Professor Teresa Lambe

Research Area: Immunology
Technology Exchange: Cellular immunology, Flow cytometry and Vaccine production and evaluation
Scientific Themes: Immunology & Infectious Disease and Tropical Medicine & Global Health
Keywords: Viral vectored vaccines, Pandemic and Emerging diseases

Vaccines are efficacious and cost-effective preventative healthcare measures and a global mainstay in protecting millions from infectious disease. The establishment of protective immune responses following vaccination and the formation of adaptive immune memory are crucial for ensuring long-lived immunity toward infectious disease.

Works based at The Jenner Institute have generated viral vectored vaccines efficacious in the establishment of immune memory toward candidate antigens. These include novel simian adenovirus and Modified vaccinia Ankara, which when used in heterologous prime-boost regimens are particularly successful at eliciting strong immune responses.

Increased global interdependence and the ease of human, animal and trade movements facilitate the continued emergence and re-emergence of novel infectious pathogens. There are a number of novel pathogens with recognised pandemic potential. Sporadic outbreaks or pandemic spread can have devastating health impacts, particularly on vulnerable populations.

We continue to develop and augment vaccines that can induce long-lived immunity toward emerging and re-emerging pathogens, particularly focusing on the development of vaccines that can confer heterologous protection. A number of these vaccines have progressed through preclinical works and are being assessed in clinical trials.

Name Department Institution Country
Professor Sarah C Gilbert Jenner Institute Oxford University, Old Road Campus Research Building United Kingdom
Professor Adrian VS Hill Jenner Institute Oxford University, Old Road Campus Research Building United Kingdom
Dr Edward Wright University of Westminster United Kingdom
Dr Nigel Temperton University of Kent United Kingdom
Professor Sir Andrew J McMichael NDM Research Building Oxford University, NDM Research Building United Kingdom
Professor Stephen Kent University of Melbourne Australia
Professor Simon J Draper Jenner Institute Oxford University, Old Road Campus Research Building United Kingdom
Associate Professor Katie Ewer Jenner Institute Oxford University, Old Road Campus Research Building United Kingdom
Dr Alexandra J Spencer Jenner Institute Oxford University, Old Road Campus Research Building United Kingdom
Sebastian S, Lambe T. 2018. Clinical Advances in Viral-Vectored Influenza Vaccines. Vaccines (Basel), 6 (2), pp. 29-29. | Show Abstract | Read more

Influenza-virus-mediated disease can be associated with high levels of morbidity and mortality, particularly in younger children and older adults. Vaccination is the primary intervention used to curb influenza virus infection, and the WHO recommends immunization for at-risk individuals to mitigate disease. Unfortunately, influenza vaccine composition needs to be updated annually due to antigenic shift and drift in the viral immunogen hemagglutinin (HA). There are a number of alternate vaccination strategies in current development which may circumvent the need for annual re-vaccination, including new platform technologies such as viral-vectored vaccines. We discuss the different vectored vaccines that have been or are currently in clinical trials, with a forward-looking focus on immunogens that may be protective against seasonal and pandemic influenza infection, in the context of viral-vectored vaccines. We also discuss future perspectives and limitations in the field that will need to be addressed before new vaccines can significantly impact disease levels.

Coughlan L, Sridhar S, Payne R, Edmans M, Milicic A, Venkatraman N, Lugonja B, Clifton L, Qi C, Folegatti PM et al. 2018. Corrigendum to "Heterologous Two-dose Vaccination with Simian Adenovirus and Poxvirus Vectors Elicits Long-lasting Cellular Immunity to Influenza Virus A in Healthy Adults" [EBioMedicine 29 (2018) 146-154]. EBioMedicine, 31 pp. 321. | Show Abstract | Read more

© 2018 The Authors The authors wish to point out that L. Coughlan and S. Sridhar were both at the Jenner Institute, University of Oxford, OX3 7DQ, UK, when the work for the paper was completed.

Coughlan L, Sridhar S, Payne R, Edmans M, Milicic A, Venkatraman N, Lugonja B, Clifton L, Qi C, Folegatti PM et al. 2018. Heterologous Two-Dose Vaccination with Simian Adenovirus and Poxvirus Vectors Elicits Long-Lasting Cellular Immunity to Influenza Virus A in Healthy Adults. EBioMedicine, 29 pp. 146-154. | Show Abstract | Read more

BACKGROUND: T-cell responses against highly conserved influenza antigens have been previously associated with protection. However, these immune responses are poorly maintained following recovery from influenza infection and are not boosted by inactivated influenza vaccines. We have previously demonstrated the safety and immunogenicity of two viral vectored vaccines, modified vaccinia virus Ankara (MVA) and the chimpanzee adenovirus ChAdOx1 expressing conserved influenza virus antigens, nucleoprotein (NP) and matrix protein-1 (M1). We now report on the safety and long-term immunogenicity of multiple combination regimes of these vaccines in young and older adults. METHODS: We conducted a Phase I open-label, randomized, multi-center study in 49 subjects aged 18-46years and 24 subjects aged 50years or over. Following vaccination, adverse events were recorded and the kinetics of the T cell response determined at multiple time points for up to 18months. FINDINGS: Both vaccines were well tolerated. A two dose heterologous vaccination regimen significantly increased the magnitude of pre-existing T-cell responses to NP and M1 after both doses in young and older adults. The fold-increase and peak immune responses after a single MVA-NP+M1 vaccination was significantly higher compared to ChAdOx1 NP+M1. In a mixed regression model, T-cell responses over 18months were significantly higher following the two dose vaccination regimen of MVA/ChAdOx1 NP+M1. INTERPRETATION: A two dose heterologous vaccination regimen of MVA/ChAdOx1 NP+M1 was safe and immunogenic in young and older adults, offering a promising vaccination strategy for inducing long-term broadly cross-reactive protection against influenza A. FUNDING SOURCE: Medical Research Council UK, NIHR BMRC Oxford.

Ewer K, Sebastian S, Spencer AJ, Gilbert S, Hill AVS, Lambe T. 2017. Chimpanzee adenoviral vectors as vaccines for outbreak pathogens. Hum Vaccin Immunother, 13 (12), pp. 3020-3032. | Show Abstract | Read more

The 2014-15 Ebola outbreak in West Africa highlighted the potential for large disease outbreaks caused by emerging pathogens and has generated considerable focus on preparedness for future epidemics. Here we discuss drivers, strategies and practical considerations for developing vaccines against outbreak pathogens. Chimpanzee adenoviral (ChAd) vectors have been developed as vaccine candidates for multiple infectious diseases and prostate cancer. ChAd vectors are safe and induce antigen-specific cellular and humoral immunity in all age groups, as well as circumventing the problem of pre-existing immunity encountered with human Ad vectors. For these reasons, such viral vectors provide an attractive platform for stockpiling vaccines for emergency deployment in response to a threatened outbreak of an emerging pathogen. Work is already underway to develop vaccines against a number of other outbreak pathogens and we will also review progress on these approaches here, particularly for Lassa fever, Nipah and MERS.

Munster VJ, Wells D, Lambe T, Wright D, Fischer RJ, Bushmaker T, Saturday G, van Doremalen N, Gilbert SC, de Wit E, Warimwe GM. 2017. Protective efficacy of a novel simian adenovirus vaccine against lethal MERS-CoV challenge in a transgenic human DPP4 mouse model. NPJ Vaccines, 2 (1), pp. 28. | Show Abstract | Read more

Middle East respiratory syndrome coronavirus (MERS-CoV) is a novel zoonotic virus that causes severe respiratory disease in humans with a case fatality rate close to 40%, but for which no vaccines are available. Here, we evaluated the utility of ChAdOx1, a promising replication-deficient simian adenovirus vaccine vector platform with an established safety profile in humans and dromedary camels, for MERS-CoV vaccine development. Using a transgenic lethal BALB/c MERS-CoV mouse model we showed that single dose intranasal or intramuscular immunisation with ChAdOx1 MERS, encoding full-length MERS-CoV Spike glycoprotein, is highly immunogenic and confers protection against lethal viral challenge. Immunogenicity and efficacy were comparable between immunisation routes. Together these data provide support for further evaluation of ChAdOx1 MERS vaccine in humans and dromedary camels, the animal reservoir of infection.

Tully CM, Chinnakannan S, Mullarkey CE, Ulaszewska M, Ferrara F, Temperton N, Gilbert SC, Lambe T. 2017. Novel Bivalent Viral-Vectored Vaccines Induce Potent Humoral and Cellular Immune Responses Conferring Protection against Stringent Influenza A Virus Challenge. J Immunol, 199 (4), pp. 1333-1341. | Show Abstract | Read more

Seasonal influenza viruses are a common cause of acute respiratory illness worldwide and generate a significant socioeconomic burden. Influenza viruses mutate rapidly, necessitating annual vaccine reformulation because traditional vaccines do not typically induce broad-spectrum immunity. In addition to seasonal infections, emerging pandemic influenza viruses present a continued threat to global public health. Pandemic influenza viruses have consistently higher attack rates and are typically associated with greater mortality compared with seasonal strains. Ongoing strategies to improve vaccine efficacy typically focus on providing broad-spectrum immunity; although B and T cells can mediate heterosubtypic responses, typical vaccine development will augment either humoral or cellular immunity. However, multipronged approaches that target several Ags may limit the generation of viral escape mutants. There are few vaccine platforms that can deliver multiple Ags and generate robust cellular and humoral immunity. In this article, we describe a novel vaccination strategy, tested preclinically in mice, for the delivery of novel bivalent viral-vectored vaccines. We show this strategy elicits potent T cell responses toward highly conserved internal Ags while simultaneously inducing high levels of Abs toward hemagglutinin. Importantly, these humoral responses generate long-lived plasma cells and generate Abs capable of neutralizing variant hemagglutinin-expressing pseudotyped lentiviruses. Significantly, these novel viral-vectored vaccines induce strong immune responses capable of conferring protection in a stringent influenza A virus challenge. Thus, this vaccination regimen induces lasting efficacy toward influenza. Importantly, the simultaneous delivery of dual Ags may alleviate the selective pressure that is thought to potentiate antigenic diversity in avian influenza viruses.

Alharbi NK, Padron-Regalado E, Thompson CP, Kupke A, Wells D, Sloan MA, Grehan K, Temperton N, Lambe T, Warimwe G et al. 2017. ChAdOx1 and MVA based vaccine candidates against MERS-CoV elicit neutralising antibodies and cellular immune responses in mice. Vaccine, 35 (30), pp. 3780-3788. | Show Abstract | Read more

The Middle East respiratory syndrome coronavirus (MERS-CoV) has infected more than 1900 humans, since 2012. The syndrome ranges from asymptomatic and mild cases to severe pneumonia and death. The virus is believed to be circulating in dromedary camels without notable symptoms since the 1980s. Therefore, dromedary camels are considered the only animal source of infection. Neither antiviral drugs nor vaccines are approved for veterinary or medical use despite active research on this area. Here, we developed four vaccine candidates against MERS-CoV based on ChAdOx1 and MVA viral vectors, two candidates per vector. All vaccines contained the full-length spike gene of MERS-CoV; ChAdOx1 MERS vaccines were produced with or without the leader sequence of the human tissue plasminogen activator gene (tPA) where MVA MERS vaccines were produced with tPA, but either the mH5 or F11 promoter driving expression of the spike gene. All vaccine candidates were evaluated in a mouse model in prime only or prime-boost regimens. ChAdOx1 MERS with tPA induced higher neutralising antibodies than ChAdOx1 MERS without tPA. A single dose of ChAdOx1 MERS with tPA elicited cellular immune responses as well as neutralising antibodies that were boosted to a significantly higher level by MVA MERS. The humoral immunogenicity of a single dose of ChAdOx1 MERS with tPA was equivalent to two doses of MVA MERS (also with tPA). MVA MERS with mH5 or F11 promoter induced similar antibody levels; however, F11 promoter enhanced the cellular immunogenicity of MVA MERS to significantly higher magnitudes. In conclusion, our study showed that MERS-CoV vaccine candidates could be optimized by utilising different viral vectors, various genetic designs of the vectors, or different regimens to increase immunogenicity. ChAdOx1 and MVA vectored vaccines have been safely evaluated in camels and humans and these MERS vaccine candidates should now be tested in camels and in clinical trials.

