, Old Road Campus Research Building United Kingdom
Dr David Wyllie Jenner Institute Oxford University, Henry Wellcome Building for Molecular Physiology United Kingdom
Professor Arturo Reyes-Sandoval Jenner Institute Oxford University, Henry Wellcome Building for Molecular Physiology United Kingdom
Dr Anita Milicic Jenner Institute Oxford University, Old Road Campus Research Building United Kingdom
Professor Martin Maiden Jenner Institute Oxford University, Zoology United Kingdom
Dr Ian Feavers National Institute for Biological Standards and Controls United Kingdom
Dr Fergal Hill Imaxio France
Dr Jeremy Derrick University of Manchester United Kingdom
Professor Ray Owens Structural Biology Oxford University, Oxford Protein Production Facility United Kingdom
Professor Dan Granoff Children Hospital Oakland Research Institute United States
Dr Peter Beernink Children Hospital Oakland Research Institute United States
Dr Barbara Bolgiano National Institute for Biological Standards and Controls United Kingdom
Norheim G, Mueller JE, Njanpop-Lafourcade B-M, Delrieu I, Findlow H, Borrow R, Xie O, Nagaputra J, Ramasamy R, Dold C et al. 2018. Natural immunity against capsular group X N. meningitidis following an outbreak in Togo, 2007. Vaccine, 36 (10), pp. 1297-1303. | Show Abstract | Read more

BACKGROUND: Capsular group X N. meningitidis (MenX) has emerged as a cause of localized disease outbreaks in sub-Saharan Africa, but the human immune response following exposure to MenX antigens is poorly described. We therefore assessed the natural immunity against MenX in individuals who were living in an area affected by a MenX outbreak during 2007 in Togo, West Africa. During 2009, 300 healthy individuals (100 aged 3-5 years, 100 aged 13-19 years and 100 aged 20-25 years) were included in the study, and serum responses were compared with sera from age-matched controls from the U.K. and Burkina Faso. METHODS: MenX serum bactericidal antibody (SBA) was measured using rabbit complement, and antibodies against MenX polysaccharide (XPS) and outer membrane vesicles (XOMVs) were quantified by ELISA. RESULTS: The proportion of Togolese individuals with an SBA titer of ≥8 against the MenX strain was 29% (95% confidence interval (CI) 18-41) among those aged 3-5 years, 34% (95% CI 9-60) among those aged 13-19 years and 32% (95% CI 24-40) among those aged 20-25 years. These were significantly higher than observed in the control populations from the U.K (range 13-16%) and Burkina Faso (range 2-6%). CONCLUSION: In Togolese individuals, the concentration of serum IgG against XPS was higher among the two older age groups as compared to the youngest age group. Antibody concentrations against MenX PS correlated significantly with SBA titers. This supports further development of a MenX PS based conjugate vaccine. Further studies are needed to verify the ability of MenX PS to induce SBA in humans.

Flaxman A, van Diemen PM, Yamaguchi Y, Allen E, Lindemann C, Rollier CS, Milicic A, Wyllie DH. 2017. Development of persistent gastrointestinal S. aureus carriage in mice. Sci Rep, 7 (1), pp. 12415. | Show Abstract | Read more

One fifth to one quarter of the human population is asymptomatically, naturally and persistently colonised by Staphylococcus aureus. Observational human studies indicate that although the whole population is intermittently exposed, some individuals lose S. aureus rapidly. Others become persistent carriers, as assessed by nasal cultures, with many individuals colonised for decades. Current animal models of S. aureus colonisation are expensive and normally require antibiotics. Importantly, these animal models have not yet contributed to our poor understanding of the dichotomy in human colonisation status. Here, we identify a single strain of S. aureus found to be persistently colonising the gastrointestinal tract of BALB/c mice. Phylogenetic analyses suggest it diverged from a human ST15 lineage in the recent past. We show that murine carriage of this organism occurs in the bowel and nares, is acquired early in life, and can persist for months. Importantly, we observe the development of persistent and non-persistent gastrointestinal carriage states in genetically identical mice. We developed a needle- and antibiotic-free model in which we readily induced S. aureus colonisation of the gastrointestinal tract experimentally by environmental exposure. Using our experimental model, impact of adaptive immunity on S. aureus colonisation could be assessed. Vaccine efficacy to eliminate colonisation could also be investigated using this model.

Weissmueller NT, Marsay L, Schiffter HA, Carlisle RC, Rollier CS, Prud'homme RK, Pollard AJ. 2017. Alternative vaccine administration by powder injection: Needle-free dermal delivery of the glycoconjugate meningococcal group Y vaccine. PLoS One, 12 (8), pp. e0183427. | Show Abstract | Read more

Powder-injectors use gas propulsion to deposit lyophilised drug or vaccine particles in the epidermal and sub epidermal layers of the skin. We prepared dry-powder (Tg = 45.2 ± 0.5°C) microparticles (58.1 μm) of a MenY-CRM197 glyconjugate vaccine (0.5% wt.) for intradermal needle-free powder injection (NFPI). SFD used ultrasound atomisation of the liquid vaccine-containing excipient feed, followed by lyophilisation above the glass transition temperature (Tg' = - 29.9 ± 0.3°C). This resulted in robust particles (density~ 0.53 ±0.09 g/cm3) with a narrow volume size distribution (mean diameter 58.1 μm, and span = 1.2), and an impact parameter (ρvr ~ 11.5 kg/m·s) sufficient to breach the Stratum corneum (sc). The trehalose, manitol, dextran (10 kDa), dextran (150 kDa) formulation, or TMDD (3:3:3:1), protected the MenY-CRM197 glyconjugate during SFD with minimal loss, no detectable chemical degradation or physical aggregation. In a capsular group Y Neisseria meningitidis serum bactericidal assay (SBA) with human serum complement, the needle free vaccine, which contained no alum adjuvant, induced functional protective antibody responses in vivo of similar magnitude to the conventional vaccine injected by hypodermic needle and syringe and containing alum adjuvant. These results demonstrate that needle-free vaccination is both technically and immunologically valid, and could be considered for vaccines in development.

Milicic A, S Rollier C, Tang CK, Longley R, Hill AVS, Reyes-Sandoval A. 2017. Adjuvanting a viral vectored vaccine against pre-erythrocytic malaria. Sci Rep, 7 (1), pp. 7284. | Show Abstract | Read more

The majority of routinely given vaccines require two or three immunisations for full protective efficacy. Single dose vaccination has long been considered a key solution to improving the global immunisation coverage. Recent infectious disease outbreaks have further highlighted the need for vaccines that can achieve full efficacy after a single administration. Viral vectors are a potent immunisation platform, benefiting from intrinsic immuno-stimulatory features while retaining excellent safety profile through the use of non-replicating viruses. We investigated the scope for enhancing the protective efficacy of a single dose adenovirus-vectored malaria vaccine in a mouse model of malaria by co-administering it with vaccine adjuvants. Out of 11 adjuvants, only two, Abisco®-100 and CoVaccineHTTM, enhanced vaccine efficacy and sterile protection following malaria challenge. The CoVaccineHTTM adjuvanted vaccine induced significantly higher proportion of antigen specific central memory CD8+ cells, and both adjuvants resulted in increased proportion of CD8+ T cells expressing the CD107a degranulation marker in the absence of IFNγ, TNFα and IL2 production. Our results show that the efficacy of vaccines designed to induce protective T cell responses can be positively modulated with chemical adjuvants and open the possibility of achieving full protection with a single dose immunisation.

Diemen PMV, Leneghan DB, Brian IJ, Miura K, Long CA, Milicic A, Biswas S, Rollier CS, Wyllie DH. 2017. The S. aureus 4-oxalocrotonate tautomerase SAR1376 enhances immune responses when fused to several antigens. Sci Rep, 7 (1), pp. 1745. | Show Abstract | Read more

A persistent goal of vaccine development is the enhancement of the immunogenicity of antigens while maintaining safety. One strategy involves alteration of the presentation of the antigen by combining antigens with a multimeric scaffold. Multi-antigen vaccines are under development, and there are presently far more candidate antigens than antigen scaffolding strategies. This is potentially problematic, since prior immunity to a scaffold may inhibit immune responses to the antigen-scaffold combination. In this study, a series of domains from S. aureus which have been shown to crystallise into multimeric structures have been examined for their scaffolding potential. Of these domains, SAR1376, a 62 amino acid member of the 4-oxalocrotonate tautomerase (4-OT) family, was pro-immunogenic in mice when fused to a range of pathogen antigens from both S. aureus and P. falciparum, and delivered by either DNA vaccination, viral vector vaccines or as protein-in-adjuvant formulations. The adjuvant effect did not depend on enzymatic activity, but was abrogated by mutations disrupting the hexameric structure of the protein. We therefore propose that SAR1376, and perhaps other members of the 4-OT protein family, represent very small domains which can be fused to a wide range of antigens, enhancing immune responses against them.

Giuntini S, Lujan E, Gibani MM, Dold C, Rollier CS, Pollard AJ, Granoff DM. 2017. Serum Bactericidal Antibody Responses of Adults Immunized with the MenB-4C Vaccine against Genetically Diverse Serogroup B Meningococci. Clin Vaccine Immunol, 24 (1), pp. e00430-16-e00430-16. | Show Abstract | Read more

MenB-4C is a meningococcal vaccine for the prevention of serogroup B disease. The vaccine contains factor H binding protein (FHbp) and three other antigens that can elicit serum bactericidal antibodies (SBA). For vaccine licensure, efficacy was inferred from the SBA responses against three antigen-specific indicator strains. The relation between those results and broad protection against circulating genetically diverse strains is not known. Twenty adults were immunized with two doses of MenB-4C given 1 to 2 months apart. SBA activity against 3 reference strains and 15 serogroup B test strains (6 from college outbreaks) was measured. Compared to the preimmunization titers, 70%, 95%, and 95% of subjects had ≥4-fold increases in the titers of anti-PorA P1.4, anti-NadA, and anti-FHbp antibodies against the reference strains, respectively. In contrast, only 25 to 45% of the subjects had ≥4-fold increases in responses to 10 of the 15 test strains, including 8 that expressed one to three of the antigens in the vaccine. At 1 month, the majority of subjects with <4-fold titer increases had serum titers of ≥1:4, which are considered sufficient for protection. However, the titers against four strains declined to <1:4 by 4 to 6 months in one-third to greater than 50% of the subjects tested. Clinically relevant isolates are often more resistant to SBA than the indicator strains used to measure antigen-specific SBA. A working model is that the percentage of subjects with titers of ≥1:4 at 1 month postimmunization correlates with short-term protection against that strain, whereas the percentage of subjects with ≥4-fold titer increases represents a more robust response. (The protocol used at the Oxford Vaccine Group has been registered at ClinicalTrials.gov under registration no. NCT02398396.).