Lambe T, Bowyer G, Ewer KJ. 2017. A review of Phase I trials of Ebola virus vaccines: what can we learn from the race to develop novel vaccines? Philos Trans R Soc Lond B Biol Sci, 372 (1721), pp. 20160295-20160295. | Show Abstract | Read more

Sporadic outbreaks of Ebola virus infection have been documented since the mid-Seventies and viral exposure can lead to lethal haemorrhagic fever with case fatalities as high as 90%. There is now a comprehensive body of data from both ongoing and completed clinical trials assessing various vaccine strategies, which were rapidly advanced through clinical trials in response to the 2013-2016 Ebola virus disease (EVD) public health emergency. Careful consideration of immunogenicity post vaccination is essential but has been somewhat stifled because of the wide array of immunological assays and outputs that have been used in the numerous clinical trials. We discuss here the different aspects of the immune assays currently used in the Phase I clinical trials for Ebola virus vaccines, and draw comparisons across the immune outputs where possible; various trials have examined both cellular and humoral immunity in European and African cohorts. Assessment of the safety data, the immunological outputs and the ease of field deployment for the various vaccine modalities will help both the scientific community and policy-makers prioritize and potentially license vaccine candidates. If this can be achieved, the next outbreak of Ebola virus, or other emerging pathogen, can be more readily contained and will not have such widespread and devastating consequences.This article is part of the themed issue 'The 2013-2016 West African Ebola epidemic: data, decision-making and disease control'.

Spencer AJ, Longley RJ, Gola A, Ulaszewska M, Lambe T, Hill AVS. 2017. The Threshold of Protection from Liver-Stage Malaria Relies on a Fine Balance between the Number of Infected Hepatocytes and Effector CD8+ T Cells Present in the Liver. J Immunol, 198 (5), pp. 2006-2016. | Show Abstract | Read more

Since the demonstration of sterile protection afforded by injection of irradiated sporozoites, CD8+ T cells have been shown to play a significant role in protection from liver-stage malaria. This is, however, dependent on the presence of an extremely high number of circulating effector cells, thought to be necessary to scan, locate, and kill infected hepatocytes in the short time that parasites are present in the liver. We used an adoptive transfer model to elucidate the kinetics of the effector CD8+ T cell response in the liver following Plasmodium berghei sporozoite challenge. Although effector CD8+ T cells require <24 h to find, locate, and kill infected hepatocytes, active migration of Ag-specific CD8+ T cells into the liver was not observed during the 2-d liver stage of infection, as divided cells were only detected from day 3 postchallenge. However, the percentage of donor cells recruited into division was shown to indicate the level of Ag presentation from infected hepatocytes. By titrating the number of transferred Ag-specific effector CD8+ T cells and sporozoites, we demonstrate that achieving protection toward liver-stage malaria is reliant on CD8+ T cells being able to locate infected hepatocytes, resulting in a protection threshold dependent on a fine balance between the number of infected hepatocytes and CD8+ T cells present in the liver. With such a fine balance determining protection, achieving a high number of CD8+ T cells will be critical to the success of a cell-mediated vaccine against liver-stage malaria.

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.

Vanderven HA, Ana-Sosa-Batiz F, Jegaskanda S, Rockman S, Laurie K, Barr I, Chen W, Wines B, Hogarth PM, Lambe T et al. 2016. What Lies Beneath: Antibody Dependent Natural Killer Cell Activation by Antibodies to Internal Influenza Virus Proteins. EBioMedicine, 8 pp. 277-290. | Show Abstract | Read more

The conserved internal influenza proteins nucleoprotein (NP) and matrix 1 (M1) are well characterised for T cell immunity, but whether they also elicit functional antibodies capable of activating natural killer (NK) cells has not been explored. We studied NP and M1-specific ADCC activity using biochemical, NK cell activation and killing assays with plasma from healthy and influenza-infected subjects. Healthy adults had antibodies to M1 and NP capable of binding dimeric FcγRIIIa and activating NK cells. Natural symptomatic and experimental influenza infections resulted in a rise in antibody dependent NK cell activation post-infection to the hemagglutinin of the infecting strain, but changes in NK cell activation to M1 and NP were variable. Although antibody dependent killing of target cells infected with vaccinia viruses expressing internal influenza proteins was not detected, opsonising antibodies to NP and M1 likely contribute to an antiviral microenvironment by stimulating innate immune cells to secrete cytokines early in infection. We conclude that effector cell activating antibodies to conserved internal influenza proteins are common in healthy and influenza-infected adults. Given the significance of such antibodies in animal models of heterologous influenza infection, the definition of their importance and mechanism of action in human immunity to influenza is essential.

Ewer K, Rampling T, Venkatraman N, Bowyer G, Wright D, Lambe T, Imoukhuede EB, Payne R, Fehling SK, Strecker T et al. 2016. A Monovalent Chimpanzee Adenovirus Ebola Vaccine Boosted with MVA. N Engl J Med, 374 (17), pp. 1635-1646. | Show Abstract | Read more

BACKGROUND: The West African outbreak of Ebola virus disease that peaked in 2014 has caused more than 11,000 deaths. The development of an effective Ebola vaccine is a priority for control of a future outbreak. METHODS: In this phase 1 study, we administered a single dose of the chimpanzee adenovirus 3 (ChAd3) vaccine encoding the surface glycoprotein of Zaire ebolavirus (ZEBOV) to 60 healthy adult volunteers in Oxford, United Kingdom. The vaccine was administered in three dose levels--1×10(10) viral particles, 2.5×10(10) viral particles, and 5×10(10) viral particles--with 20 participants in each group. We then assessed the effect of adding a booster dose of a modified vaccinia Ankara (MVA) strain, encoding the same Ebola virus glycoprotein, in 30 of the 60 participants and evaluated a reduced prime-boost interval in another 16 participants. We also compared antibody responses to inactivated whole Ebola virus virions and neutralizing antibody activity with those observed in phase 1 studies of a recombinant vesicular stomatitis virus-based vaccine expressing a ZEBOV glycoprotein (rVSV-ZEBOV) to determine relative potency and assess durability. RESULTS: No safety concerns were identified at any of the dose levels studied. Four weeks after immunization with the ChAd3 vaccine, ZEBOV-specific antibody responses were similar to those induced by rVSV-ZEBOV vaccination, with a geometric mean titer of 752 and 921, respectively. ZEBOV neutralization activity was also similar with the two vaccines (geometric mean titer, 14.9 and 22.2, respectively). Boosting with the MVA vector increased virus-specific antibodies by a factor of 12 (geometric mean titer, 9007) and increased glycoprotein-specific CD8+ T cells by a factor of 5. Significant increases in neutralizing antibodies were seen after boosting in all 30 participants (geometric mean titer, 139; P<0.001). Virus-specific antibody responses in participants primed with ChAd3 remained positive 6 months after vaccination (geometric mean titer, 758) but were significantly higher in those who had received the MVA booster (geometric mean titer, 1750; P<0.001). CONCLUSIONS: The ChAd3 vaccine boosted with MVA elicited B-cell and T-cell immune responses to ZEBOV that were superior to those induced by the ChAd3 vaccine alone. (Funded by the Wellcome Trust and others; ClinicalTrials.gov number, NCT02240875.).

Mullin J, Ahmed MS, Sharma R, Upile N, Beer H, Achar P, Puksuriwong S, Ferrara F, Temperton N, McNamara P et al. 2016. Activation of cross-reactive mucosal T and B cell responses in human nasopharynx-associated lymphoid tissue in vitro by Modified Vaccinia Ankara-vectored influenza vaccines. Vaccine, 34 (14), pp. 1688-1695. | Show Abstract | Read more

UNLABELLED: Recent efforts have been focused on the development of vaccines that could induce broad immunity against influenza virus, either through T cell responses to conserved internal antigens or B cell response to cross-reactive haemagglutinin (HA). We studied the capacity of Modified Vaccinia Ankara (MVA)-vectored influenza vaccines to induce cross-reactive immunity to influenza virus in human nasopharynx-associated lymphoid tissue (NALT) in vitro. Adenotonsillar cells were isolated and stimulated with MVA vaccines expressing either conserved nucleoprotein (NP) and matrix protein 1 (M1) (MVA-NP-M1) or pandemic H1N1 HA (MVA-pdmH1HA). The MVA vaccine uptake and expression, and T and B cell responses were analyzed. MVA-vectored vaccines were highly efficient infecting NALT and vaccine antigens were highly expressed by B cells. MVA-NP-M1 elicited T cell response with greater numbers of IFNγ-producing CD4+ T cells and tissue-resident memory T cells than controls. MVA-pdmH1HA induced cross-reactive anti-HA antibodies to a number of influenza subtypes, in an age-dependent manner. The cross-reactive antibodies include anti-avian H5N1 and mainly target HA2 domain. CONCLUSION: MVA vaccines are efficient in infecting NALT and the vaccine antigen is highly expressed by B cells. MVA vaccines expressing conserved influenza antigens induce cross-reactive T and B cell responses in human NALT in vitro, suggesting the potential as mucosal vaccines for broader immunity against influenza.

Alharbi NK, Spencer AJ, Salman AM, Tully CM, Chinnakannan SK, Lambe T, Yamaguchi Y, Morris SJ, Orubu T, Draper SJ et al. 2016. Enhancing cellular immunogenicity of MVA-vectored vaccines by utilizing the F11L endogenous promoter. Vaccine, 34 (1), pp. 49-55. | Show Abstract | Read more

Modified vaccinia virus Ankara (MVA)-vectored vaccines against malaria, influenza, tuberculosis and recently Ebola virus are in clinical development. Although this vector is safe and immunogenic in humans, efforts remain on-going to enhance immunogenicity through various approaches such as using stronger promoters to boost transgene expression. We previously reported that endogenous MVA promoters such as pB8 and pF11 increased transgene expression and immunogenicity, as compared to the conventional p7.5 promoter. Here, we show that both promoters also rivalled the mH5 promoter in enhancing MVA immunogenicity. We investigated the mechanisms behind this improved immunogenicity and show that it was a result of strong early transgene expression in vivo, rather than in vitro as would normally be assessed. Moreover, keeping the TK gene intact resulted in a modest improvement in immunogenicity. Utilizing pB8 or pF11 as ectopic promoters at the TK locus instead of their natural loci also increased transgene expression and immunogenicity. In addition to a reporter antigen, the pF11 promoter was tested with the expression of two vaccine antigens for which cellular immunogenicity was significantly increased as compared to the p7.5 promoter. Our data support the use of the pF11 and pB8 promoters for improved immunogenicity in future MVA-vectored candidate vaccines.

Lambe T, Rampling T, Samuel D, Bowyer G, Ewer KJ, Venkatraman N, Edmans M, Dicks S, Hill AVS, Tedder RS, Gilbert SC. 2016. Detection of Vaccine-Induced Antibodies to Ebola Virus in Oral Fluid. Open Forum Infect Dis, 3 (1), pp. ofw031. | Show Abstract | Read more

Blood sampling to assess production of antigen-specific antibodies after immunization is commonly performed, but it presents logistical difficulties for trials carried out during an infectious disease outbreak. In this study, we show that antibodies may be reliably detected in oral fluid collected in a minimally invasive manner without use of sharps. Clinical Trials Registration. NCT02240875.

Coughlan L, Lambe T. 2015. Measuring Cellular Immunity to Influenza: Methods of Detection, Applications and Challenges. Vaccines (Basel), 3 (2), pp. 293-319. | Show Abstract | Read more

Influenza A virus is a respiratory pathogen which causes both seasonal epidemics and occasional pandemics; infection continues to be a significant cause of mortality worldwide. Current influenza vaccines principally stimulate humoral immune responses that are largely directed towards the variant surface antigens of influenza. Vaccination can result in an effective, albeit strain-specific antibody response and there is a need for vaccines that can provide superior, long-lasting immunity to influenza. Vaccination approaches targeting conserved viral antigens have the potential to provide broadly cross-reactive, heterosubtypic immunity to diverse influenza viruses. However, the field lacks consensus on the correlates of protection for cellular immunity in reducing severe influenza infection, transmission or disease outcome. Furthermore, unlike serological methods such as the standardized haemagglutination inhibition assay, there remains a large degree of variation in both the types of assays and method of reporting cellular outputs. T-cell directed immunity has long been known to play a role in ameliorating the severity and/or duration of influenza infection, but the precise phenotype, magnitude and longevity of the requisite protective response is unclear. In order to progress the development of universal influenza vaccines, it is critical to standardize assays across sites to facilitate direct comparisons between clinical trials.

Tully CM, Lambe T, Gilbert SC, Hill AVS. 2015. Emergency Ebola response. a new approach to the rapid design and development of vaccines against emerging diseases (vol 15, pg 356, 2015) LANCET INFECTIOUS DISEASES, 15 (3), pp. 263-263. | Read more

Tully CM, Lambe T, Gilbert SC, Hill AVS. 2015. Corrections. Emergency Ebola response: a new approach to the rapid design and development of vaccines against emerging diseases. Lancet Infect Dis, 15 (3), pp. 263. | Read more

Tully CM, Lambe T, Gilbert SC, Hill AVS. 2015. Emergency Ebola response: A new approach to the rapid design and development of vaccines against emerging diseases The Lancet Infectious Diseases, 15 (3), pp. 356-359. | Show Abstract | Read more

© 2015 Elsevier Ltd. The epidemic of Ebola virus disease has spread at an alarming rate despite containment efforts. As a result, unprecedented large-scale international response efforts have been made in an attempt to gain control of the outbreak and reduce transmission. Several international consortia have been formed in a remarkable worldwide collaborative effort to expedite trials of two candidate Ebola virus vaccines: cAd3-EBOZ and rVSV-EBOV. In parallel, both vaccines are being manufactured in large amounts to enable future rapid deployment for management of the crisis.

Tully CM, Lambe T, Gilbert SC, Hill AVS. 2015. Emergency Ebola response: a new approach to the rapid design and development of vaccines against emerging diseases. Lancet Infect Dis, 15 (3), pp. 356-359. | Show Abstract | Read more

The epidemic of Ebola virus disease has spread at an alarming rate despite containment efforts. As a result, unprecedented large-scale international response efforts have been made in an attempt to gain control of the outbreak and reduce transmission. Several international consortia have been formed in a remarkable worldwide collaborative effort to expedite trials of two candidate Ebola virus vaccines: cAd3-EBOZ and rVSV-EBOV. In parallel, both vaccines are being manufactured in large amounts to enable future rapid deployment for management of the crisis.

Mullin J, Ahmed MS, Upile N, Vaughan C, McNamara P, Beer H, McCormick M, Lambe T, Gilbert SC, Zhang Q. 2014. Evaluating the potential of novel virus-vectored vaccines as intranasal vaccine candidates for broad immunity against influenza IMMUNOLOGY, 143 pp. 78-78.

Cited:

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Antrobus RD, Coughlan L, Berthoud TK, Dicks MD, Hill AVS, Lambe T, Gilbert SC. 2014. Clinical assessment of a novel recombinant simian adenovirus ChAdOx1 as a vectored vaccine expressing conserved influenza a antigens Molecular Therapy, 22 (3), pp. 668-674. | Show Abstract | Read more

Adenoviruses are potent vectors for inducing and boosting cellular immunity to encoded recombinant antigens. However, the widespread seroprevalence of neutralizing antibodies to common human adenovirus serotypes limits their use. Simian adenoviruses do not suffer from the same drawbacks. We have constructed a replication-deficient chimpanzee adenovirus-vectored vaccine expressing the conserved influenza antigens, nucleoprotein (NP), and matrix protein 1 (M1). Here, we report safety and T-cell immunogenicity following vaccination with this novel recombinant simian adenovirus, ChAdOx1 NP+M1, in a first in human dose-escalation study using a 3+3 study design, followed by boosting with modified vaccinia virus Ankara expressing the same antigens in some volunteers. We demonstrate ChAdOx1 NP+M1 to be safe and immunogenic. ChAdOx1 is a promising vaccine vector that could be used to deliver vaccine antigens where strong cellular immune responses are required for protection. © The American Society of Gene & Cell Therapy.

Antrobus RD, Coughlan L, Berthoud TK, Dicks MD, Hill AV, Lambe T, Gilbert SC. 2014. Clinical assessment of a novel recombinant simian adenovirus ChAdOx1 as a vectored vaccine expressing conserved Influenza A antigens. Mol Ther, 22 (3), pp. 668-674. | Show Abstract | Read more

Adenoviruses are potent vectors for inducing and boosting cellular immunity to encoded recombinant antigens. However, the widespread seroprevalence of neutralizing antibodies to common human adenovirus serotypes limits their use. Simian adenoviruses do not suffer from the same drawbacks. We have constructed a replication-deficient chimpanzee adenovirus-vectored vaccine expressing the conserved influenza antigens, nucleoprotein (NP), and matrix protein 1 (M1). Here, we report safety and T-cell immunogenicity following vaccination with this novel recombinant simian adenovirus, ChAdOx1 NP+M1, in a first in human dose-escalation study using a 3+3 study design, followed by boosting with modified vaccinia virus Ankara expressing the same antigens in some volunteers. We demonstrate ChAdOx1 NP+M1 to be safe and immunogenic. ChAdOx1 is a promising vaccine vector that could be used to deliver vaccine antigens where strong cellular immune responses are required for protection.

Crawford G, Enders A, Gileadi U, Stankovic S, Zhang Q, Lambe T, Crockford TL, Lockstone HE, Freeman A, Arkwright PD et al. 2013. DOCK8 is critical for the survival and function of NKT cells. Blood, 122 (12), pp. 2052-2061. | Show Abstract | Read more

Patients with the dedicator of cytokinesis 8 (DOCK8) immunodeficiency syndrome suffer from recurrent viral and bacterial infections, hyper-immunoglobulin E levels, eczema, and greater susceptibility to cancer. Because natural killer T (NKT) cells have been implicated in these diseases, we asked if these cells were affected by DOCK8 deficiency. Using a mouse model, we found that DOCK8 deficiency resulted in impaired NKT cell development, principally affecting the formation and survival of long-lived, differentiated NKT cells. In the thymus, DOCK8-deficient mice lack a terminally differentiated subset of NK1.1(+) NKT cells expressing the integrin CD103, whereas in the liver, DOCK8-deficient NKT cells express reduced levels of the prosurvival factor B-cell lymphoma 2 and the integrin lymphocyte function-associated antigen 1. Although the initial NKT cell response to antigen is intact in the absence of DOCK8, their ongoing proliferative and cytokine responses are impaired. Importantly, a similar defect in NKT cell numbers was detected in DOCK8-deficient humans, highlighting the relevance of the mouse model. In conclusion, our data demonstrate that DOCK8 is required for the development and survival of mature NKT cells, consistent with the idea that DOCK8 mediates survival signals within a specialized niche. Accordingly, impaired NKT cell numbers and function are likely to contribute to the susceptibility of DOCK8-deficient patients to recurrent infections and malignant disease.

Mullarkey CE, Boyd A, van Laarhoven A, Lefevre EA, Veronica Carr B, Baratelli M, Molesti E, Temperton NJ, Butter C, Charleston B et al. 2013. Improved adjuvanting of seasonal influenza vaccines: preclinical studies of MVA-NP+M1 coadministration with inactivated influenza vaccine. Eur J Immunol, 43 (7), pp. 1940-1952. | Show Abstract | Read more

Licensed seasonal influenza vaccines induce antibody (Ab) responses against influenza hemagglutinin (HA) that are limited in their ability to protect against different strains of influenza. Cytotoxic T lymphocytes recognizing the conserved internal nucleoprotein (NP) and matrix protein (M1) are capable of mediating a cross-subtype immune response against influenza. Modified vaccinia Ankara (MVA) virus encoding NP and M1 (MVA-NP+M1) is designed to boost preexisting T-cell responses in adults in order to elicit a cross-protective immune response. We examined the coadministration of HA protein formulations and candidate MVA-NP+M1 influenza vaccines in murine, avian, and swine models. Ab responses postimmunization were measured by ELISA and pseudotype neutralization assays. Here, we demonstrate that MVA-NP+M1 can act as an adjuvant enhancing Ab responses to HA while simultaneously inducing potent T-cell responses to conserved internal Ags. We show that this regimen leads to the induction of cytophilic Ab isotypes that are capable of inhibiting hemagglutination and in the context of H5 exhibit cross-clade neutralization. The simultaneous induction of T cells and Ab responses has the potential to improve seasonal vaccine performance and could be employed in pandemic situations.

Lambe T, Carey JB, Li Y, Spencer AJ, van Laarhoven A, Mullarkey CE, Vrdoljak A, Moore AC, Gilbert SC. 2013. Immunity against heterosubtypic influenza virus induced by adenovirus and MVA expressing nucleoprotein and matrix protein-1. Sci Rep, 3 (1), pp. 1443. | Show Abstract | Read more

Alternate prime/boost vaccination regimens employing recombinant replication-deficient adenovirus or MVA, expressing Influenza A virus nucleoprotein and matrix protein 1, induced antigen-specific T cell responses in intradermally (ID) vaccinated mice; with the strongest responses resulting from Ad/MVA immunization. In BALB/C mice the immunodominant response was shifted from the previously identified immunodominant epitope to a novel epitope when the antigen was derived from A/Panama/2007/1999 rather than A/PR/8. Alternate immunization routes did not affect the magnitude of antigen-specific systemic IFN-γ response, but higher CD8(+) T-cell IFN-γ immune responses were seen in the bronchoalveolar lavage following intransal (IN) boosting after intramuscular (IM) priming, whilst higher splenic antigen-specific CD8(+) T cell IFN-γ was seen following IM boosting. Partial protection against heterologous influenza virus challenge was achieved following either IM/IM or IM/IN but not ID/ID immunization. These data may be of relevance for the design of optimal immunization regimens for human influenza vaccines, especially for influenza-naïve infants.

Wegmann F, Gartlan KH, Harandi AM, Brinckmann SA, Coccia M, Hillson WR, Kok WL, Cole S, Ho L-P, Lambe T et al. 2012. Polyethyleneimine is a potent mucosal adjuvant for viral glycoprotein antigens. Nat Biotechnol, 30 (9), pp. 883-888. | Show Abstract | Read more

Protection against mucosally transmitted infections probably requires immunity at the site of pathogen entry, yet there are no mucosal adjuvant formulations licensed for human use. Polyethyleneimine (PEI) represents a family of organic polycations used as nucleic acid transfection reagents in vitro and DNA vaccine delivery vehicles in vivo. Here we show that diverse PEI forms have potent mucosal adjuvant activity for viral subunit glycoprotein antigens. A single intranasal administration of influenza hemagglutinin or herpes simplex virus type-2 (HSV-2) glycoprotein D with PEI elicited robust antibody-mediated protection from an otherwise lethal infection, and was superior to existing experimental mucosal adjuvants. PEI formed nanoscale complexes with antigen, which were taken up by antigen-presenting cells in vitro and in vivo, promoted dendritic cell trafficking to draining lymph nodes and induced non-proinflammatory cytokine responses. PEI adjuvanticity required release of host double-stranded DNA that triggered Irf3-dependent signaling. PEI therefore merits further investigation as a mucosal adjuvant for human use.