Rollier CS, Verschoor EJ, Verstrepen BE, Drexhage JAR, Paranhos-Baccala G, Liljeström P, Sutter G, Arribillaga L, Lasarte JJ, Bartosch B et al. 2016. T- and B-cell responses to multivalent prime-boost DNA and viral vectored vaccine combinations against hepatitis C virus in non-human primates. Gene Ther, 23 (10), pp. 753-759. | Show Abstract | Read more

Immune responses against multiple epitopes are required for the prevention of hepatitis C virus (HCV) infection, and the progression to phase I trials of candidates may be guided by comparative immunogenicity studies in non-human primates. Four vectors, DNA, SFV, human serotype 5 adenovirus (HuAd5) and Modified Vaccinia Ankara (MVA) poxvirus, all expressing hepatitis C virus Core, E1, E2 and NS3, were combined in three prime-boost regimen, and their ability to elicit immune responses against HCV antigens in rhesus macaques was explored and compared. All combinations induced specific T-cell immune responses, including high IFN-γ production. The group immunized with the SFV+MVA regimen elicited higher E2-specific responses as compared with the two other modalities, while animals receiving HuAd5 injections elicited lower IL-4 responses as compared with those receiving MVA. The IFN-γ responses to NS3 were remarkably similar between groups. Only the adenovirus induced envelope-specific antibody responses, but these failed to show neutralizing activity. Therefore, the two novel regimens failed to induce superior responses as compared with already existing HCV vaccine candidates. Differences were found in response to envelope proteins, but the relevance of these remain uncertain given the surprisingly poor correlation with immunogenicity data in chimpanzees, underlining the difficulty to predict efficacy from immunology studies.

Rollier CS, Hill AVS, Reyes-Sandoval A. 2016. Influence of adenovirus and MVA vaccines on the breadth and hierarchy of T cell responses. Vaccine, 34 (38), pp. 4470-4474. | Show Abstract | Read more

Viral-vectored vaccines are in clinical development for several infectious diseases where T-cell responses can mediate protection, and responses to sub-dominant epitopes is needed. Little is known about the influence of MVA or adenoviral vectors on the hierarchy of the dominant and sub-dominant T-cell epitopes. We investigated this aspect in mice using a malaria immunogen. Our results demonstrate that the T-cell hierarchy is influenced by the timing of analysis, rather than by the vector after a single immunization, with hierarchy changing over time. Repeated homologous immunization reduced the breadth of responses, while heterologous prime-boost induced the strongest response to the dominant epitope, albeit with only modest response to the sub-dominant epitopes.

Ewer KJ, Lambe T, Rollier CS, Spencer AJ, Hill AV, Dorrell L. 2016. Viral vectors as vaccine platforms: from immunogenicity to impact. Curr Opin Immunol, 41 pp. 47-54. | Show Abstract | Read more

Viral vectors are the vaccine platform of choice for many pathogens that have thwarted efforts towards control using conventional vaccine approaches. Although the STEP trial encumbered development of recombinant human adenovirus vectors only a few years ago, replication-deficient simian adenoviruses have since emerged as a crucial component of clinically effective prime-boost regimens. The vectors discussed here elicit functionally relevant cellular and humoral immune responses, at extremes of age and in diverse populations. The recent Ebola virus outbreak highlighted the utility of viral vectored vaccines in facilitating a rapid response to public health emergencies. Meanwhile, technological advances in manufacturing to support scale-up of viral vectored vaccines have helped to consolidate their position as a leading approach to tackling 'old' and emerging infections.

Das S, Lindemann C, Young BC, Muller J, Österreich B, Ternette N, Winkler A-C, Paprotka K, Reinhardt R, Förstner KU et al. 2016. Natural mutations in a Staphylococcus aureus virulence regulator attenuate cytotoxicity but permit bacteremia and abscess formation. Proc Natl Acad Sci U S A, 113 (22), pp. E3101-E3110. | Show Abstract | Read more

Staphylococcus aureus is a major bacterial pathogen, which causes severe blood and tissue infections that frequently emerge by autoinfection with asymptomatically carried nose and skin populations. However, recent studies report that bloodstream isolates differ systematically from those found in the nose and skin, exhibiting reduced toxicity toward leukocytes. In two patients, an attenuated toxicity bloodstream infection evolved from an asymptomatically carried high-toxicity nasal strain by loss-of-function mutations in the gene encoding the transcription factor repressor of surface proteins (rsp). Here, we report that rsp knockout mutants lead to global transcriptional and proteomic reprofiling, and they exhibit the greatest signal in a genome-wide screen for genes influencing S. aureus survival in human cells. This effect is likely to be mediated in part via SSR42, a long-noncoding RNA. We show that rsp controls SSR42 expression, is induced by hydrogen peroxide, and is required for normal cytotoxicity and hemolytic activity. Rsp inactivation in laboratory- and bacteremia-derived mutants attenuates toxin production, but up-regulates other immune subversion proteins and reduces lethality during experimental infection. Crucially, inactivation of rsp preserves bacterial dissemination, because it affects neither formation of deep abscesses in mice nor survival in human blood. Thus, we have identified a spontaneously evolving, attenuated-cytotoxicity, nonhemolytic S. aureus phenotype, controlled by a pleiotropic transcriptional regulator/noncoding RNA virulence regulatory system, capable of causing S. aureus bloodstream infections. Such a phenotype could promote deep infection with limited early clinical manifestations, raising concerns that bacterial evolution within the human body may contribute to severe infection.

Whitehouse J, Flaxman A, Rollier C, O'Shea MK, Fallowfield J, Lindsay M, Gunner F, Knox K, Wyllie DH, Yamaguchi Y. 2016. Population variation in anti-S. aureus IgG isotypes influences surface protein A mediated immune subversion. Vaccine, 34 (15), pp. 1792-1799. | Show Abstract | Read more

BACKGROUND: Staphylococcus aureus is a pathogen which causes life-threatening infection, the incidence of which rises during adult life. This, together with the emergence of drug-resistant strains and the expansion of more susceptible elderly populations, represents the rationale for the ongoing development of S. aureus vaccines targeting adult populations. Humoral responses to S. aureus naturally develop early in life, influence susceptibility to infection, and potentially influence the effect of vaccination. Despite this, the nature of pre-existing anti-S. aureus antibodies in healthy adult populations is not fully characterised. METHODS: Immunoglobulin levels against S. aureus surface antigens were measured by a filter membrane enzyme-linked immunosorbent assay using fixed ΔSpA S. aureus as an antigen in serum samples obtained from three clinical cohorts comprising 133 healthy adult volunteers from 19 to 65 years of age. Functional capacity of antibody was also assessed, using antibody-mediated attachment of FITC-stained S. aureus to differentiated HL-60 cells. RESULTS: Wide variation in the concentrations of immunoglobulins recognising S. aureus surface antigens was observed among individuals in all three cohorts. There was a decline of anti-S. aureus IgG1 with age, and a similar trend was observed in IgM, but not in IgA or other IgG sub-classes. Antibody mediated bacterial attachment to cells was associated with IgG1 and IgG3 concentrations in serum. The presence of SpA on the bacterial cell surface reduced antibody-mediated binding of bacteria to phagocytes in serum with low, but not high, levels of naturally occurring anti-S. aureus IgG3 antibodies. CONCLUSIONS: Naturally acquired immunoglobulin responses to S. aureus are heterogeneous in populations and their concentrations alter during adulthood. Elevated IgG1 or IgG3 titres against S. aureus enhance S. aureus recognition by phagocytosis and may be correlates of natural protection and/or vaccine efficacy in adult populations.

Allen ER, van Diemen P, Yamaguchi Y, Lindemann C, Soilleux E, Rollier C, Hill F, Schneider J, Wyllie DH. 2016. MRI Based Localisation and Quantification of Abscesses following Experimental S. aureus Intravenous Challenge: Application to Vaccine Evaluation. PLoS One, 11 (5), pp. e0154705. | Show Abstract | Read more

PURPOSE: To develop and validate a sensitive and specific method of abscess enumeration and quantification in a preclinical model of Staphylococcus aureus infection. METHODS: S. aureus infected murine kidneys were fixed in paraformaldehyde, impregnated with gadolinium, and embedded in agar blocks, which were subjected to 3D magnetic resonance microscopy on a 9.4T MRI scanner. Image analysis techniques were developed, which could identify and quantify abscesses. The result of this imaging was compared with histological examination. The impact of a S. aureus Sortase A vaccination regime was assessed using the technique. RESULTS: Up to 32 murine kidneys could be imaged in a single MRI run, yielding images with voxels of about 25 μm3. S. aureus abscesses could be readily identified in blinded analyses of the kidneys after 3 days of infection, with low inter-observer variability. Comparison with histological sections shows a striking correlation between the two techniques: all presumptive abscesses identified by MRI were confirmed histologically, and histology identified no abscesses not evident on MRI. In view of this, simulations were performed assuming that both MRI reconstruction, and histology examining all sections of the tissue, were fully sensitive and specific at abscess detection. This simulation showed that MRI provided more sensitive and precise estimates of abscess numbers and volume than histology, unless at least 5 histological sections are taken through the long axis of the kidney. We used the MRI technique described to investigate the impact of a S. aureus Sortase A vaccine. CONCLUSION: Post mortem MRI scanning of large batches of fixed organs has application in the preclinical assessment of S. aureus vaccines.