Lambe T. 2012. Novel viral vectored vaccines for the prevention of influenza. Mol Med, 18 (8), pp. 1153-1160. | Show Abstract | Read more

Influenza represents a substantial global healthcare burden, with annual epidemics resulting in 3-5 million cases of severe illness with a significant associated mortality. In addition, the risk of a virulent and lethal influenza pandemic has generated widespread and warranted concern. Currently licensed influenza vaccines are limited in their ability to induce efficacious and long-lasting herd immunity. In addition, and as evidenced by the H1N1 pandemic in 2009, there can be a significant delay between the emergence of a pandemic influenza and an effective, antibody-inducing vaccine. There is, therefore, a continued need for new, efficacious vaccines conferring cross-clade protection-obviating the need for biannual reformulation of seasonal influenza vaccines. Development of such a vaccine would yield enormous health benefits to society and also greatly reduce the associated global healthcare burden. There are a number of alternative influenza vaccine technologies being assessed both preclinically and clinically. In this review we discuss viral vectored vaccines, either recombinant live-attenuated or replication-deficient viruses, which are current lead candidates for inducing efficacious and long-lasting immunity toward influenza viruses. These alternate influenza vaccines offer real promise to deliver viable alternatives to currently deployed vaccines and more importantly may confer long-lasting and universal protection against influenza viral infection.

Lambe T, Spencer AJ, Mullarkey CE, Antrobus RD, Yu L-M, de Whalley P, Thompson BAV, Jones C, Chalk J, Kerridge S et al. 2012. T-cell responses in children to internal influenza antigens, 1 year after immunization with pandemic H1N1 influenza vaccine, and response to revaccination with seasonal trivalent-inactivated influenza vaccine. Pediatr Infect Dis J, 31 (6), pp. e86-e91. | Show Abstract | Read more

BACKGROUND: During seasonal influenza epidemics, 5-15% of the population are affected with an illness having a nontrivial mortality, morbidity and economic burden. Inactivated influenza vaccines are routinely used to prevent influenza infection, primarily by inducing humoral immunity. In addition, trivalent-inactivated influenza vaccines have previously been shown to boost influenza-specific T-cell responses in a small percentage of adults. We investigate here the influenza-specific T-cell response, in children, 1 year after pandemic H1N1 vaccination and the ability to boost the T-cell response with trivalent-inactivated influenza immunization. METHODS: Peripheral blood mononuclear cells (PBMCs) were isolated from children previously vaccinated with pandemic H1N1 vaccine, pre- and postseasonal 2010-2011 trivalent influenza vaccine (TIV) vaccination. Samples were analyzed by interferon-gamma enzyme-linked immunosorbent spot for reactogenicity toward internal influenza antigens (nucleoprotein, matrix protein 1 and nonstructural protein 1). RESULTS: Basal ex vivo T-cell responses to nucleoprotein, matrix protein 1 and nonstructural protein 1 measured by interferon-gamma enzyme-linked immunosorbent spot assay were significantly higher in those children who had previously received an AS03B-adjuvanted split virion pandemic vaccine 12 months earlier rather than a nonadjuvanted whole virion vaccine. Boosting of these responses, 21 days after 2010/2011 seasonal TIV vaccination was observed regardless of age or prior pandemic vaccination regime, although boosting was greater in those groups with the lowest initial response. CONCLUSIONS: We show here that children previously vaccinated with the 2009 pandemic H1N1 vaccine have measurable T-cell responses 1 year after vaccination. The magnitudes of these responses are dependent on both age of vaccine and type of pandemic H1N1 vaccine used. After 2010/2011 seasonal TIV vaccination, these T-cell responses undergo a small but significant boost.

Lillie PJ, Berthoud TK, Powell TJ, Lambe T, Mullarkey C, Spencer AJ, Hamill M, Peng Y, Blais M-E, Duncan CJA et al. 2012. Preliminary assessment of the efficacy of a T-cell-based influenza vaccine, MVA-NP+M1, in humans. Clin Infect Dis, 55 (1), pp. 19-25. | Show Abstract | Read more

BACKGROUND: The novel influenza vaccine MVA-NP+M1 is designed to boost cross-reactive T-cell responses to internal antigens of the influenza A virus that are conserved across all subtypes, providing protection against both influenza disease and virus shedding against all influenza A viruses. Following a phase 1 clinical study that demonstrated vaccine safety and immunogenicity, a phase 2a vaccination and influenza challenge study has been conducted in healthy adult volunteers. METHODS: Volunteers with no measurable serum antibodies to influenza A/Wisconsin/67/2005 received either a single vaccination with MVA-NP+M1 or no vaccination. T-cell responses to the vaccine antigens were measured at enrollment and again prior to virus challenge. All volunteers underwent intranasal administration of influenza A/Wisconsin/67/2005 while in a quarantine unit and were monitored for symptoms of influenza disease and virus shedding. RESULTS: Volunteers had a significantly increased T-cell response to the vaccine antigens following a single dose of the vaccine, with an increase in cytolytic effector molecules. Intranasal influenza challenge was undertaken without safety issues. Two of 11 vaccinees and 5 of 11 control subjects developed laboratory-confirmed influenza (symptoms plus virus shedding). Symptoms of influenza were less pronounced in the vaccinees and there was a significant reduction in the number of days of virus shedding in those vaccinees who developed influenza (mean, 1.09 days in controls, 0.45 days in vaccinees, P = .036). CONCLUSIONS: This study provides the first demonstration of clinical efficacy of a T-cell-based influenza vaccine and indicates that further clinical development should be undertaken. CLINICAL TRIALS REGISTRATION: NCT00993083.

Lambe T, Spencer AJ, Mullarkey CE, Antrobus RD, Yu LM, de Whalley P, Thompson BAV, Jones C, Chalk J, Kerridge S et al. 2012. T-Cell Responses in Children to Internal Influenza Antigens, 1 Year After Immunization with Pandemic H1N1 Influenza Vaccine, and Response to Revaccination with Seasonal Trivalent-inactivated Influenza Vaccine Pediatric Infectious Disease Journal, 31 (6), pp. e86-e91. | Read more

Orubu T, Alharbi NK, Lambe T, Gilbert SC, Cottingham MG. 2012. Expression and cellular immunogenicity of a transgenic antigen driven by endogenous poxviral early promoters at their authentic loci in MVA. PLoS One, 7 (6), pp. e40167. | Show Abstract | Read more

CD8(+) T cell responses to vaccinia virus are directed almost exclusively against early gene products. The attenuated strain modified vaccinia virus Ankara (MVA) is under evaluation in clinical trials of new vaccines designed to elicit cellular immune responses against pathogens including Plasmodium spp., M. tuberculosis and HIV-1. All of these recombinant MVAs (rMVA) utilize the well-established method of linking the gene of interest to a cloned poxviral promoter prior to insertion into the viral genome at a suitable locus by homologous recombination in infected cells. Using BAC recombineering, we show that potent early promoters that drive expression of non-functional or non-essential MVA open reading frames (ORFs) can be harnessed for immunogenic expression of recombinant antigen. Precise replacement of the MVA orthologs of C11R, F11L, A44L and B8R with a model antigen positioned to use the same translation initiation codon allowed early transgene expression similar to or slightly greater than that achieved by the commonly-used p7.5 or short synthetic promoters. The frequency of antigen-specific CD8(+) T cells induced in mice by single shot or adenovirus-prime, rMVA-boost vaccination were similarly equal or marginally enhanced using endogenous promoters at their authentic genomic loci compared to the traditional constructs. The enhancement in immunogenicity observed using the C11R or F11L promoters compared with p7.5 was similar to that obtained with the mH5 promoter compared with p7.5. Furthermore, the growth rates of the viruses were unimpaired and the insertions were genetically stable. Insertion of a transgenic ORF in place of a viral ORF by BAC recombineering can thus provide not only a potent promoter, but also, concomitantly, a suitable insertion site, potentially facilitating development of MVA vaccines expressing multiple recombinant antigens.

Antrobus RD, Lillie PJ, Berthoud TK, Spencer AJ, McLaren JE, Ladell K, Lambe T, Milicic A, Price DA, Hill AVS, Gilbert SC. 2012. A T cell-inducing influenza vaccine for the elderly: safety and immunogenicity of MVA-NP+M1 in adults aged over 50 years. PLoS One, 7 (10), pp. e48322. | Show Abstract | Read more

BACKGROUND: Current influenza vaccines have reduced immunogenicity and are of uncertain efficacy in older adults. We assessed the safety and immunogenicity of MVA-NP+M1, a viral-vectored influenza vaccine designed to boost memory T cell responses, in a group of older adults. METHODS: Thirty volunteers (aged 50-85) received a single intramuscular injection of MVA-NP+M1 at a dose of 1·5×10(8) plaque forming units (pfu). Safety and immunogenicity were assessed over a period of one year. The frequency of T cells specific for nucleoprotein (NP) and matrix protein 1 (M1) was determined by interferon-gamma (IFN-γ) ELISpot, and their phenotypic and functional properties were characterized by polychromatic flow cytometry. In a subset of M1-specific CD8(+) T cells, T cell receptor (TCR) gene expression was evaluated using an unbiased molecular approach. RESULTS: Vaccination with MVA-NP+M1 was well tolerated. ELISpot responses were boosted significantly above baseline following vaccination. Increases were detected in both CD4(+) and CD8(+) T cell subsets. Clonality studies indicated that MVA-NP+M1 expanded pre-existing memory CD8(+) T cells, which displayed a predominant CD27(+)CD45RO(+)CD57(-)CCR7(-) phenotype both before and after vaccination. CONCLUSIONS: MVA-NP+M1 is safe and immunogenic in older adults. Unlike seasonal influenza vaccination, the immune responses generated by MVA-NP+M1 are similar between younger and older individuals. A T cell-inducing vaccine such as MVA-NP+M1 may therefore provide a way to circumvent the immunosenescence that impairs routine influenza vaccination. TRIAL REGISTRATION: ClinicalTrials.gov NCT00942071.

Randall KL, Chan SS-Y, Ma CS, Fung I, Mei Y, Yabas M, Tan A, Arkwright PD, Al Suwairi W, Lugo Reyes SO et al. 2011. DOCK8 deficiency impairs CD8 T cell survival and function in humans and mice. J Exp Med, 208 (11), pp. 2305-2320. | Show Abstract | Read more

In humans, DOCK8 immunodeficiency syndrome is characterized by severe cutaneous viral infections. Thus, CD8 T cell function may be compromised in the absence of DOCK8. In this study, by analyzing mutant mice and humans, we demonstrate a critical, intrinsic role for DOCK8 in peripheral CD8 T cell survival and function. DOCK8 mutation selectively diminished the abundance of circulating naive CD8 T cells in both species, and in DOCK8-deficient humans, most CD8 T cells displayed an exhausted CD45RA(+)CCR7(-) phenotype. Analyses in mice revealed the CD8 T cell abnormalities to be cell autonomous and primarily postthymic. DOCK8 mutant naive CD8 T cells had a shorter lifespan and, upon encounter with antigen on dendritic cells, exhibited poor LFA-1 synaptic polarization and a delay in the first cell division. Although DOCK8 mutant T cells underwent near-normal primary clonal expansion after primary infection with recombinant influenza virus in vivo, they showed greatly reduced memory cell persistence and recall. These findings highlight a key role for DOCK8 in the survival and function of human and mouse CD8 T cells.

Berthoud TK, Hamill M, Lillie PJ, Hwenda L, Collins KA, Ewer KJ, Milicic A, Poyntz HC, Lambe T, Fletcher HA et al. 2011. Potent CD8+ T-cell immunogenicity in humans of a novel heterosubtypic influenza A vaccine, MVA-NP+M1. Clin Infect Dis, 52 (1), pp. 1-7. | Show Abstract | Read more

BACKGROUND: Influenza A viruses cause occasional pandemics and frequent epidemics. Licensed influenza vaccines that induce high antibody titers to the highly polymorphic viral surface antigen hemagglutinin must be re-formulated and readministered annually. A vaccine providing protective immunity to the highly conserved internal antigens could provide longer-lasting protection against multiple influenza subtypes. METHODS: We prepared a Modified Vaccinia virus Ankara (MVA) vector encoding nucleoprotein and matrix protein 1 (MVA-NP+M1) and conducted a phase I clinical trial in healthy adults. RESULTS: The vaccine was generally safe and well tolerated, with significantly fewer local side effects after intramuscular rather than intradermal administration. Systemic side effects increased at the higher dose in both frequency and severity, with 5 out of 8 volunteers experiencing severe nausea/vomiting, malaise, or rigors. Ex vivo T-cell responses to NP and M1 measured by IFN-γ ELISPOT assay were significantly increased after vaccination (prevaccination median of 123 spot-forming units/million peripheral blood mononuclear cells, postvaccination peak response median 339, 443, and 1443 in low-dose intradermal, low-dose intramuscular, and high-dose intramuscular groups, respectively), and the majority of the antigen-specific T cells were CD8(+). CONCLUSIONS: We conclude that the vaccine was both safe and remarkably immunogenic, leading to frequencies of responding T cells that appear to be much higher than those induced by any other influenza vaccination approach. Further studies will be required to find the optimum dose and to assess whether the increased T-cell response to conserved influenza proteins results in protection from influenza disease.

Lambe T, Crawford G, Johnson AL, Crockford TL, Bouriez-Jones T, Smyth AM, Pham THM, Zhang Q, Freeman AF, Cyster JG et al. 2011. DOCK8 is essential for T-cell survival and the maintenance of CD8+ T-cell memory. Eur J Immunol, 41 (12), pp. 3423-3435. | Show Abstract | Read more

Deficiency in the guanine nucleotide exchange factor dedicator of cytokinesis 8 (DOCK8) causes a human immunodeficiency syndrome associated with recurrent sinopulmonary and viral infections. We have recently identified a DOCK8-deficient mouse strain, carrying an ethylnitrosourea-induced splice-site mutation that shows a failure to mature a humoral immune response due to the loss of germinal centre B cells. In this study, we turned to T-cell immunity to investigate further the human immunodeficiency syndrome and its association with decreased peripheral CD4(+) and CD8(+) T cells. Characterisation of the DOCK8-deficient mouse revealed T-cell lymphopenia, with increased T-cell turnover and decreased survival. Egress of mature CD4(+) thymocytes was reduced with increased migration of these cells to the chemokine CXCL12. However, despite the two-fold reduction in peripheral naïve T cells, the DOCK8-deficient mice generated a normal primary CD8(+) immune response and were able to survive acute influenza virus infection. The limiting effect of DOCK8 was in the normal survival of CD8(+) memory T cells after infection. These findings help to explain why DOCK8-deficient patients are susceptible to recurrent infections and provide new insights into how T-cell memory is sustained.

Randall KL, Lambe T, Johnson AL, Treanor B, Kucharska E, Domaschenz H, Whittle B, Tze LE, Enders A, Crockford TL et al. 2010. Dock8 mutations cripple B cell immunological synapses, germinal centers and long-lived antibody production (vol 10, pg 1283, 2009) NATURE IMMUNOLOGY, 11 (7), pp. 644-644. | Read more

Johnson AL, Aravind L, Shulzhenko N, Morgun A, Choi SY, Crockford TL, Lambe T, Domaschenz H, Kucharska EM, Zheng L et al. 2010. Themis is a member of a new metazoan gene family and is required for the completion of thymocyte positive selection. Nat Immunol, 11 (1), pp. 97-97. | Read more

Randall KL, Lambe T, Goodnow CC, Cornall RJ. 2010. The essential role of DOCK8 in humoral immunity. Dis Markers, 29 (3-4), pp. 141-150. | Show Abstract | Read more

The processes that normally generate and maintain adaptive immunity and immunological memory are poorly understood, and yet of fundamental importance when infectious diseases place such a major economic and social burden on the world's health and agriculture systems. Defects in these mechanisms also underlie the many forms of human primary immunodeficiency. Identifying these mechanisms in a systematic way is therefore important if we are to develop better strategies for treating and preventing infection, inherited disease, transplant rejection and autoimmunity. In this review we describe a genome-wide screen in mice for the genes important for generating these adaptive responses, and describe two independent DOCK8 mutant mice strains identified by this screen. DOCK 8 was found to play an essential role in humoral immune responses and to be important in the proper formation of the B cell immunological synapse.

Johnson AL, Aravind L, Shulzhenko N, Morgun A, Choi SY, Crockford TL, Lambe T, Domaschenz H, Kucharska EM, Zheng L et al. 2010. Erratum: Themis is a member of a new metazoan gene family and is required for the completion of thymocyte positive selection (Nature Immunology (2009) 10 (831-839)) Nature Immunology, 11 (1), pp. 97. | Read more

Randall KL, Lambe T, Johnson A, Treanor B, Kucharska E, Domaschenz H, Whittle B, Tze LE, Enders A, Crockford TL et al. 2010. Corrigendum: Dock8 mutations cripple B cell immunological synapses, germinal centers and long-lived antibody production Nature Immunology, 11 (7), pp. 644-644. | Read more

Randall KL, Lambe T, Johnson AL, Treanor B, Kucharska E, Domaschenz H, Whittle B, Tze LE, Enders A, Crockford TL et al. 2009. Dock8 mutations cripple B cell immunological synapses, germinal centers and long-lived antibody production. Nat Immunol, 10 (12), pp. 1283-1291. | Show Abstract | Read more

To identify genes and mechanisms involved in humoral immunity, we did a mouse genetic screen for mutations that do not affect the first wave of antibody to immunization but disrupt response maturation and persistence. The first two mutants identified had loss-of-function mutations in the gene encoding a previously obscure member of a family of Rho-Rac GTP-exchange factors, DOCK8. DOCK8-mutant B cells were unable to form marginal zone B cells or to persist in germinal centers and undergo affinity maturation. Dock8 mutations disrupted accumulation of the integrin ligand ICAM-1 in the B cell immunological synapse but did not alter other aspects of B cell antigen receptor signaling. Humoral immunodeficiency due to Dock8 mutation provides evidence that organization of the immunological synapse is critical for signaling the survival of B cell subsets required for long-lasting immunity.

Johnson AL, Aravind L, Shulzhenko N, Morgun A, Choi S-Y, Crockford TL, Lambe T, Domaschenz H, Kucharska EM, Zheng L et al. 2009. Themis is a member of a new metazoan gene family and is required for the completion of thymocyte positive selection. Nat Immunol, 10 (8), pp. 831-839. | Show Abstract | Read more

T cell antigen receptor (TCR) signaling in CD4(+)CD8(+) double-positive thymocytes determines cell survival and lineage commitment, but the genetic and molecular basis of this process is poorly defined. To address this issue, we used ethylnitrosourea mutagenesis to identify a previously unknown T lineage-specific gene, Themis, which is critical for the completion of positive selection. Themis contains a tandem repeat of a unique globular domain (called 'CABIT' here) that includes a cysteine motif that defines a family of five uncharacterized vertebrate proteins with orthologs in most animal species. Themis-deficient thymocytes showed no substantial impairment in early TCR signaling but did show altered expression of genes involved in the cell cycle and survival before and during positive selection. Our data suggest a unique function for Themis in sustaining positive selection.

Lambe T, Simpson RJ, Dawson S, Bouriez-Jones T, Crockford TL, Lepherd M, Latunde-Dada GO, Robinson H, Raja KB, Campagna DR et al. 2009. Identification of a Steap3 endosomal targeting motif essential for normal iron metabolism. Blood, 113 (8), pp. 1805-1808. | Show Abstract | Read more

Hereditary forms of iron-deficiency anemia, including animal models, have taught us much about the normal physiologic control of iron metabolism. However, the discovery of new informative mutants is limited by the natural mutation frequency. To address this limitation, we have developed a screen for heritable abnormalities of red blood cell morphology in mice with single-nucleotide changes induced by the chemical mutagen ethylnitrosourea (ENU). We now describe the first strain, fragile-red, with hypochromic microcytic anemia resulting from a Y228H substitution in the ferrireductase Steap3 (Steap3(Y288H)). Analysis of the Steap3(Y288H) mutant identifies a conserved motif required for targeting Steap3 to internal compartments and highlights how phenotypic screens linked to mutagenesis can identify new functional variants in erythropoiesis and ascribe function to previously unidentified motifs.

Forrester JV, Xu H, Lambe T, Cornall R. 2008. Immune privilege or privileged immunity? Mucosal Immunol, 1 (5), pp. 372-381. | Show Abstract | Read more

Immune privilege is a concept that has come of age. Where previously it was considered to be a passive phenomenon restricted to certain specialized tissues, it is now viewed as comprising several mechanisms, both active and passive, shared in many aspects with emerging notions of the mechanisms of peripheral tolerance. The relative degrees of immune privilege vary from tissue to tissue depending on the number and strength of each of the mechanisms contained in that tissue. Immune privilege can be generated in non-privileged sites such as the skin and allografts, and is a property of the tissue itself. We therefore propose that, in addition to canonical central and peripheral tolerance mechanisms, there is a third route whereby the organism promotes self-antigen non-reactivity centered on the specific properties of each tissue and varying accordingly (relative degrees of immune privilege). This third mechanism of inducing immunological tolerance, as it is a local tissue phenomenon, might have particular therapeutic significance, for instance in devising strategies for induction of immunity to tumors by disrupting immune privilege or in preventing graft rejection by promoting immune privilege.

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Schallreuter KU, Bahadoran P, Picardo M, Slominski A, Elassiuty YE, Kemp EH, Giachino C, Liu JB, Luiten RM, Lambe T et al. 2008. Vitiligo pathogenesis: autoimmune disease, genetic defect, excessive reactive oxygen species, calcium imbalance, or what else? EXPERIMENTAL DERMATOLOGY, 17 (2), pp. 139-140. | Read more

Lambe T, Cornall RJ. 2008. Vitiligo pathogenesis: autoimmune disease, genetic defect, excessive reactive oxygen species, calcium imbalance, or what else? Commentary 7 EXPERIMENTAL DERMATOLOGY, 17 (2), pp. 157-158. | Read more

Schallreuter KU, Bahadoran P, Picardo M, Slominski A, Elassiuty YE, Kemp EH, Giachino C, Liu JB, Luiten RM, Lambe T et al. 2008. Vitiligo pathogenesis: autoimmune disease, genetic defect, excessive reactive oxygen species, calcium imbalance, or what else? Exp Dermatol, 17 (2), pp. 139-140. | Show Abstract | Read more

The pathobiology of vitiligo has been hotly disputed for as long as one remembers, and has been a magnet for endless speculation. Evidently, the different schools of thought--ranging, e.g. from the concept that vitiligo essentially is a free-radical disorder to that of vitiligo being a primary autoimmune disease--imply very different consequences for the best therapeutic strategies that one should adopt. As a more effective therapy for this common, often disfiguring pigmentary disorder is direly needed, we must strive harder to settle the pathogenesis debate definitively--on the basis of sound experimental evidence, rather than by a war of dogmatic theories. Recognizing, however, that it is theories which tend to guide our experimental designs and choice of study parameters, the various pathogenesis theories on the market deserve to be critically, yet unemotionally re-evaluated. This Controversies feature invites you to do so, and to ask yourself: is there something important or worthwhile exploring in other pathogenesis scenarios than those already favoured by you that may help you improve your own study design, next time you have a fresh look at vitiligo? Vitiligo provides a superb model for the study of many fundamental problems in skin biology and pathology. Therefore, even if it later turns out that, as far as your own vitiligo pathogenesis concept is concerned, you have barked-up the wrong tree most of the time, chances are that you shall anyway have generated priceless new insights into skin function along the way.