Daniels-Treffandier H, de Nie K, Marsay L, Dold C, Sadarangani M, Reyes-Sandoval A, Langford PR, Wyllie D, Hill F, Pollard AJ, Rollier CS. 2016. Impact of Reducing Complement Inhibitor Binding on the Immunogenicity of Native Neisseria meningitidis Outer Membrane Vesicles. PLoS One, 11 (2), pp. e0148840. | Show Abstract | Read more

Neisseria meningitidis recruits host human complement inhibitors to its surface to down-regulate complement activation and enhance survival in blood. We have investigated whether such complement inhibitor binding occurs after vaccination with native outer membrane vesicles (nOMVs), and limits immunogenicity of such vaccines. To this end, nOMVs reactogenic lipopolysaccharide was detoxified by deletion of the lpxl1 gene (nOMVlpxl1). nOMVs unable to bind human complement factor H (hfH) were generated by additional deletions of the genes encoding factor H binding protein (fHbp) and neisserial surface protein A (NspA) (nOMVdis). Antibody responses elicited in mice with nOMVdis were compared to those elicited with nOMVlpxl1 in the presence of hfH. Results demonstrate that the administration of human fH to mice immunized with fHbp containing OMVlpxl1 decreased immunogenicity against fHbp (but not against the OMV as a whole). The majority of the OMV-induced bactericidal immune response (OMVlpxl1 or OMVdis) was versus PorA. Despite a considerable reduction of hfH binding to nOMVdis, and the absence of the vaccine antigen fHbp, immunogenicity in mice was not different from nOMVlpxl1, in the absence or presence of hfH (serum bactericidal titers of 1:64 vs 1:128 after one dose in the nOMVdis and nOMVlpxl1-immunized groups respectively). Therefore, partial inhibition of fH binding did not enhance immunity in this model.

Marsay L, Dold C, Green CA, Rollier CS, Norheim G, Sadarangani M, Shanyinde M, Brehony C, Thompson AJ, Sanders H et al. 2015. A novel meningococcal outer membrane vesicle vaccine with constitutive expression of FetA: A phase I clinical trial. J Infect, 71 (3), pp. 326-337. | Show Abstract | Read more

OBJECTIVES: Outer membrane vesicle (OMV) vaccines are used against outbreaks of capsular group B Neisseria meningitidis (MenB) caused by strains expressing particular PorA outer membrane proteins (OMPs). Ferric enterobactin receptor (FetA) is another variable OMP that induces type-specific bactericidal antibodies, and the combination of judiciously chosen PorA and FetA variants in vaccine formulations is a potential approach to broaden protection of such vaccines. METHODS: The OMV vaccine MenPF-1 was generated by genetically modifying N. meningitidis strain 44/76 to constitutively express FetA. Three doses of 25 μg or 50 μg of MenPF-1 were delivered intra-muscularly to 52 healthy adults. RESULTS: MenPF-1 was safe and well tolerated. Immunogenicity was measured by serum bactericidal assay (SBA) against wild-type and isogenic mutant strains. After 3 doses, the proportion of volunteers with SBA titres ≥1:4 (the putative protective titre) was 98% for the wild-type strain, and 77% for the strain 44/76 FetA(on)PorA(off) compared to 51% in the strain 44/76 FetA(off)PorA(off), demonstrating that vaccination with MenPF-1 simultaneously induced FetA and PorA bactericidal antibodies. CONCLUSION: This study provides a proof-of-concept for generating bactericidal antibodies against FetA after OMV vaccination in humans. Prevalence-based choice of PorA and FetA types can be used to formulate a vaccine for broad protection against MenB disease.

Weissmueller NT, Schiffter HA, Carlisle RC, Rollier CS, Pollard AJ. 2015. Needle-Free Dermal Delivery of a Diphtheria Toxin CRM197 Mutant on Potassium-Doped Hydroxyapatite Microparticles. Clin Vaccine Immunol, 22 (5), pp. 586-592. | Show Abstract | Read more

Injections with a hypodermic needle and syringe (HNS) are the current standard of care globally, but the use of needles is not without limitation. While a plethora of needle-free injection devices exist, vaccine reformulation is costly and presents a barrier to their widespread clinical application. To provide a simple, needle-free, and broad-spectrum protein antigen delivery platform, we developed novel potassium-doped hydroxyapatite (K-Hap) microparticles with improved protein loading capabilities that can provide sustained local antigen presentation and release. K-Hap showed increased protein adsorption over regular hydroxyapatite (P < 0.001), good structural retention of the model antigen (CRM197) with 1% decrease in α-helix content and no change in β-sheet content upon adsorption, and sustained release in vitro. Needle-free intradermal powder inoculation with K-Hap-CRM197 induced significantly higher IgG1 geometric mean titers (GMTs) than IgG2a GMTs in a BALB/c mouse model (P < 0.001) and induced IgG titer levels that were not different from the current clinical standard (P > 0.05), namely, alum-adsorbed CRM197 by intramuscular (i.m.) delivery. The presented results suggest that K-Hap microparticles may be used as a novel needle-free delivery vehicle for some protein antigens.

Marsay L, Dold C, Green CA, Rollier CS, Norheim G, Sadarangani M, Shanyinde M, Brehony C, Thompson AJ, Sanders H et al. 2015. A novel meningococcal outer membrane vesicle vaccine with constitutive expression of FetA: A phase I clinical trial Journal of Infection, 71 (3), pp. 326-337. | Show Abstract | Read more

© 2015 The Authors. Objectives: Outer membrane vesicle (OMV) vaccines are used against outbreaks of capsular group B Neisseria meningitidis (MenB) caused by strains expressing particular PorA outer membrane proteins (OMPs). Ferric enterobactin receptor (FetA) is another variable OMP that induces type-specific bactericidal antibodies, and the combination of judiciously chosen PorA and FetA variants in vaccine formulations is a potential approach to broaden protection of such vaccines. Methods: The OMV vaccine MenPF-1 was generated by genetically modifying N. meningitidis strain 44/76 to constitutively express FetA. Three doses of 25 μg or 50 μg of MenPF-1 were delivered intra-muscularly to 52 healthy adults. Results: MenPF-1 was safe and well tolerated. Immunogenicity was measured by serum bactericidal assay (SBA) against wild-type and isogenic mutant strains. After 3 doses, the proportion of volunteers with SBA titres ≥1:4 (the putative protective titre) was 98% for the wild-type strain, and 77% for the strain 44/76 FetA on PorA off compared to 51% in the strain 44/76 FetA off PorA off , demonstrating that vaccination with MenPF-1 simultaneously induced FetA and PorA bactericidal antibodies. Conclusion: This study provides a proof-of-concept for generating bactericidal antibodies against FetA after OMV vaccination in humans. Prevalence-based choice of PorA and FetA types can be used to formulate a vaccine for broad protection against MenB disease.

Rollier CS, Dold C, Marsay L, Sadarangani M, Pollard AJ. 2015. The capsular group B meningococcal vaccine, 4CMenB : clinical experience and potential efficacy. Expert Opin Biol Ther, 15 (1), pp. 131-142. | Show Abstract | Read more

INTRODUCTION: Capsular group B meningococcal disease is a leading cause of childhood meningitis and septicaemia. Up to 10% of sufferers die, and sequelae remain in > 30% of survivors. A vaccine, four component meningococcal group B ( 4CMenB ), designed with the aim to induce broad coverage against this highly variable bacterium, has been licensed in countries including in the European Union, Canada and Australia. AREAS COVERED: Immunogenicity and safety data, published in peer-reviewed literature between 2004 and 2014, are presented in the context of the recent recommendation for the use of the vaccine in infants in the UK. EXPERT OPINION: 4CMenB induces significant reactogenicity when administered with routine infant vaccines, in particular with respect to fever rates. Fevers can be somewhat reduced using paracetamol. The efficacy of the vaccine is unknown but has been extrapolated from effectiveness data obtained from use of one of its components in New Zealand, immunogenicity data from clinical trials and estimation of coverage from in vitro studies. These data suggest that the vaccine will prevent a proportion of invasive meningococcal disease cases in infants and young children. Implementation and well-planned post-marketing surveillance will address uncertainties over field effectiveness.

Mehta OH, Norheim G, Hoe JC, Rollier CS, Nagaputra JC, Makepeace K, Saleem M, Chan H, Ferguson DJP, Jones C et al. 2014. Adjuvant effects elicited by novel oligosaccharide variants of detoxified meningococcal lipopolysaccharides on Neisseria meningitidis recombinant PorA protein: a comparison in mice. PLoS One, 9 (12), pp. e115713. | Show Abstract | Read more

Neisseria meningitidis lipopolysaccharide (LPS) has adjuvant properties that can be exploited to assist vaccine immunogenicity. The modified penta-acylated LPS retains the adjuvant properties of hexa-acylated LPS but has a reduced toxicity profile. In this study we investigated whether two modified glycoform structures (LgtE and IcsB) of detoxified penta-acylated LPS exhibited differential adjuvant properties when formulated as native outer membrane vesicles (nOMVs) as compared to the previously described LgtB variant. Detoxified penta-acylated LPS was obtained by disruption of the lpxL1 gene (LpxL1 LPS), and three different glycoforms were obtained by disruption of the lgtB, lgtE or icsB genes respectively. Mice (mus musculus) were immunized with a recombinant PorA P1.7-2,4 (rPorA) protein co-administered with different nOMVs (containing a different PorA serosubtype P1.7,16), each of which expressed one of the three penta-acylated LPS glycoforms. All nOMVs induced IgG responses against the rPorA, but the nOMVs containing the penta-acylated LgtB-LpxL1 LPS glycoform induced significantly greater bactericidal activity compared to the other nOMVs or when the adjuvant was Alhydrogel. Compared to LgtE or IcsB LPS glycoforms, these data support the use of nOMVs containing detoxified, modified LgtB-LpxL1 LPS as a potential adjuvant for future meningococcal protein vaccines.