Lambe T, Cornall RJ. 2008. Commmentary 7. Exp Dermatol, 17 (2), pp. 157-158. | Read more

Silver KL, Crockford TL, Bouriez-Jones T, Milling S, Lambe T, Cornall RJ. 2007. MyD88-dependent autoimmune disease in Lyn-deficient mice. Eur J Immunol, 37 (10), pp. 2734-2743. | Show Abstract | Read more

Recent evidence suggests that systemic autoimmune disease depends on signals from TLR ligands, but little is known about how TLR-dependent pathways lead to the loss of self tolerance in vivo. To address this, we have examined the role of TLR signaling in Lyn-deficient mice, which develop an autoimmune disease similar to SLE. We found that absence of the TLR signaling adaptor molecule MyD88 suppresses plasma cell differentiation of switched and unswitched B cells, and prevents the generation of antinuclear IgG antibodies and glomerulonephritis. In mixed chimeras the increased IgM and IgG antibody secretion in Lyn-deficient mice is at least partially due to B cell-independent effects of Lyn. We now show that MyD88 deficiency blocks the expansion and activation of DC in which Lyn is also normally expressed, and prevents the hypersecretion of proinflammatory cytokines IL-6 and IL-12 by Lyn-deficient DC. These findings further highlight the important role of TLR-dependent signals in both lymphocyte activation and autoimmune pathogenesis.

Nijnik A, Woodbine L, Marchetti C, Dawson S, Lambe T, Liu C, Rodrigues NP, Crockford TL, Cabuy E, Vindigni A et al. 2007. DNA repair is limiting for haematopoietic stem cells during ageing. Nature, 447 (7145), pp. 686-690. | Show Abstract | Read more

Accumulation of DNA damage leading to adult stem cell exhaustion has been proposed to be a principal mechanism of ageing. Here we address this question by taking advantage of the highly specific role of DNA ligase IV in the repair of DNA double-strand breaks by non-homologous end-joining, and by the discovery of a unique mouse strain with a hypomorphic Lig4(Y288C) mutation. The Lig4(Y288C) mouse, identified by means of a mutagenesis screening programme, is a mouse model for human LIG4 syndrome, showing immunodeficiency and growth retardation. Diminished DNA double-strand break repair in the Lig4(Y288C) strain causes a progressive loss of haematopoietic stem cells and bone marrow cellularity during ageing, and severely impairs stem cell function in tissue culture and transplantation. The sensitivity of haematopoietic stem cells to non-homologous end-joining deficiency is therefore a key determinant of their ability to maintain themselves against physiological stress over time and to withstand culture and transplantation.

Lambe T, Leung JCH, Ferry H, Bouriez-Jones T, Makinen K, Crockford TL, Jiang HR, Nickerson JM, Peltonen L, Forrester JV, Cornall RJ. 2007. Limited peripheral T cell anergy predisposes to retinal autoimmunity. J Immunol, 178 (7), pp. 4276-4283. | Show Abstract | Read more

Autoimmune uveoretinitis accounts for at least 10% of worldwide blindness, yet it is unclear why tolerance to retinal Ags is so fragile and, particularly, to what extent this might be due to defects in peripheral tolerance. To address this issue, we generated double-transgenic mice expressing hen egg lysozyme, under the retinal interphotoreceptor retinoid-binding promoter, and a hen egg lysozyme-specific CD4(+) TCR transgene. In this manner, we have tracked autoreactive CD4(+) T cells from their development in the thymus to their involvement in uveoretinitis and compared tolerogenic mechanisms induced in a variety of organs to the same self-Ag. Our findings show that central tolerance to retinal and pancreatic Ags is qualitatively similar and equally dependent on the transcriptional regulator protein AIRE. However, the lack of Ag presentation in the eye-draining lymph nodes results in a failure to induce high levels of T cell anergy. Under these circumstances, despite considerable central deletion, low levels of retinal-specific autoreactive CD4(+) T cells can induce severe autoimmune disease. The relative lack of anergy induction by retinal Ags, in contrast to the same Ag in other organs, helps to explain the unique susceptibility of the eye to spontaneous and experimentally induced autoimmune disease.

Lambe T, Leung JCH, Bouriez-Jones T, Silver K, Makinen K, Crockford TL, Ferry H, Forrester JV, Cornall RJ. 2006. CD4 T cell-dependent autoimmunity against a melanocyte neoantigen induces spontaneous vitiligo and depends upon Fas-Fas ligand interactions. J Immunol, 177 (5), pp. 3055-3062. | Show Abstract | Read more

Better understanding of tolerance and autoimmunity toward melanocyte-specific Ags is needed to develop effective treatment for vitiligo and malignant melanoma; yet, a systematic assessment of these mechanisms has been hampered by the difficulty in tracking autoreactive T cells. To address this issue, we have generated transgenic mice that express hen egg lysozyme as a melanocyte-specific neoantigen. By crossing these animals to a hen egg lysozyme-specific CD4 TCR transgenic line we have been able to track autoreactive CD4+ T cells from their development in the thymus to their involvement in spontaneous autoimmune disease with striking similarity to human vitiligo vulgaris and Vogt-Koyanagi-Harada syndrome. Our findings show that CD4-dependent destruction of melanocytes is partially inhibited by blocking Fas-Fas ligand interactions and also highlights the importance of local control of autoimmunity, as vitiligo remains patchy and never proceeds to confluence even when Ag and autoreactive CD4+ T cells are abundant. Immune therapy to enhance or suppress melanocyte-specific T cells can be directed at a series of semiredundant pathways involving tolerance and cell death.

Nijnik A, Ferry H, Lewis G, Rapsomaniki E, Leung JCH, Daser A, Lambe T, Goodnow CC, Cornall RJ. 2006. Spontaneous B cell hyperactivity in autoimmune-prone MRL mice. Int Immunol, 18 (7), pp. 1127-1137. | Show Abstract | Read more

The MRL-lpr/lpr mouse strain is a commonly used model of the human autoimmune disease systemic lupus erythematosus (SLE). Although much is known about the contribution of the lpr Fas mutation to B cell tolerance breakdown, the role of the genetic background of the MRL strain itself is less well explored. In this study, we use the MD4 anti-hen egg lysozyme Ig (IgHEL) transgenic system to explore B cell function in MRL+/+ and non-autoimmune mice. We demonstrate that MRL IgHEL B cells show spontaneous hyperactivity in the absence of self-antigen, which is associated with low total B cell numbers but an expansion of the marginal zone B cell population. However, B cell anergy is normal in the presence of soluble lysozyme [soluble hen egg lysozyme (sHEL)], and MRL IgHEL B cells undergo normal elimination in the presence of sHEL when competing with a polyclonal C57BL/6 B cell repertoire. We conclude that B cell hyperactivity may contribute to the autoimmune phenotype of MRL+/+ and MRL-lpr/lpr strains when it initiates antibody responses to rare or sequestered antigens that are below the threshold for tolerance induction, but that there is no B cell intrinsic defect in anergy in MRL mice.

Lambe T, Finlay D, Murphy M, Martin F. 2006. Differential expression of connexin 43 in mouse mammary cells. Cell Biol Int, 30 (5), pp. 472-479. | Show Abstract | Read more

In this study we have employed suppressive subtractive hybridization (SSH) analysis to investigate differential gene expression in primary mouse mammary epithelial cells (PMMEC) cultured under mildly apoptotic/quiescent and differentiating conditions. Among a small group of genes whose expression was differentially regulated was connexin 43. In vitro, connexin 43 mRNA and protein were detectable in PMMEC cultured under proliferative or mildly apoptotic conditions. The level of connexin 43 mRNA expression in vivo was also investigated. High levels of expression were found to be associated with the periods of greatest glandular plasticity (pubertal expansion of the mammary tree, early pregnancy and during early involution). Thus, terminally differentiated cells in vivo and in vitro did not express connexin 43 mRNA suggesting that connexin 43 expression, and perhaps facilitated gap junction communication, is associated with undifferentiated progenitor cell populations.

Howlin J, McBryan J, Napoletano S, Lambe T, McArdle E, Shioda T, Martin F. 2006. CITED1 homozygous null mice display aberrant pubertal mammary ductal morphogenesis. Oncogene, 25 (10), pp. 1532-1542. | Show Abstract | Read more

Expression microarray analysis identified CITED1 among a group of genes specifically upregulated in the pubertal mouse mammary gland. At puberty, CITED1 localizes to the luminal epithelial cell population of the mammary ducts and the body cells of the terminal end buds. Generation of CITED1 gene knockout mice showed that homozygous null mutants exhibit retarded mammary ductal growth at puberty and, in addition, dilated ductal structures with a lack of spatial restriction of the subtending branches. Analysis of CITED1 homozygous null and heterozygous null mammary gland gene expression using microarrays suggested that the mammary-specific phenotype seen in the homozygous null females is due to a disturbance in the transcription of a number of key mediators of pubertal ductal morphogenesis. These include estrogen and TGFbeta responsive genes, such as the EGFR/ErbB2 ligand, amphiregulin, whose transcription we suggest is directly or indirectly regulated by CITED1.

Ferry H, Leung JCH, Lewis G, Nijnik A, Silver K, Lambe T, Cornall RJ. 2006. B-cell tolerance. Transplantation, 81 (3), pp. 308-315. | Show Abstract | Read more

Autoreactive B cells are actively tolerized to more abundant self-antigens by a series of checkpoints involving receptor editing, deletion, anergy and competition for growth factors. In contrast, B cells reactive against rare, sequestered or tissue specific self-antigens remain functionally naïve. During an immune response, the autoimmune danger from these cells is countered by a variety of mechanisms comprising control of self-antigen presentation, limitation of immunogenic and tolerogenic costimuli including T cell help, homeostatic control of growth and strict regulation of germinal centre reactions. In this overview we consider how knowledge of these checkpoints may be used to gain a better understanding of transplant tolerance and the generation of alloantibodies.

Vinuesa CG, Cook MC, Angelucci C, Athanasopoulos V, Rui L, Hill KM, Yu D, Domaschenz H, Whittle B, Lambe T et al. 2005. A RING-type ubiquitin ligase family member required to repress follicular helper T cells and autoimmunity. Nature, 435 (7041), pp. 452-458. | Show Abstract | Read more

Despite the sequencing of the human and mouse genomes, few genetic mechanisms for protecting against autoimmune disease are currently known. Here we systematically screen the mouse genome for autoimmune regulators to isolate a mouse strain, sanroque, with severe autoimmune disease resulting from a single recessive defect in a previously unknown mechanism for repressing antibody responses to self. The sanroque mutation acts within mature T cells to cause formation of excessive numbers of follicular helper T cells and germinal centres. The mutation disrupts a repressor of ICOS, an essential co-stimulatory receptor for follicular T cells, and results in excessive production of the cytokine interleukin-21. sanroque mice fail to repress diabetes-causing T cells, and develop high titres of autoantibodies and a pattern of pathology consistent with lupus. The causative mutation is in a gene of previously unknown function, roquin (Rc3h1), which encodes a highly conserved member of the RING-type ubiquitin ligase protein family. The Roquin protein is distinguished by the presence of a CCCH zinc-finger found in RNA-binding proteins, and localization to cytosolic RNA granules implicated in regulating messenger RNA translation and stability.

Black AP, Wee E, Lambe T, Bailey A, Jones L, Ogg GS. 2005. Combined oligonucleotide and protein microarray temporal analysis of p53-mediated apoptosis Cancer Genomics and Proteomics, 2 (2), pp. 61-69. | Show Abstract

Background: P53-mediated apoptosis involves a complex process induced largely by p53 acting as a transcription factor. We hypothesised that p53 expression would lead to transcriptional events that rapidly change during apoptosis and that protein array analysis would give a more comprehensive picture of p53-mediated apoptosis than mRNA alone. Materials and Methods: We over-expressed p53 in lymphoblastoid cell lines and performed temporal analysis of functional apoptosis, assessing mRNA levels by oligo microarray and protein levels by novel antibody microarray and Western blot. Results: mRNA levels varied over time. At least 10 genes that showed enhanced expression, such as APAF-1 and GADD45, contained the p53 target sequence confirming their nature as primary p53 targets. Changes in mRNA expression did not correlate directly with changes in protein expression. Conclusion: Array analysis of protein expression in addition to mRNA expression gave a more complete assessment of p53-mediated apoptosis; for example, enhanced levels of bcl-2 and E2F2 protein were detected which, along with APAF-1, are involved in the mitochondrial apoptotic process. These data extend existing p53 array findings to correlate with protein microarray data and functional apoptosis. Furthermore they emphasize the importance of combined proteomic and genomic approaches to the investigation of p53-mediated apoptosis and other cellular processes.

Lewis G, Rapsomaniki E, Bouriez T, Crockford T, Ferry H, Rigby R, Vyse T, Lambe T, Cornall R. 2004. Hyper IgE in New Zealand black mice due to a dominant-negative CD23 mutation. Immunogenetics, 56 (8), pp. 564-571. | Show Abstract | Read more

Immunoglobulin E (IgE) plays a critical role in both resistance to parasitic infection and allergy to environmental antigens. The IgE response is in turn regulated by the B-cell co-receptor CD23, and CD23-deficient mice show exaggerated IgE responses and airway hyper-responsiveness. In this report, we show that New Zealand black (NZB) mice express a variant CD23 allele, with mutations in both the C-lectin-binding domain and stalk region, which fails to bind IgE at high affinity and has reduced expression on the cell surface. Expression of the variant CD23 chain interferes with trimerisation of the receptor and has a dominant-negative effect leading to reduced IgE binding in crosses between NZB and other strains. Genetic mapping shows that the variant CD23 leads to an exaggerated primary IgE response, which is independent of other strain-specific effects. These results suggest that NZB mice or mice carrying the variant allele will be useful models for studying both allergy and quantitative traits associated with atopy. The exaggerated IgE response provides an explanation for the natural resistance of NZB mice to parasitic infection by Leishmania.

Clarkson MR, Murphy M, Gupta S, Lambe T, Mackenzie HS, Godson C, Martin F, Brady HR. 2002. High glucose-altered gene expression in mesangial cells. Actin-regulatory protein gene expression is triggered by oxidative stress and cytoskeletal disassembly (vol 277, pg 9707, 2002) JOURNAL OF BIOLOGICAL CHEMISTRY, 277 (25), pp. 23100-23102.

Clarkson MR, Murphy M, Gupta S, Lambe T, Mackenzie HS, Godson C, Martin F, Brady HR. 2002. Erratum: High glucose-altered gene expression in mesangial cells. Actin-regulatory protein gene expression is triggered by oxidative stress and cytoskeletal disassembly (The Journal of Biological Chemistry (2002) 277 (9707-9712)) Journal of Biological Chemistry, 277 (25), pp. 23100-23102.

Clarkson MR, Murphy M, Gupta S, Lambe T, Mackenzie HS, Godson C, Martin F, Brady HR. 2002. High glucose-altered gene expression in mesangial cells. Actin-regulatory protein gene expression is triggered by oxidative stress and cytoskeletal disassembly. J Biol Chem, 277 (12), pp. 9707-9712. | Show Abstract | Read more

High extracellular glucose plays a pivotal role in the pathophysiology of diabetic nephropathy. Here we report 200 genes, identified using suppression-subtractive hybridization, that are differentially expressed when human mesangial cells are propagated in high ambient glucose in vitro. The major functional classes of genes identified included modulators and products of extracellular matrix protein metabolism, regulators of cell growth and turnover, and a cohort of actin cytoskeleton regulatory proteins. Actin cytoskeletal disassembly is a prominent feature of diabetic nephropathy. The induction of actin cytoskeleton regulatory gene expression by high glucose was attenuated by the inhibitor of reactive oxygen species generation, carbonyl cyanide m-chlorophenylhydrazone but not by the protein kinase C inhibitor GF 109203X and was not mimicked by the addition of transforming growth factor beta. Enhanced expression of actin cytoskeleton regulatory genes was also observed following disruption of the mesangial cell actin cytoskeleton by cytochalasin D. In aggregate, these results suggest that the induction of genes encoding actin cytoskeleton regulatory proteins (a) is a prominent component of the mesangial cell transcriptomic response in diabetic nephropathy and (b) is dependent on oxidative stress, is independent of protein kinase C and transforming growth factor-beta, and represents an adaptive response to actin cytoskeleton disassembly.

Tully CM, Chinnakannan S, Mullarkey CE, Ulaszewska M, Ferrara F, Temperton N, Gilbert SC, Lambe T. 2017. Novel Bivalent Viral-Vectored Vaccines Induce Potent Humoral and Cellular Immune Responses Conferring Protection against Stringent Influenza A Virus Challenge. J Immunol, 199 (4), pp. 1333-1341. | Show Abstract | Read more

Seasonal influenza viruses are a common cause of acute respiratory illness worldwide and generate a significant socioeconomic burden. Influenza viruses mutate rapidly, necessitating annual vaccine reformulation because traditional vaccines do not typically induce broad-spectrum immunity. In addition to seasonal infections, emerging pandemic influenza viruses present a continued threat to global public health. Pandemic influenza viruses have consistently higher attack rates and are typically associated with greater mortality compared with seasonal strains. Ongoing strategies to improve vaccine efficacy typically focus on providing broad-spectrum immunity; although B and T cells can mediate heterosubtypic responses, typical vaccine development will augment either humoral or cellular immunity. However, multipronged approaches that target several Ags may limit the generation of viral escape mutants. There are few vaccine platforms that can deliver multiple Ags and generate robust cellular and humoral immunity. In this article, we describe a novel vaccination strategy, tested preclinically in mice, for the delivery of novel bivalent viral-vectored vaccines. We show this strategy elicits potent T cell responses toward highly conserved internal Ags while simultaneously inducing high levels of Abs toward hemagglutinin. Importantly, these humoral responses generate long-lived plasma cells and generate Abs capable of neutralizing variant hemagglutinin-expressing pseudotyped lentiviruses. Significantly, these novel viral-vectored vaccines induce strong immune responses capable of conferring protection in a stringent influenza A virus challenge. Thus, this vaccination regimen induces lasting efficacy toward influenza. Importantly, the simultaneous delivery of dual Ags may alleviate the selective pressure that is thought to potentiate antigenic diversity in avian influenza viruses.

Alharbi NK, Padron-Regalado E, Thompson CP, Kupke A, Wells D, Sloan MA, Grehan K, Temperton N, Lambe T, Warimwe G et al. 2017. ChAdOx1 and MVA based vaccine candidates against MERS-CoV elicit neutralising antibodies and cellular immune responses in mice. Vaccine, 35 (30), pp. 3780-3788. | Show Abstract | Read more

The Middle East respiratory syndrome coronavirus (MERS-CoV) has infected more than 1900 humans, since 2012. The syndrome ranges from asymptomatic and mild cases to severe pneumonia and death. The virus is believed to be circulating in dromedary camels without notable symptoms since the 1980s. Therefore, dromedary camels are considered the only animal source of infection. Neither antiviral drugs nor vaccines are approved for veterinary or medical use despite active research on this area. Here, we developed four vaccine candidates against MERS-CoV based on ChAdOx1 and MVA viral vectors, two candidates per vector. All vaccines contained the full-length spike gene of MERS-CoV; ChAdOx1 MERS vaccines were produced with or without the leader sequence of the human tissue plasminogen activator gene (tPA) where MVA MERS vaccines were produced with tPA, but either the mH5 or F11 promoter driving expression of the spike gene. All vaccine candidates were evaluated in a mouse model in prime only or prime-boost regimens. ChAdOx1 MERS with tPA induced higher neutralising antibodies than ChAdOx1 MERS without tPA. A single dose of ChAdOx1 MERS with tPA elicited cellular immune responses as well as neutralising antibodies that were boosted to a significantly higher level by MVA MERS. The humoral immunogenicity of a single dose of ChAdOx1 MERS with tPA was equivalent to two doses of MVA MERS (also with tPA). MVA MERS with mH5 or F11 promoter induced similar antibody levels; however, F11 promoter enhanced the cellular immunogenicity of MVA MERS to significantly higher magnitudes. In conclusion, our study showed that MERS-CoV vaccine candidates could be optimized by utilising different viral vectors, various genetic designs of the vectors, or different regimens to increase immunogenicity. ChAdOx1 and MVA vectored vaccines have been safely evaluated in camels and humans and these MERS vaccine candidates should now be tested in camels and in clinical trials.

Lambe T, Bowyer G, Ewer KJ. 2017. A review of Phase I trials of Ebola virus vaccines: what can we learn from the race to develop novel vaccines? Philos Trans R Soc Lond B Biol Sci, 372 (1721), pp. 20160295-20160295. | Show Abstract | Read more

Sporadic outbreaks of Ebola virus infection have been documented since the mid-Seventies and viral exposure can lead to lethal haemorrhagic fever with case fatalities as high as 90%. There is now a comprehensive body of data from both ongoing and completed clinical trials assessing various vaccine strategies, which were rapidly advanced through clinical trials in response to the 2013-2016 Ebola virus disease (EVD) public health emergency. Careful consideration of immunogenicity post vaccination is essential but has been somewhat stifled because of the wide array of immunological assays and outputs that have been used in the numerous clinical trials. We discuss here the different aspects of the immune assays currently used in the Phase I clinical trials for Ebola virus vaccines, and draw comparisons across the immune outputs where possible; various trials have examined both cellular and humoral immunity in European and African cohorts. Assessment of the safety data, the immunological outputs and the ease of field deployment for the various vaccine modalities will help both the scientific community and policy-makers prioritize and potentially license vaccine candidates. If this can be achieved, the next outbreak of Ebola virus, or other emerging pathogen, can be more readily contained and will not have such widespread and devastating consequences.This article is part of the themed issue 'The 2013-2016 West African Ebola epidemic: data, decision-making and disease control'.

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.

Ewer K, Rampling T, Venkatraman N, Bowyer G, Wright D, Lambe T, Imoukhuede EB, Payne R, Fehling SK, Strecker T et al. 2016. A Monovalent Chimpanzee Adenovirus Ebola Vaccine Boosted with MVA. N Engl J Med, 374 (17), pp. 1635-1646. | Show Abstract | Read more

BACKGROUND: The West African outbreak of Ebola virus disease that peaked in 2014 has caused more than 11,000 deaths. The development of an effective Ebola vaccine is a priority for control of a future outbreak. METHODS: In this phase 1 study, we administered a single dose of the chimpanzee adenovirus 3 (ChAd3) vaccine encoding the surface glycoprotein of Zaire ebolavirus (ZEBOV) to 60 healthy adult volunteers in Oxford, United Kingdom. The vaccine was administered in three dose levels--1×10(10) viral particles, 2.5×10(10) viral particles, and 5×10(10) viral particles--with 20 participants in each group. We then assessed the effect of adding a booster dose of a modified vaccinia Ankara (MVA) strain, encoding the same Ebola virus glycoprotein, in 30 of the 60 participants and evaluated a reduced prime-boost interval in another 16 participants. We also compared antibody responses to inactivated whole Ebola virus virions and neutralizing antibody activity with those observed in phase 1 studies of a recombinant vesicular stomatitis virus-based vaccine expressing a ZEBOV glycoprotein (rVSV-ZEBOV) to determine relative potency and assess durability. RESULTS: No safety concerns were identified at any of the dose levels studied. Four weeks after immunization with the ChAd3 vaccine, ZEBOV-specific antibody responses were similar to those induced by rVSV-ZEBOV vaccination, with a geometric mean titer of 752 and 921, respectively. ZEBOV neutralization activity was also similar with the two vaccines (geometric mean titer, 14.9 and 22.2, respectively). Boosting with the MVA vector increased virus-specific antibodies by a factor of 12 (geometric mean titer, 9007) and increased glycoprotein-specific CD8+ T cells by a factor of 5. Significant increases in neutralizing antibodies were seen after boosting in all 30 participants (geometric mean titer, 139; P<0.001). Virus-specific antibody responses in participants primed with ChAd3 remained positive 6 months after vaccination (geometric mean titer, 758) but were significantly higher in those who had received the MVA booster (geometric mean titer, 1750; P<0.001). CONCLUSIONS: The ChAd3 vaccine boosted with MVA elicited B-cell and T-cell immune responses to ZEBOV that were superior to those induced by the ChAd3 vaccine alone. (Funded by the Wellcome Trust and others; ClinicalTrials.gov number, NCT02240875.).