Nagaputra JC, Rollier CS, Sadarangani M, Hoe JC, Mehta OH, Norheim G, Saleem M, Chan H, Derrick JP, Feavers I et al. 2014. Neisseria meningitidis native outer membrane vesicles containing different lipopolysaccharide glycoforms as adjuvants for meningococcal and nonmeningococcal antigens. Clin Vaccine Immunol, 21 (2), pp. 234-242. | Show Abstract | Read more

We evaluated the adjuvant effect of a modified glycoform of lipopolysaccharide (LPS) (LgtB-LpxL1) compared to that of the nonmodified glycoform Lpxl1 serogroup B meningococcal H44/76 native outer membrane vesicles (nOMVs) on immune responses to vaccination with the recombinant meningococcal protein, rPorA, tetanus toxoid, or meningococcal serogroup C capsular polysaccharide. We used LgtB-LpxL1 LPS because the disruption of the lgtB gene, which results in the exposure of N-acetylglucosamine-galactose-glucose residues in the LPS outer core, has been shown to enhance the activation of human dendritic cells in vitro. The responses were compared to those of a monophosphoryl lipid A (MPL)-based adjuvant and to an aluminum hydroxide suspension. The nOMVs induced blood serum IgG responses against each of the three antigens comparable to those obtained with MPL or aluminum salt. However, nOMVs elicited (i) a lower IgG1/IgG2a ratio against rPorA and (ii) serum bactericidal antibody titers superior to those achieved with aluminum salt, reaching similar titers to those obtained with MPL. Similarly, bactericidal antibody titers induced by immunization with meningococcal serogroup C polysaccharide and nOMVs were similar to those obtained using MPL but were better than those with aluminum salt. Immunization with tetanus toxoid and nOMVs resulted in tetanus toxoid-specific IgG responses similar to those obtained when adjuvanted with aluminum salt. These results highlight the potential utility of meningococcal LpxL1 LPS-containing nOMVs as an adjuvant for recombinant meningococcal protein vaccines and suggest their possible use with a variety of other antigens.

van Diemen PM, Yamaguchi Y, Paterson GK, Rollier CS, Hill AVS, Wyllie DH. 2013. Irradiated wild-type and Spa mutant Staphylococcus aureus induce anti-S. aureus immune responses in mice which do not protect against subsequent intravenous challenge. Pathog Dis, 68 (1), pp. 20-26. | Show Abstract | Read more

Staphylococcus aureus remains an important human and animal pathogen. Its pathogenicity is determined in part by expression of the Spa-immune subversion protein, neutralising the activity of which provides partial protection in murine models, as does experimental infection with live S. aureus with Spa gene deletions followed by antibiotic-mediated cure in mice. Together, these data raise the question of whether Spa mutant S. aureus might represent a viable vaccine. Here, we find that gamma-irradiated S. aureus strains, both wild-type and null mutant of spa, are immunogenic in mice when administered intramuscularly, eliciting large amounts of anti-S. aureus antibodies, as judged by whole-cell immunoassay on fixed microorganisms. We used an intravenous challenge system to assess vaccine efficacy, the sensitivity of which was increased by studying renal bacterial concentrations in both kidneys. Despite this, protection from intravenous challenge was not observed (mean difference between vaccinated and unvaccinated mice 0.27 log(10) with 95% confidence interval -0.922 to 1.467). Surprisingly, antibody responses elicited against a panel of protective cell surface proteins were very low, indicating that most antibody induced is not protective. Additionally, these data suggest a limited role for irradiated wild-type or spa mutant S. aureus as vaccines.

Reyes-Sandoval A, Rollier CS, Milicic A, Bauza K, Cottingham MG, Tang C-K, Dicks MD, Wang D, Longley RJ, Wyllie DH, Hill AVS. 2012. Mixed vector immunization with recombinant adenovirus and MVA can improve vaccine efficacy while decreasing antivector immunity. Mol Ther, 20 (8), pp. 1633-1647. | Show Abstract | Read more

Substantial protection can be provided against the pre-erythrocytic stages of malaria by vaccination first with an adenoviral and then with an modified vaccinia virus Ankara (MVA) poxviral vector encoding the same ME.TRAP transgene. We investigated whether the two vaccine components adenovirus (Ad) and MVA could be coinjected as a mixture to enhance protection against malaria. A single-shot mixture at specific ratios of Ad and MVA (Ad+MVA) enhanced CD8(+) T cell-dependant protection of mice against challenge with Plasmodium berghei. Moreover, the degree of protection could be enhanced after homologous boosting with the same Ad+MVA mixture to levels comparable with classic heterologous Ad prime-MVA boost regimes. The mixture increased transgene-specific responses while decreasing the CD8(+) T cell antivector immunity compared to each vector used alone, particularly against the MVA backbone. Mixed vector immunization led to increased early circulating interferon-γ (IFN-γ) response levels and altered transcriptional microarray profiles. Furthermore, we found that sequential immunizations with the Ad+MVA mixture led to consistent boosting of the transgene-specific CD8(+) response for up to three mixture immunizations, whereas each vector used alone elicited progressively lower responses. Our findings offer the possibility of simplifying the deployment of viral vectors as a single mixture product rather than in heterologous prime-boost regimens.

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Reyes-Sandoval A, Rollier CS, Milicic A, Bauza K, Cottingham MG, Tang CK, Dicks MD, Wang D, Longley RJ, Wyllie DH, Hill AVS. 2012. Mixed vector immunization with recombinant adenovirus and MVA can improve vaccine efficacy while decreasing antivector immunity Molecular Therapy, 20 (8), pp. 1633-1647. | Show Abstract | Read more

Substantial protection can be provided against the pre-erythrocytic stages of malaria by vaccination first with an adenoviral and then with an modified vaccinia virus Ankara (MVA) poxviral vector encoding the same ME.TRAP transgene. We investigated whether the two vaccine components adenovirus (Ad) and MVA could be coinjected as a mixture to enhance protection against malaria. A single-shot mixture at specific ratios of Ad and MVA (AdMVA) enhanced CD8 ++ T cell-dependant protection of mice against challenge with Plasmodium berghei. Moreover, the degree of protection could be enhanced after homologous boosting with the same AdMVA mixture to levels comparable with classic heterologous Ad prime-MVA boost regimes. The mixture increased transgene-specific responses while decreasing the CD8 ++ T cell antivector immunity compared to each vector used alone, particularly against the MVA backbone. Mixed vector immunization led to increased early circulating interferon-γ (IFN-γ) response levels and altered transcriptional microarray profiles. Furthermore, we found that sequential immunizations with the AdMVA mixture led to consistent boosting of the transgene-specific CD8 ++ response for up to three mixture immunizations, whereas each vector used alone elicited progressively lower responses. Our findings offer the possibility of simplifying the deployment of viral vectors as a single mixture product rather than in heterologous prime-boost regimens. © The American Society of Gene & Cell Therapy.

Spencer AJ, Hill F, Honeycutt JD, Cottingham MG, Bregu M, Rollier CS, Furze J, Draper SJ, Søgaard KC, Gilbert SC et al. 2012. Fusion of the Mycobacterium tuberculosis antigen 85A to an oligomerization domain enhances its immunogenicity in both mice and non-human primates. PLoS One, 7 (3), pp. e33555. | Show Abstract | Read more

To prevent important infectious diseases such as tuberculosis, malaria and HIV, vaccines inducing greater T cell responses are required. In this study, we investigated whether fusion of the M. tuberculosis antigen 85A to recently described adjuvant IMX313, a hybrid avian C4bp oligomerization domain, could increase T cell responses in pre-clinical vaccine model species. In mice, the fused antigen 85A showed consistent increases in CD4(+) and CD8(+) T cell responses after DNA and MVA vaccination. In rhesus macaques, higher IFN-γ responses were observed in animals vaccinated with MVA-Ag85A IMX313 after both primary and secondary immunizations. In both animal models, fusion to IMX313 induced a quantitative enhancement in the response without altering its quality: multifunctional cytokines were uniformly increased and differentiation into effector and memory T cell subsets was augmented rather than skewed. An extensive in vivo characterization suggests that IMX313 improves the initiation of immune responses as an increase in antigen 85A specific cells was observed as early as day 3 after vaccination. This report demonstrates that antigen multimerization using IMX313 is a simple and effective cross-species method to improve vaccine immunogenicity with potentially broad applicability.

Reyes-Sandoval A, Wyllie DH, Bauza K, Milicic A, Forbes EK, Rollier CS, Hill AVS. 2011. CD8+ T effector memory cells protect against liver-stage malaria. J Immunol, 187 (3), pp. 1347-1357. | Show Abstract | Read more

Identification of correlates of protection for infectious diseases including malaria is a major challenge and has become one of the main obstacles in developing effective vaccines. We investigated protection against liver-stage malaria conferred by vaccination with adenoviral (Ad) and modified vaccinia Ankara (MVA) vectors expressing pre-erythrocytic malaria Ags. By classifying CD8(+) T cells into effector, effector memory (T(EM)), and central memory subsets using CD62L and CD127 markers, we found striking differences in T cell memory generation. Although MVA induced accelerated central memory T cell generation, which could be efficiently boosted by subsequent Ad administration, it failed to protect against malaria. In contrast, Ad vectors, which permit persistent Ag delivery, elicit a prolonged effector T cell and T(EM) response that requires long intervals for an efficient boost. A preferential T(EM) phenotype was maintained in liver, blood, and spleen after Ad/MVA prime-boost regimens, and animals were protected against malaria sporozoite challenge. Blood CD8(+) T(EM) cells correlated with protection against malaria liver-stage infection, assessed by estimation of number of parasites emerging from the liver into the blood. The protective ability of Ag-specific T(EM) cells was confirmed by transfer experiments into naive recipient mice. Thus, we identify persistent CD8 T(EM) populations as essential for vaccine-induced pre-erythrocytic protection against malaria, a finding that has important implications for vaccine design.

Rollier CS, Reyes-Sandoval A, Cottingham MG, Ewer K, Hill AVS. 2011. Viral vectors as vaccine platforms: deployment in sight. Curr Opin Immunol, 23 (3), pp. 377-382. | Show Abstract | Read more

A little more than a decade after the explosion of research into recombinant live-attenuated or replication-deficient viruses as vaccine platforms, many viral vector-based vaccines have been licensed for animals. Progress has been slower for humans but 2011 will see the licensure of the first viral-vectored vaccine for humans, against Japanese Encephalitis. In addition a vaccine with a viral-vectored component showed efficacy against HIV infection in humans. Viral-based vaccines have an excellent safety profile but must deal with the potential problem of pre-existing anti-vector immunity. Recent successes reflect diverse improvements such as development of new adenovirus serotypes and better prime-boost approaches, suggesting that many viral vectors are approaching their final years as vaccine 'candidates' rather than vaccines.

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Reyes-Sandoval A, Wyllie DH, Bauza K, Milicic A, Forbes EK, Rollier CS, Hill AVS. 2011. CD8<sup>+</sup>T effector memory cells protect against liver-stage malaria Journal of Immunology, 187 (3), pp. 1347-1357. | Show Abstract | Read more

Identification of correlates of protection for infectious diseases including malaria is a major challenge and has become one of the main obstacles in developing effective vaccines.We investigated protection against liver-stage malaria conferred by vaccination with adenoviral (Ad) and modified vaccinia Ankara (MVA) vectors expressing pre-erythrocytic malaria Ags. By classifying CD8+ T cells into effector, effector memory (TEM), and central memory subsets using CD62L and CD127 markers, we found striking differences in T cell memory generation. Although MVA induced accelerated central memory T cell generation, which could be efficiently boosted by subsequent Ad administration, it failed to protect against malaria. In contrast, Ad vectors, which permit persistent Ag delivery, elicit a prolonged effector T cell and TEM response that requires long intervals for an efficient boost. A preferential TEM phenotype was maintained in liver, blood, and spleen after Ad/MVA prime-boost regimens, and animals were protected against malaria sporozoite challenge. Blood CD8+ TEM cells correlated with protection against malaria liver-stage infection, assessed by estimation of number of parasites emerging from the liver into the blood. The protective ability of Agspecific TEM cells was confirmed by transfer experiments into naive recipient mice. Thus, we identify persistent CD8 TEMpopulations as essential for vaccine-induced pre-erythrocytic protection against malaria, a finding that has important implications for vaccine design. Copyright © 2011 by The American Association of Immunologists, Inc.

Verstrepen BE, Depla E, Rollier CS, Mares G, Drexhage JAR, Priem S, Verschoor EJ, Koopman G, Granier C, Dreux M et al. 2011. Clearance of genotype 1b hepatitis C virus in chimpanzees in the presence of vaccine-induced E1-neutralizing antibodies. J Infect Dis, 204 (6), pp. 837-844. | Show Abstract | Read more

Accumulating evidence indicates that neutralizing antibodies play an important role in protection from chronic hepatitis C virus (HCV) infection. Efforts to elicit such responses by immunization with intact heterodimeric E1E2 envelope proteins have met with limited success. To determine whether antigenic sites, which are not exposed by the combined E1E2 heterodimer structure, are capable of eliciting neutralizing antibody responses, we expressed and purified each as separate recombinant proteins E1 and E2, from which the immunodominant hypervariable region (HVR-1) was deleted. Immunization of chimpanzees with either E1 or E2 alone induced antigen-specific T-helper cytokines of similar magnitude. Unexpectedly, the capacity to neutralize HCV was observed in E1 but not in animals immunized with E2 devoid of HVR-1. Furthermore, in vivo only E1-vaccinated animals exposed to the heterologous HCV-1b inoculum cleared HCV infection.

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Rollier CS, Reyes-Sandoval A, Cottingham MG, Ewer K, Hill AVS. 2011. Viral vectors as vaccine platforms: Deployment in sight Current Opinion in Immunology, 23 (3), pp. 377-382. | Show Abstract | Read more

A little more than a decade after the explosion of research into recombinant live-attenuated or replication-deficient viruses as vaccine platforms, many viral vector-based vaccines have been licensed for animals. Progress has been slower for humans but 2011 will see the licensure of the first viral-vectored vaccine for humans, against Japanese Encephalitis. In addition a vaccine with a viral-vectored component showed efficacy against HIV infection in humans. Viral-based vaccines have an excellent safety profile but must deal with the potential problem of pre-existing anti-vector immunity. Recent successes reflect diverse improvements such as development of new adenovirus serotypes and better prime-boost approaches, suggesting that many viral vectors are approaching their final years as vaccine 'candidates' rather than vaccines. © 2011 Elsevier Ltd.

Alcock R, Cottingham MG, Rollier CS, Furze J, De Costa SD, Hanlon M, Spencer AJ, Honeycutt JD, Wyllie DH, Gilbert SC et al. 2010. Long-term thermostabilization of live poxviral and adenoviral vaccine vectors at supraphysiological temperatures in carbohydrate glass. Sci Transl Med, 2 (19), pp. 19ra12. | Show Abstract | Read more

Live recombinant viral vectors based on adenoviruses and poxviruses are among the most promising platforms for development of new vaccines against diseases such as malaria, tuberculosis, and HIV-AIDS. Vaccines based on live viruses must remain infectious to be effective, so therefore need continuous refrigeration to maintain stability and viability, a requirement that can be costly and difficult, especially in developing countries. The sugars sucrose and trehalose are commonly used as stabilizing agents and cryoprotectants for biological products. Here, we have exploited the ability of these sugars to vitrify on desiccation to develop a thermostabilization technique for live viral vaccine vectors. By slowly drying vaccines suspended in solutions of these disaccharide stabilizers onto a filter-like support membrane at ambient temperature, an ultrathin glass is deposited on the fibers of the inert matrix. Immobilization of two recombinant vaccine vectors-E1/E3-deleted human adenovirus type 5 and modified vaccinia virus Ankara-in this glass on the membranes enabled complete recovery of viral titer and immunogenicity after storage at up to 45 degrees C for 6 months and even longer with minimal losses. Furthermore, the membrane carrying the stabilized vaccine can be incorporated into a holder attached to a syringe for almost simultaneous reconstitution and injection at point of use. The technology may potentially be developed for the deployment of viral vector-based biopharmaceuticals in resource-poor settings.

Capone S, Reyes-Sandoval A, Naddeo M, Siani L, Ammendola V, Rollier CS, Nicosia A, Colloca S, Cortese R, Folgori A, Hill AVS. 2010. Immune responses against a liver-stage malaria antigen induced by simian adenoviral vector AdCh63 and MVA prime-boost immunisation in non-human primates. Vaccine, 29 (2), pp. 256-265. | Show Abstract | Read more

Malaria is a major health problem as nearly half of the human population is exposed to this parasite causing around 600 million clinical cases annually. Prime-boost regimes using simian adenoviral vectors and MVA expressing the clinically relevant Plasmodium falciparum ME.TRAP antigen have shown outstanding protective efficacy in mouse models. We now extend those observations to macaque monkeys. Immunisation with AdCh63 elicited a median response of 869 IFN-γ SFC/million PBMCs to ME.TRAP and responses were boosted by MVA to reach 5256 SFC/million PBMCs, increasing at the same time the breadth of the T cell responses to cover the complete ME.TRAP antigen. Intramuscular vaccination was more immunogenic than the intradermal route, and MVA could be used repeatedly for up to 3 times to boost adenovirus-primed responses. An interval of 16 weeks between repeated MVA injections was optimal to enhance cytokine production by T cells and improve the CD8 multifunctional responses. Antibodies to TRAP were exceptionally high and maintained for a long period of time after the prime-boost regime. These results in non-human primates highlight the potential of this vaccination regime and encourage its future use in clinical trials.

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Capone S, Reyes-Sandoval A, Naddeo M, Siani L, Ammendola V, Rollier CS, Nicosia A, Colloca S, Cortese R, Folgori A, Hill AVS. 2010. Immune responses against a liver-stage malaria antigen induced by simian adenoviral vector AdCh63 and MVA prime-boost immunisation in non-human primates Vaccine, 29 (2), pp. 256-265. | Show Abstract | Read more

Malaria is a major health problem as nearly half of the human population is exposed to this parasite causing around 600 million clinical cases annually. Prime-boost regimes using simian adenoviral vectors and MVA expressing the clinically relevant Plasmodium falciparum ME.TRAP antigen have shown outstanding protectiv e efficacy in mouse models. We now extend those observations to macaque monkeys. Immunisation with AdCh63 elicited a median response of 869 IFN-γ SFC/million PBMCs to ME.TRAP and responses were boosted by MVA to reach 5256 SFC/million PBMCs, increasing at the same time the breadth of the T cell responses to cover the complete ME.TRAP antigen. Intramuscular vaccination was more immunogenic than the intradermal route, and MVA could be used repeatedly for up to 3 times to boost adenovirus-primed responses. An interval of 16 weeks between repeated MVA injections was optimal to enhance cytokine production by T cells and improve the CD8 multifunctional responses. Antibodies to TRAP were exceptionally high and maintained for a long period of time after the prime-boost regime. These results in non-human primates highlight the potential of this vaccination regime and encourage its future use in clinical trials. © 2010 Elsevier Ltd.

Larsen KC, Spencer AJ, Goodman AL, Gilchrist A, Furze J, Rollier CS, Kiss-Toth E, Gilbert SC, Bregu M, Soilleux EJ et al. 2009. Expression of tak1 and tram induces synergistic pro-inflammatory signalling and adjuvants DNA vaccines. Vaccine, 27 (41), pp. 5589-5598. | Show Abstract | Read more

Improving vaccine immunogenicity remains a major challenge in the fight against developing country diseases like malaria and AIDS. We describe a novel strategy to identify new DNA vaccine adjuvants. We have screened components of the Toll-like receptor signalling pathways for their ability to activate pro-inflammatory target genes in transient transfection assays and assessed in vivo adjuvant activity by expressing the activators from the DNA backbone of vaccines. We find that a robust increase in the immune response necessitates co-expression of two activators. Accordingly, the combination of tak1 and tram elicits synergistic reporter activation in transient transfection assays. In a mouse model this combination, but not the individual molecules, induced approximately twofold increases in CD8+ T-cell immune responses. These results indicate that optimal immunogenicity may require activation of distinct innate immune signalling pathways. Thus this strategy offers a novel route to the discovery of a new generation of adjuvants.

Verstrepen BE, Bins AD, Rollier CS, Mooij P, Koopman G, Sheppard NC, Sattentau Q, Wagner R, Wolf H, Schumacher TNM et al. 2008. Improved HIV-1 specific T-cell responses by short-interval DNA tattooing as compared to intramuscular immunization in non-human primates. Vaccine, 26 (26), pp. 3346-3351. | Show Abstract | Read more

The new intradermal DNA delivery technique, termed DNA tattooing might overcome the discrepancy between the encouraging immunogenicity results obtained with DNA vaccines in murine studies and the poor results obtained in non-human primates and humans, the so called "simian barrier". Here, we demonstrate a 10- to 100-fold increase in the magnitude of vaccine specific T-cell responses in peripheral blood from DNA tattooed rhesus macaques, as compared to T-cell responses in animals immunized via intramuscular (IM) route. A marked increase in the magnitude of the antigen specific T-cell responses as well as an increase in the number of animals responding to the immunogens was observed. These findings in non-human primates suggest that similar results may be observed in humans. Clinical trials are planned to validate tattooing as an optimal method of DNA vaccine delivery in humans.