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Antrobus RD, Coughlan L, Berthoud TK, Dicks MD, Hill AVS, Lambe T, Gilbert SC. 2014. Clinical assessment of a novel recombinant simian adenovirus ChAdOx1 as a vectored vaccine expressing conserved influenza a antigens Molecular Therapy, 22 (3), pp. 668-674. | Show Abstract | Read more

Adenoviruses are potent vectors for inducing and boosting cellular immunity to encoded recombinant antigens. However, the widespread seroprevalence of neutralizing antibodies to common human adenovirus serotypes limits their use. Simian adenoviruses do not suffer from the same drawbacks. We have constructed a replication-deficient chimpanzee adenovirus-vectored vaccine expressing the conserved influenza antigens, nucleoprotein (NP), and matrix protein 1 (M1). Here, we report safety and T-cell immunogenicity following vaccination with this novel recombinant simian adenovirus, ChAdOx1 NP+M1, in a first in human dose-escalation study using a 3+3 study design, followed by boosting with modified vaccinia virus Ankara expressing the same antigens in some volunteers. We demonstrate ChAdOx1 NP+M1 to be safe and immunogenic. ChAdOx1 is a promising vaccine vector that could be used to deliver vaccine antigens where strong cellular immune responses are required for protection. © The American Society of Gene & Cell Therapy.

Mullarkey CE, Boyd A, van Laarhoven A, Lefevre EA, Veronica Carr B, Baratelli M, Molesti E, Temperton NJ, Butter C, Charleston B et al. 2013. Improved adjuvanting of seasonal influenza vaccines: preclinical studies of MVA-NP+M1 coadministration with inactivated influenza vaccine. Eur J Immunol, 43 (7), pp. 1940-1952. | Show Abstract | Read more

Licensed seasonal influenza vaccines induce antibody (Ab) responses against influenza hemagglutinin (HA) that are limited in their ability to protect against different strains of influenza. Cytotoxic T lymphocytes recognizing the conserved internal nucleoprotein (NP) and matrix protein (M1) are capable of mediating a cross-subtype immune response against influenza. Modified vaccinia Ankara (MVA) virus encoding NP and M1 (MVA-NP+M1) is designed to boost preexisting T-cell responses in adults in order to elicit a cross-protective immune response. We examined the coadministration of HA protein formulations and candidate MVA-NP+M1 influenza vaccines in murine, avian, and swine models. Ab responses postimmunization were measured by ELISA and pseudotype neutralization assays. Here, we demonstrate that MVA-NP+M1 can act as an adjuvant enhancing Ab responses to HA while simultaneously inducing potent T-cell responses to conserved internal Ags. We show that this regimen leads to the induction of cytophilic Ab isotypes that are capable of inhibiting hemagglutination and in the context of H5 exhibit cross-clade neutralization. The simultaneous induction of T cells and Ab responses has the potential to improve seasonal vaccine performance and could be employed in pandemic situations.

Lambe T. 2012. Novel viral vectored vaccines for the prevention of influenza. Mol Med, 18 (8), pp. 1153-1160. | Show Abstract | Read more

Influenza represents a substantial global healthcare burden, with annual epidemics resulting in 3-5 million cases of severe illness with a significant associated mortality. In addition, the risk of a virulent and lethal influenza pandemic has generated widespread and warranted concern. Currently licensed influenza vaccines are limited in their ability to induce efficacious and long-lasting herd immunity. In addition, and as evidenced by the H1N1 pandemic in 2009, there can be a significant delay between the emergence of a pandemic influenza and an effective, antibody-inducing vaccine. There is, therefore, a continued need for new, efficacious vaccines conferring cross-clade protection-obviating the need for biannual reformulation of seasonal influenza vaccines. Development of such a vaccine would yield enormous health benefits to society and also greatly reduce the associated global healthcare burden. There are a number of alternative influenza vaccine technologies being assessed both preclinically and clinically. In this review we discuss viral vectored vaccines, either recombinant live-attenuated or replication-deficient viruses, which are current lead candidates for inducing efficacious and long-lasting immunity toward influenza viruses. These alternate influenza vaccines offer real promise to deliver viable alternatives to currently deployed vaccines and more importantly may confer long-lasting and universal protection against influenza viral infection.

Lambe T, Spencer AJ, Mullarkey CE, Antrobus RD, Yu L-M, de Whalley P, Thompson BAV, Jones C, Chalk J, Kerridge S et al. 2012. T-cell responses in children to internal influenza antigens, 1 year after immunization with pandemic H1N1 influenza vaccine, and response to revaccination with seasonal trivalent-inactivated influenza vaccine. Pediatr Infect Dis J, 31 (6), pp. e86-e91. | Show Abstract | Read more

BACKGROUND: During seasonal influenza epidemics, 5-15% of the population are affected with an illness having a nontrivial mortality, morbidity and economic burden. Inactivated influenza vaccines are routinely used to prevent influenza infection, primarily by inducing humoral immunity. In addition, trivalent-inactivated influenza vaccines have previously been shown to boost influenza-specific T-cell responses in a small percentage of adults. We investigate here the influenza-specific T-cell response, in children, 1 year after pandemic H1N1 vaccination and the ability to boost the T-cell response with trivalent-inactivated influenza immunization. METHODS: Peripheral blood mononuclear cells (PBMCs) were isolated from children previously vaccinated with pandemic H1N1 vaccine, pre- and postseasonal 2010-2011 trivalent influenza vaccine (TIV) vaccination. Samples were analyzed by interferon-gamma enzyme-linked immunosorbent spot for reactogenicity toward internal influenza antigens (nucleoprotein, matrix protein 1 and nonstructural protein 1). RESULTS: Basal ex vivo T-cell responses to nucleoprotein, matrix protein 1 and nonstructural protein 1 measured by interferon-gamma enzyme-linked immunosorbent spot assay were significantly higher in those children who had previously received an AS03B-adjuvanted split virion pandemic vaccine 12 months earlier rather than a nonadjuvanted whole virion vaccine. Boosting of these responses, 21 days after 2010/2011 seasonal TIV vaccination was observed regardless of age or prior pandemic vaccination regime, although boosting was greater in those groups with the lowest initial response. CONCLUSIONS: We show here that children previously vaccinated with the 2009 pandemic H1N1 vaccine have measurable T-cell responses 1 year after vaccination. The magnitudes of these responses are dependent on both age of vaccine and type of pandemic H1N1 vaccine used. After 2010/2011 seasonal TIV vaccination, these T-cell responses undergo a small but significant boost.

Lillie PJ, Berthoud TK, Powell TJ, Lambe T, Mullarkey C, Spencer AJ, Hamill M, Peng Y, Blais M-E, Duncan CJA et al. 2012. Preliminary assessment of the efficacy of a T-cell-based influenza vaccine, MVA-NP+M1, in humans. Clin Infect Dis, 55 (1), pp. 19-25. | Show Abstract | Read more

BACKGROUND: The novel influenza vaccine MVA-NP+M1 is designed to boost cross-reactive T-cell responses to internal antigens of the influenza A virus that are conserved across all subtypes, providing protection against both influenza disease and virus shedding against all influenza A viruses. Following a phase 1 clinical study that demonstrated vaccine safety and immunogenicity, a phase 2a vaccination and influenza challenge study has been conducted in healthy adult volunteers. METHODS: Volunteers with no measurable serum antibodies to influenza A/Wisconsin/67/2005 received either a single vaccination with MVA-NP+M1 or no vaccination. T-cell responses to the vaccine antigens were measured at enrollment and again prior to virus challenge. All volunteers underwent intranasal administration of influenza A/Wisconsin/67/2005 while in a quarantine unit and were monitored for symptoms of influenza disease and virus shedding. RESULTS: Volunteers had a significantly increased T-cell response to the vaccine antigens following a single dose of the vaccine, with an increase in cytolytic effector molecules. Intranasal influenza challenge was undertaken without safety issues. Two of 11 vaccinees and 5 of 11 control subjects developed laboratory-confirmed influenza (symptoms plus virus shedding). Symptoms of influenza were less pronounced in the vaccinees and there was a significant reduction in the number of days of virus shedding in those vaccinees who developed influenza (mean, 1.09 days in controls, 0.45 days in vaccinees, P = .036). CONCLUSIONS: This study provides the first demonstration of clinical efficacy of a T-cell-based influenza vaccine and indicates that further clinical development should be undertaken. CLINICAL TRIALS REGISTRATION: NCT00993083.

Berthoud TK, Hamill M, Lillie PJ, Hwenda L, Collins KA, Ewer KJ, Milicic A, Poyntz HC, Lambe T, Fletcher HA et al. 2011. Potent CD8+ T-cell immunogenicity in humans of a novel heterosubtypic influenza A vaccine, MVA-NP+M1. Clin Infect Dis, 52 (1), pp. 1-7. | Show Abstract | Read more

BACKGROUND: Influenza A viruses cause occasional pandemics and frequent epidemics. Licensed influenza vaccines that induce high antibody titers to the highly polymorphic viral surface antigen hemagglutinin must be re-formulated and readministered annually. A vaccine providing protective immunity to the highly conserved internal antigens could provide longer-lasting protection against multiple influenza subtypes. METHODS: We prepared a Modified Vaccinia virus Ankara (MVA) vector encoding nucleoprotein and matrix protein 1 (MVA-NP+M1) and conducted a phase I clinical trial in healthy adults. RESULTS: The vaccine was generally safe and well tolerated, with significantly fewer local side effects after intramuscular rather than intradermal administration. Systemic side effects increased at the higher dose in both frequency and severity, with 5 out of 8 volunteers experiencing severe nausea/vomiting, malaise, or rigors. Ex vivo T-cell responses to NP and M1 measured by IFN-γ ELISPOT assay were significantly increased after vaccination (prevaccination median of 123 spot-forming units/million peripheral blood mononuclear cells, postvaccination peak response median 339, 443, and 1443 in low-dose intradermal, low-dose intramuscular, and high-dose intramuscular groups, respectively), and the majority of the antigen-specific T cells were CD8(+). CONCLUSIONS: We conclude that the vaccine was both safe and remarkably immunogenic, leading to frequencies of responding T cells that appear to be much higher than those induced by any other influenza vaccination approach. Further studies will be required to find the optimum dose and to assess whether the increased T-cell response to conserved influenza proteins results in protection from influenza disease.

Lambe T, Crawford G, Johnson AL, Crockford TL, Bouriez-Jones T, Smyth AM, Pham THM, Zhang Q, Freeman AF, Cyster JG et al. 2011. DOCK8 is essential for T-cell survival and the maintenance of CD8+ T-cell memory. Eur J Immunol, 41 (12), pp. 3423-3435. | Show Abstract | Read more

Deficiency in the guanine nucleotide exchange factor dedicator of cytokinesis 8 (DOCK8) causes a human immunodeficiency syndrome associated with recurrent sinopulmonary and viral infections. We have recently identified a DOCK8-deficient mouse strain, carrying an ethylnitrosourea-induced splice-site mutation that shows a failure to mature a humoral immune response due to the loss of germinal centre B cells. In this study, we turned to T-cell immunity to investigate further the human immunodeficiency syndrome and its association with decreased peripheral CD4(+) and CD8(+) T cells. Characterisation of the DOCK8-deficient mouse revealed T-cell lymphopenia, with increased T-cell turnover and decreased survival. Egress of mature CD4(+) thymocytes was reduced with increased migration of these cells to the chemokine CXCL12. However, despite the two-fold reduction in peripheral naïve T cells, the DOCK8-deficient mice generated a normal primary CD8(+) immune response and were able to survive acute influenza virus infection. The limiting effect of DOCK8 was in the normal survival of CD8(+) memory T cells after infection. These findings help to explain why DOCK8-deficient patients are susceptible to recurrent infections and provide new insights into how T-cell memory is sustained.

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