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Verstrepen BE, Bins AD, Rollier CS, Mooij P, Koopman G, Sheppard NC, Sattentau Q, Wagner R, Wolf H, Schumacher TNM et al. 2008. Improved HIV-1 specific T-cell responses by short-interval DNA tattooing as compared to intramuscular immunization in non-human primates Vaccine, 26 (26), pp. 3346-3351. | Show Abstract | Read more

The new intradermal DNA delivery technique, termed DNA tattooing might overcome the discrepancy between the encouraging immunogenicity results obtained with DNA vaccines in murine studies and the poor results obtained in non-human primates and humans, the so called "simian barrier". Here, we demonstrate a 10- to 100-fold increase in the magnitude of vaccine specific T-cell responses in peripheral blood from DNA tattooed rhesus macaques, as compared to T-cell responses in animals immunized via intramuscular (IM) route. A marked increase in the magnitude of the antigen specific T-cell responses as well as an increase in the number of animals responding to the immunogens was observed. These findings in non-human primates suggest that similar results may be observed in humans. Clinical trials are planned to validate tattooing as an optimal method of DNA vaccine delivery in humans. © 2008 Elsevier Ltd. All rights reserved.

Larrea E, Riezu-Boj JI, Gil-Guerrero L, Casares N, Aldabe R, Sarobe P, Civeira MP, Heeney JL, Rollier C, Verstrepen B et al. 2007. Upregulation of indoleamine 2,3-dioxygenase in hepatitis C virus infection. J Virol, 81 (7), pp. 3662-3666. | Show Abstract | Read more

Indoleamine 2,3-dioxygenase (IDO) is induced by proinflammatory cytokines and by CTLA-4-expressing T cells and constitutes an important mediator of peripheral immune tolerance. In chronic hepatitis C, we found upregulation of IDO expression in the liver and an increased serum kynurenine/tryptophan ratio (a reflection of IDO activity). Huh7 cells supporting hepatitis C virus (HCV) replication expressed higher levels of IDO mRNA than noninfected cells when stimulated with gamma interferon or when cocultured with activated T cells. In infected chimpanzees, hepatic IDO expression decreased in animals that cured the infection, while it remained high in those that progressed to chronicity. For both patients and chimpanzees, hepatic expression of IDO and CTLA-4 correlated directly. Induction of IDO may dampen T-cell reactivity to viral antigens in chronic HCV infection.

Rollier CS, Paranhos-Baccala G, Verschoor EJ, Verstrepen BE, Drexhage JAR, Fagrouch Z, Berland J-L, Komurian-Pradel F, Duverger B, Himoudi N et al. 2007. Vaccine-induced early control of hepatitis C virus infection in chimpanzees fails to impact on hepatic PD-1 and chronicity. Hepatology, 45 (3), pp. 602-613. | Show Abstract | Read more

UNLABELLED: Broad T cell and B cell responses to multiple HCV antigens are observed early in individuals who control or clear HCV infection. The prevailing hypothesis has been that similar immune responses induced by prophylactic immunization would reduce acute virus replication and protect exposed individuals from chronic infection. Here, we demonstrate that immunization of naïve chimpanzees with a multicomponent HCV vaccine induced robust HCV-specific immune responses, and that all vaccinees exposed to heterologous chimpanzee-adapted HCV 1b J4 significantly reduced viral RNA in serum by 84%, and in liver by 99% as compared to controls (P=0.024 and 0.028, respectively). However, despite control of HCV in plasma and liver in the acute period, in the chronic phase, 3 of 4 vaccinated animals developed persistent infection. Analysis of expression levels of proinflammatory cytokines in serial hepatic biopsies failed to reveal an association with vaccine outcome. However, expression of IDO, CTLA-4 [corrected] and PD-1 levels in liver correlated with clearance or chronicity. CONCLUSION: Despite early control of virus load, a virus-associated tolerogenic-like state can develop in certain individuals independent of vaccination history.

Rollier C, Verschoor EJ, Paranhos-Baccala G, Drexhage JAR, Verstrepen BE, Berland J-L, Himoudi N, Barnfield C, Liljestrom P, Lasarte JJ et al. 2005. Modulation of vaccine-induced immune responses to hepatitis C virus in rhesus macaques by altering priming before adenovirus boosting. J Infect Dis, 192 (5), pp. 920-929. | Show Abstract | Read more

BACKGROUND: Preventive and therapeutic vaccine strategies aimed at controlling hepatitis C virus (HCV) infection should mimic the immune responses observed in patients who control or clear HCV, specifically T helper (Th) type 1 and CD8+ cell responses to multiple antigens, including nonstructural protein (NS) 3. Given the experience with human immunodeficiency virus, the best candidates for this are based on DNA prime, pox, or adenovirus boost regimens. METHODS: In rhesus macaques, we compared NS3-expressing DNA prime and adenovirus boost strategy with 2 alternative priming approaches aimed at modifying Th1 and CD8+ responses: DNA adjuvanted with interleukin (IL)-2- and -12-encoding plasmids or Semliki Forest virus (SFV). RESULTS: All prime-boost regimens elicited NS3-specific B and T cell responses in rhesus macaques, including CD8+ responses. SFV priming induced higher lymphoproliferation and longer Th1 memory responses. The use of IL-2- and IL-12-expressing vectors resulted in reduced Th2 and antibody responses, which led to increased Th1 skewing but not to an increase in the magnitude of the IFN- gamma and CD8+ responses. CONCLUSIONS: All strategies induced Th1 cellular responses to HCV NS3, with fine modulations depending on the different priming approaches. When they are developed for more HCV antigens, these strategies could be beneficial in therapeutic vaccine approaches.

Thermet A, Robaczewska M, Rollier C, Hantz O, Trepo C, Deleage G, Cova L. 2004. Identification of antigenic regions of duck hepatitis B virus core protein with antibodies elicited by DNA immunization and chronic infection. J Virol, 78 (4), pp. 1945-1953. | Show Abstract | Read more

The induction of humoral response in ducks by DNA-based immunization against duck hepatitis B virus (DHBV) core protein (DHBc) was investigated. In addition, the amino acid specificity of the induced response was compared by using peptide scanning to that elicited either by protein immunization or during chronic DHBV infection. Immunization of ducks with a plasmid expressing DHBc protein led to the induction of a long-lasting antibody response able to specifically recognize viral protein in chronically infected duck livers. Peptide scanning analysis of anti-DHBc response induced during chronic DHBV infection allowed us to identify six major antigenic regions (AR1 to AR6). The reactivity spectrum of duck sera elicited by protein immunization appeared narrower and was restricted to only four of these antigenic regions in spite of higher anti-DHBc antibody titers. Interestingly, anti-DHBc antibodies induced by DNA-based immunization recognized five of six antigenic regions, and the epitope pattern was broader and more closely related to that observed in chronic viral infections. To gain more insight into the location of antigenic regions, we built a three-dimensional (3-D) model of DHBc protein based on human and duck core sequence alignment data and the HBc 3-D crystal structure. The results suggest that two identified antigenic regions (AR2, amino acids [aa] (64)T-P(84), and AR5, aa (183)A-R(210)) are located at positions on the protein surface equivalent to those of the two HBc major epitopes. Moreover, we identified another antigenic region (AR3, aa (99)I-I(112)) that was recognized by all sera from chronically infected, DNA- or protein-immunized ducks within the large 45-aa insertion in DHBc protein, suggesting that this region, which lacks HBc, is externally exposed.

Rollier C, Depla E, Drexhage JAR, Verschoor EJ, Verstrepen BE, Fatmi A, Brinster C, Fournillier A, Whelan JA, Whelan M et al. 2004. Control of heterologous hepatitis C virus infection in chimpanzees is associated with the quality of vaccine-induced peripheral T-helper immune response. J Virol, 78 (1), pp. 187-196. | Show Abstract | Read more

Prophylactic hepatitis C virus (HCV) vaccine trials with human volunteers are pending. There is an important need for immunological end points which correlate with vaccine efficacy and which do not involve invasive procedures, such as liver biopsies. By using a multicomponent DNA priming-protein boosting vaccine strategy, naïve chimpanzees were immunized against HCV structural proteins (core, E1, and E2) as well as a nonstructural (NS3) protein. Following immunization, exposure to the heterologous HCV 1b J4 subtype resulted in a peak of plasma viremia which was lower in both immunized animals. Compared to the naïve infection control and nine additional historical controls which became chronic, vaccinee 2 (Vac2) rapidly resolved the infection, while the other (Vac1) clearly controlled HCV infection. Immunization induced antibodies, peptide-specific gamma interferon (IFN-gamma), protein-specific lymphoproliferative responses, IFN-gamma, interleukin-2 (IL-2), and IL-4 T-helper responses in both vaccinees. However, the specificities were markedly different: Vac2 developed responses which were lower in magnitude than those of Vac1 but which were biased towards Th1-type cytokine responses for E1 and NS3. This proof-of-principle study in chimpanzees revealed that immunization with a combination of nonstructural and structural antigens elicited T-cell responses associated with an alteration of the course of infection. Our findings provide data to support the concept that the quality of the response to conserved epitopes and the specific nature of the peripheral T-helper immune response are likely pivotal factors influencing the control and clearance of HCV infection.

Rollier C, Drexhage JAR, Verstrepen BE, Verschoor EJ, Bontrop RE, Koopman G, Heeney JL. 2003. Chronic hepatitis C virus infection established and maintained in chimpanzees independent of dendritic cell impairment. Hepatology, 38 (4), pp. 851-858. | Show Abstract | Read more

Chronic hepatitis C virus (HCV) infection in humans is associated with an impairment of dendritic cells (DC). It has been hypothesized that impairment of DC function may be a central mechanism facilitating the establishment of a chronic carrier state. However, the majority of patients studied with DC impairment to date have been identified and, thus, inadvertently selected because of clinical manifestations leading to their diagnosis, which may have been many years following actual infection. We set out to determine whether impaired DC function occurred in the earlier asymptomatic phase of infection and turned to a well-defined cohort of HCV-infected chimpanzees in which the specific date of infection and the nature of the inoculum were well characterized. Results revealed that, in contrast to the observations in human subjects with advanced clinical hepatitis, there was neither impairment of the allostimulatory capacity of monocyte-derived DC from HCV chronic carriers nor impairment of the maturation process. Decreased allostimulatory capacity was only detected in 2 animals and, interestingly, in those that possessed the highest viral loads. Nevertheless, HCV sequences were undetectable in any of the DC derived from HCV-infected chimpanzees. In conclusion, these findings suggest that the mechanisms of establishing persistent HCV infection are separate and independent from those responsible for impaired DC function. Indeed, the maturation and allostimulatory impairment, as described in patient studies, are not necessary prerequisites but rather possible consequences of persistent and active HCV infection associated with disease progression.

Thermet A, Rollier C, Zoulim F, Trepo C, Cova L. 2003. Progress in DNA vaccine for prophylaxis and therapy of hepatitis B. Vaccine, 21 (7-8), pp. 659-662. | Show Abstract | Read more

Increasing lines of evidence suggest that DNA vaccine is of interest to fight chronic hepatitis B virus (HBV) infection. We used the Pekin duck infected by duck HBV (DHBV), closely related to the human virus, which is an attractive model allowing study of protective and therapeutic effectiveness of DNA vaccines against hepatitis B. Immunisation with a plasmid encoding the DHBV large (L) envelope protein induced a strong, specific, highly neutralising and long-lasting anti-preS humoral response in uninfected ducks. Importantly, maternal antibodies elicited by such DNA immunisation were vertically transmitted and protected progeny against viral challenge. Therapeutic immunisation of chronic DHBV-carrier ducks with this plasmid DNA led to the dramatic and sustained decrease in viral replication and even to clearance of intrahepatic viral covalently close circular DNA (cccDNA) pool in some animals. Our recent combination therapy data showed even a more pronounced antiviral effect of DNA vaccine to DHBV envelope protein when associated with antiviral drug (lamivudine) treatment. Therefore, DNA-based vaccine appears as a promising new approach for prophylaxis and therapy of hepatitis B.

Heeney J, Drexhage J, Verstrepen B, Rollier C, Mares G, Jacobs D, Depla E, Maertens G, Brinster C, Fournillier A et al. 2001. Vaccination of chimpanzees with a multicomponent DNA-protein prime-boost regimen provides an enhanced recovery following heterologous HCV challenge. HEPATOLOGY, 34 (4), pp. 450A-450A.

Thermet A, Robaczewska M, Charollois C, Rollier C, Kay A, Cagnon N, Geourgeon C, Deleage G, Trepo C, Cova L. 2001. DNA immunization for identification and localization of antigenic regions on duck hepatitis B virus core protein. HEPATOLOGY, 34 (4), pp. 322A-322A.

Rollier C, Charollois C, Jamard C, Trepo C, Cova L. 2000. Early life humoral response of ducks to DNA immunization against hepadnavirus large envelope protein. Vaccine, 18 (27), pp. 3091-3096. | Show Abstract | Read more

DNA vaccination may represent an interesting strategy for early life immunization. However, in some cases, this approach has been shown to induce a tolerance rather than immunity. We have compared the efficiency of neonatal DNA or protein immunization against hepadnavirus envelope protein using the duck hepatitis B virus (DHBV) model. Three-day-old ducklings were immunized with either a plasmid encoding the DHBV pre-S/S large envelope protein (L), or a recombinant preS protein, followed by sequential DNA or protein boosts at weeks 4 and 15. Our results showed that genetic immunization of duck neonates induced specific humoral response to DHBV L protein. Interestingly, an enhanced antibody response was elicited when animals received DNA priming-DNA boosting as compared to DNA priming-protein boosting.

Rollier C, Charollois C, Jamard C, Trepo C, Cova L. 2000. Maternally transferred antibodies from DNA-immunized avians protect offspring against hepadnavirus infection. J Virol, 74 (10), pp. 4908-4911. | Show Abstract | Read more

The outcome and protective efficacy of maternal antibodies elicited by DNA immunization to the large (L) hepadnavirus envelope protein were studied using the duck hepatitis B virus (DHBV) model. Following genetic immunization of breeding ducks with a DHBV L protein gene-bearing plasmid, specific and highly neutralizing antibodies were transferred from the sera of immunized ducks, via the egg yolk, to the progeny of vaccinees. Interestingly, large amounts (60 to 100 mg/egg) of high-titer and L protein-specific yolk immunoglobulins (immunoglobulin Y) accumulated in the egg yolk. These results suggest that eggs from genetically immunized avians may represent a potent source of DNA-designed antibodies specific to viral antigen. Importantly, these antibodies are vertically transmitted and protect offspring against high-titer DHBV challenge.

Rollier C, Deleage C, Trepo C, Cova L. 1999. DNA immunization for production in egg factory of protective antibodies against hepadnavirus infection HEPATOLOGY, 30 (4), pp. 300A-300A.

Sunyach C, Rollier C, Robaczewska M, Borel C, Barraud L, Kay A, Trépo C, Will H, Cova L. 1999. Residues critical for duck hepatitis B virus neutralization are involved in host cell interaction. J Virol, 73 (4), pp. 2569-2575. | Show Abstract

To date, no detailed analysis of the neutralization properties of duck hepatitis B virus (DHBV) has been reported, and it is not clear whether any of the known neutralization epitopes correspond to the viral receptor binding site or to sequences involved in the cell entry pathway. We demonstrate here that antibodies directed against two overlapping peptides (amino acids 83 to 97 and 93 to 107), covering the sequences of most DHBV pre-S neutralizing epitopes, both inhibit virus binding to primary duck hepatocytes and neutralize virus infectivity. An extensive mutagenesis of the motif 88WTP90, which is the shortest sequence of the epitope recognized by the virus-neutralizing monoclonal antibody (MAb) 900 was performed in order to define the amino acids involved in these interactions. Single point mutations within this epitope affected neither virus replication nor infectivity but abolished virus neutralization by MAb 900 completely. Interestingly, mutants with two and three consecutive residue replacements (SIP and SIH) within this epitope retained replication competence but were no longer infectious. The loss of infectivity of SIH and SIP mutant particles was associated with significantly reduced binding to primary duck hepatocytes and could be rescued by trans complementation with wild-type pre-S protein. Taken together, these results indicate that each amino acid of the DHBV pre-S sequence 88WTP90 is critical for recognition by the neutralizing MAb 900 and that replacement of the first two or all three residues strongly reduces virus interaction with hepatocytes and abrogates infectivity. These data imply that the motif 88WTP90 contains key residues which are critical for interaction with both the neutralizing MAb and the host cell.

Rollier C, Sunyach C, Barraud L, Madani N, Jamard C, Trepo C, Cova L. 1999. Protective and therapeutic effect of DNA-based immunization against hepadnavirus large envelope protein. Gastroenterology, 116 (3), pp. 658-665. | Show Abstract | Read more

BACKGROUND & AIMS: Studies in the murine model suggest that injection of DNA encoding hepatitis B virus structural proteins is promising for the induction of a specific immune response. We used the duck hepatitis B virus (DHBV) model to study the protective and therapeutic effects of naked DNA immunization against hepadnaviral large envelope protein. METHODS: A pCI-preS/S plasmid expressing the DHBV large protein was used for intramuscular immunization of ducks. The humoral response was tested by enzyme-linked immunosorbent assay, immunoblotting, neutralization, and in vivo protection tests. For DNA therapy, DHBV-carrier ducks received four injections of this plasmid. Viremia was monitored for 10 months; thereafter, liver biopsies were performed. RESULTS: Immunization with pCI-preS/S plasmid induced a specific, long-lasting, neutralizing, and highly protective anti-preS humoral response in uninfected animals. After pCI-preS/S treatment, a significant and sustained decrease in serum and liver DHBV DNA was observed for carrier ducks compared with the controls. CONCLUSIONS: DNA immunization against DHBV large protein results in a potent and protective anti-preS response in the duck model. The results of long-term follow-up of DNA-treated chronically infected ducks are promising and show the usefulness of this model for the study of genetic immunization in chronic hepatitis B therapy.

Rollier C, Sunyach C, Barraud L, Madani N, Jamard C, Trepo C, Cova L. 1998. Protective and therapeutic effect of DNA-based immunization against hepadnavirus large envelope protein. HEPATOLOGY, 28 (4), pp. 319A-319A.

Rollier CS, Dold C, Marsay L, Sadarangani M, Pollard AJ. 2015. The capsular group B meningococcal vaccine, 4CMenB : clinical experience and potential efficacy. Expert Opin Biol Ther, 15 (1), pp. 131-142. | Show Abstract | Read more

INTRODUCTION: Capsular group B meningococcal disease is a leading cause of childhood meningitis and septicaemia. Up to 10% of sufferers die, and sequelae remain in > 30% of survivors. A vaccine, four component meningococcal group B ( 4CMenB ), designed with the aim to induce broad coverage against this highly variable bacterium, has been licensed in countries including in the European Union, Canada and Australia. AREAS COVERED: Immunogenicity and safety data, published in peer-reviewed literature between 2004 and 2014, are presented in the context of the recent recommendation for the use of the vaccine in infants in the UK. EXPERT OPINION: 4CMenB induces significant reactogenicity when administered with routine infant vaccines, in particular with respect to fever rates. Fevers can be somewhat reduced using paracetamol. The efficacy of the vaccine is unknown but has been extrapolated from effectiveness data obtained from use of one of its components in New Zealand, immunogenicity data from clinical trials and estimation of coverage from in vitro studies. These data suggest that the vaccine will prevent a proportion of invasive meningococcal disease cases in infants and young children. Implementation and well-planned post-marketing surveillance will address uncertainties over field effectiveness.

Nagaputra JC, Rollier CS, Sadarangani M, Hoe JC, Mehta OH, Norheim G, Saleem M, Chan H, Derrick JP, Feavers I et al. 2014. Neisseria meningitidis native outer membrane vesicles containing different lipopolysaccharide glycoforms as adjuvants for meningococcal and nonmeningococcal antigens. Clin Vaccine Immunol, 21 (2), pp. 234-242. | Show Abstract | Read more

We evaluated the adjuvant effect of a modified glycoform of lipopolysaccharide (LPS) (LgtB-LpxL1) compared to that of the nonmodified glycoform Lpxl1 serogroup B meningococcal H44/76 native outer membrane vesicles (nOMVs) on immune responses to vaccination with the recombinant meningococcal protein, rPorA, tetanus toxoid, or meningococcal serogroup C capsular polysaccharide. We used LgtB-LpxL1 LPS because the disruption of the lgtB gene, which results in the exposure of N-acetylglucosamine-galactose-glucose residues in the LPS outer core, has been shown to enhance the activation of human dendritic cells in vitro. The responses were compared to those of a monophosphoryl lipid A (MPL)-based adjuvant and to an aluminum hydroxide suspension. The nOMVs induced blood serum IgG responses against each of the three antigens comparable to those obtained with MPL or aluminum salt. However, nOMVs elicited (i) a lower IgG1/IgG2a ratio against rPorA and (ii) serum bactericidal antibody titers superior to those achieved with aluminum salt, reaching similar titers to those obtained with MPL. Similarly, bactericidal antibody titers induced by immunization with meningococcal serogroup C polysaccharide and nOMVs were similar to those obtained using MPL but were better than those with aluminum salt. Immunization with tetanus toxoid and nOMVs resulted in tetanus toxoid-specific IgG responses similar to those obtained when adjuvanted with aluminum salt. These results highlight the potential utility of meningococcal LpxL1 LPS-containing nOMVs as an adjuvant for recombinant meningococcal protein vaccines and suggest their possible use with a variety of other antigens.

Rollier CS, Reyes-Sandoval A, Cottingham MG, Ewer K, Hill AVS. 2011. Viral vectors as vaccine platforms: deployment in sight. Curr Opin Immunol, 23 (3), pp. 377-382. | Show Abstract | Read more

A little more than a decade after the explosion of research into recombinant live-attenuated or replication-deficient viruses as vaccine platforms, many viral vector-based vaccines have been licensed for animals. Progress has been slower for humans but 2011 will see the licensure of the first viral-vectored vaccine for humans, against Japanese Encephalitis. In addition a vaccine with a viral-vectored component showed efficacy against HIV infection in humans. Viral-based vaccines have an excellent safety profile but must deal with the potential problem of pre-existing anti-vector immunity. Recent successes reflect diverse improvements such as development of new adenovirus serotypes and better prime-boost approaches, suggesting that many viral vectors are approaching their final years as vaccine 'candidates' rather than vaccines.

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Rollier CS, Reyes-Sandoval A, Cottingham MG, Ewer K, Hill AVS. 2011. Viral vectors as vaccine platforms: Deployment in sight Current Opinion in Immunology, 23 (3), pp. 377-382. | Show Abstract | Read more

A little more than a decade after the explosion of research into recombinant live-attenuated or replication-deficient viruses as vaccine platforms, many viral vector-based vaccines have been licensed for animals. Progress has been slower for humans but 2011 will see the licensure of the first viral-vectored vaccine for humans, against Japanese Encephalitis. In addition a vaccine with a viral-vectored component showed efficacy against HIV infection in humans. Viral-based vaccines have an excellent safety profile but must deal with the potential problem of pre-existing anti-vector immunity. Recent successes reflect diverse improvements such as development of new adenovirus serotypes and better prime-boost approaches, suggesting that many viral vectors are approaching their final years as vaccine 'candidates' rather than vaccines. © 2011 Elsevier Ltd.

Alcock R, Cottingham MG, Rollier CS, Furze J, De Costa SD, Hanlon M, Spencer AJ, Honeycutt JD, Wyllie DH, Gilbert SC et al. 2010. Long-term thermostabilization of live poxviral and adenoviral vaccine vectors at supraphysiological temperatures in carbohydrate glass. Sci Transl Med, 2 (19), pp. 19ra12. | Show Abstract | Read more

Live recombinant viral vectors based on adenoviruses and poxviruses are among the most promising platforms for development of new vaccines against diseases such as malaria, tuberculosis, and HIV-AIDS. Vaccines based on live viruses must remain infectious to be effective, so therefore need continuous refrigeration to maintain stability and viability, a requirement that can be costly and difficult, especially in developing countries. The sugars sucrose and trehalose are commonly used as stabilizing agents and cryoprotectants for biological products. Here, we have exploited the ability of these sugars to vitrify on desiccation to develop a thermostabilization technique for live viral vaccine vectors. By slowly drying vaccines suspended in solutions of these disaccharide stabilizers onto a filter-like support membrane at ambient temperature, an ultrathin glass is deposited on the fibers of the inert matrix. Immobilization of two recombinant vaccine vectors-E1/E3-deleted human adenovirus type 5 and modified vaccinia virus Ankara-in this glass on the membranes enabled complete recovery of viral titer and immunogenicity after storage at up to 45 degrees C for 6 months and even longer with minimal losses. Furthermore, the membrane carrying the stabilized vaccine can be incorporated into a holder attached to a syringe for almost simultaneous reconstitution and injection at point of use. The technology may potentially be developed for the deployment of viral vector-based biopharmaceuticals in resource-poor settings.

Rollier CS, Paranhos-Baccala G, Verschoor EJ, Verstrepen BE, Drexhage JAR, Fagrouch Z, Berland J-L, Komurian-Pradel F, Duverger B, Himoudi N et al. 2007. Vaccine-induced early control of hepatitis C virus infection in chimpanzees fails to impact on hepatic PD-1 and chronicity. Hepatology, 45 (3), pp. 602-613. | Show Abstract | Read more

UNLABELLED: Broad T cell and B cell responses to multiple HCV antigens are observed early in individuals who control or clear HCV infection. The prevailing hypothesis has been that similar immune responses induced by prophylactic immunization would reduce acute virus replication and protect exposed individuals from chronic infection. Here, we demonstrate that immunization of naïve chimpanzees with a multicomponent HCV vaccine induced robust HCV-specific immune responses, and that all vaccinees exposed to heterologous chimpanzee-adapted HCV 1b J4 significantly reduced viral RNA in serum by 84%, and in liver by 99% as compared to controls (P=0.024 and 0.028, respectively). However, despite control of HCV in plasma and liver in the acute period, in the chronic phase, 3 of 4 vaccinated animals developed persistent infection. Analysis of expression levels of proinflammatory cytokines in serial hepatic biopsies failed to reveal an association with vaccine outcome. However, expression of IDO, CTLA-4 [corrected] and PD-1 levels in liver correlated with clearance or chronicity. CONCLUSION: Despite early control of virus load, a virus-associated tolerogenic-like state can develop in certain individuals independent of vaccination history.

Rollier C, Depla E, Drexhage JAR, Verschoor EJ, Verstrepen BE, Fatmi A, Brinster C, Fournillier A, Whelan JA, Whelan M et al. 2004. Control of heterologous hepatitis C virus infection in chimpanzees is associated with the quality of vaccine-induced peripheral T-helper immune response. J Virol, 78 (1), pp. 187-196. | Show Abstract | Read more

Prophylactic hepatitis C virus (HCV) vaccine trials with human volunteers are pending. There is an important need for immunological end points which correlate with vaccine efficacy and which do not involve invasive procedures, such as liver biopsies. By using a multicomponent DNA priming-protein boosting vaccine strategy, naïve chimpanzees were immunized against HCV structural proteins (core, E1, and E2) as well as a nonstructural (NS3) protein. Following immunization, exposure to the heterologous HCV 1b J4 subtype resulted in a peak of plasma viremia which was lower in both immunized animals. Compared to the naïve infection control and nine additional historical controls which became chronic, vaccinee 2 (Vac2) rapidly resolved the infection, while the other (Vac1) clearly controlled HCV infection. Immunization induced antibodies, peptide-specific gamma interferon (IFN-gamma), protein-specific lymphoproliferative responses, IFN-gamma, interleukin-2 (IL-2), and IL-4 T-helper responses in both vaccinees. However, the specificities were markedly different: Vac2 developed responses which were lower in magnitude than those of Vac1 but which were biased towards Th1-type cytokine responses for E1 and NS3. This proof-of-principle study in chimpanzees revealed that immunization with a combination of nonstructural and structural antigens elicited T-cell responses associated with an alteration of the course of infection. Our findings provide data to support the concept that the quality of the response to conserved epitopes and the specific nature of the peripheral T-helper immune response are likely pivotal factors influencing the control and clearance of HCV infection.

Rollier C, Drexhage JAR, Verstrepen BE, Verschoor EJ, Bontrop RE, Koopman G, Heeney JL. 2003. Chronic hepatitis C virus infection established and maintained in chimpanzees independent of dendritic cell impairment. Hepatology, 38 (4), pp. 851-858. | Show Abstract | Read more

Chronic hepatitis C virus (HCV) infection in humans is associated with an impairment of dendritic cells (DC). It has been hypothesized that impairment of DC function may be a central mechanism facilitating the establishment of a chronic carrier state. However, the majority of patients studied with DC impairment to date have been identified and, thus, inadvertently selected because of clinical manifestations leading to their diagnosis, which may have been many years following actual infection. We set out to determine whether impaired DC function occurred in the earlier asymptomatic phase of infection and turned to a well-defined cohort of HCV-infected chimpanzees in which the specific date of infection and the nature of the inoculum were well characterized. Results revealed that, in contrast to the observations in human subjects with advanced clinical hepatitis, there was neither impairment of the allostimulatory capacity of monocyte-derived DC from HCV chronic carriers nor impairment of the maturation process. Decreased allostimulatory capacity was only detected in 2 animals and, interestingly, in those that possessed the highest viral loads. Nevertheless, HCV sequences were undetectable in any of the DC derived from HCV-infected chimpanzees. In conclusion, these findings suggest that the mechanisms of establishing persistent HCV infection are separate and independent from those responsible for impaired DC function. Indeed, the maturation and allostimulatory impairment, as described in patient studies, are not necessary prerequisites but rather possible consequences of persistent and active HCV infection associated with disease progression.

Rollier C, Sunyach C, Barraud L, Madani N, Jamard C, Trepo C, Cova L. 1999. Protective and therapeutic effect of DNA-based immunization against hepadnavirus large envelope protein. Gastroenterology, 116 (3), pp. 658-665. | Show Abstract | Read more

BACKGROUND & AIMS: Studies in the murine model suggest that injection of DNA encoding hepatitis B virus structural proteins is promising for the induction of a specific immune response. We used the duck hepatitis B virus (DHBV) model to study the protective and therapeutic effects of naked DNA immunization against hepadnaviral large envelope protein. METHODS: A pCI-preS/S plasmid expressing the DHBV large protein was used for intramuscular immunization of ducks. The humoral response was tested by enzyme-linked immunosorbent assay, immunoblotting, neutralization, and in vivo protection tests. For DNA therapy, DHBV-carrier ducks received four injections of this plasmid. Viremia was monitored for 10 months; thereafter, liver biopsies were performed. RESULTS: Immunization with pCI-preS/S plasmid induced a specific, long-lasting, neutralizing, and highly protective anti-preS humoral response in uninfected animals. After pCI-preS/S treatment, a significant and sustained decrease in serum and liver DHBV DNA was observed for carrier ducks compared with the controls. CONCLUSIONS: DNA immunization against DHBV large protein results in a potent and protective anti-preS response in the duck model. The results of long-term follow-up of DNA-treated chronically infected ducks are promising and show the usefulness of this model for the study of genetic immunization in chronic hepatitis B therapy.

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