Demonstration of the Blood-Stage Plasmodium falciparum Controlled Human Malaria Infection Model to Assess Efficacy of the P. falciparum Apical Membrane Antigen 1 Vaccine, FMP2.1/AS01

We study whether the relationship between the state unemployment rate at the time of conception and infant health, infant mortality and maternal characteristics in the United States has changed over the years 1980-2004. We use microdata on births and deaths for years 1980-2004 and find that the relationship between the state unemployment rate at the time of conception and infant mortality and birthweight changes over time and is stronger for blacks than whites. For years 1980-1989 increases in the state unemployment rate are associated with a decline in infant mortality among blacks, an effect driven by mortality from gestational development and birth weight, and complications of placenta while in utero. In contrast, state economic conditions are unrelated to black infant mortality in years 1990-2004 and white infant mortality in any period, although effects vary by cause of death. We explore potential mechanisms for our findings and, including mothers younger than 18 in the analysis, uncover evidence of age-related maternal selection in response to the business cycle. In particular, in years 1980-1989 an increase in the unemployment rate at the time of conception is associated with fewer babies born to young mothers. The magnitude and direction of the relationship between business cycles and infant mortality differs by race and period. Age-related selection into motherhood in response to the business cycle is a possible explanation for this changing relationship

Assessment of novel pre-erythrocytic malaria vaccines in humans

Falciparum malaria remains one of the leading infectious causes of morbidity and mortality worldwide. Though the search for an effective vaccine has seen unprecedented advances in recent years and one of the leading vaccine candidates, RTS,S/AS01 is going to enter pilot deployments in Africa in 2018, there has been no vaccine that has demonstrated durable high level efficacy. This thesis describes four Phase I and Phase II clinical trials assessing novel pre-erythrocytic malaria vaccines in healthy adult UK volunteers. R21 has been developed at the Jenner Institute, University of Oxford. This is an improved RTS,S construct, an antigen derived from the pre-erythrocytic circumsporozoite protein, which is an abundant coat protein involved in sporozoite development and hepatocyte invasion. R21 comprises recombinant particles expressing the central repeat and the C-terminus of the circumsporozoite protein (CSP) fused to the Hepatitis B surface antigen (HBsAg), but without the excess of unfused HBsAg protein found in RTS,S. Both RTS,S and R21 predominantly function by inducing antibodies against the circumsporozoite protein. In addition to humoral immunity, it is widely recognised that T-cell mechanisms are likely to play an important role in malaria immunity and heterologous prime-boost regimes using viral-vectored vaccines have been shown to induce potent T-cell responses. ChAd63-MVA ME-TRAP is a regime that has been developed in the Jenner Institute and the first of the clinical trials described in Chapter 3 assesses the safety and immunogenicity of adjuvanting this regime with a novel saponin-based adjuvant, Matrix-MTM. This trial showed that this regime was safe, well tolerated and did not result in any immunological interference. R21 had never been administered to humans prior to the work described in this thesis. In Chapter 4, I describe two Phase I clinical trials assessing the safety and immunogenicity of R21 given at varying doses administered with two different adjuvants, Matrix-M (MM) and AS01B. Both these trials showed that R21 was safe and well tolerated when administered with either adjuvant and elicited comparable humoral immune responses to RTS,S/AS01B even at one-fifth of the dose (10&mu;g). This formed the basis to test the efficacy of R21/MM, which had a favourable reactogenicity profile, using controlled human malaria infection in malaria-naïve UK volunteers. High level efficacy (&GT; 80%) is reported in Chapter 5 with a three-dose schedule of 10/10/10&mu;g R21/MM given 4 weeks apart. This provides the first evidence of proof-of-concept that this vaccine regime should be tested in malaria-endemic populations.</p

Assessment of novel pre-erythrocytic malaria vaccines in humans

Falciparum malaria remains one of the leading infectious causes of morbidity and mortality worldwide. Though the search for an effective vaccine has seen unprecedented advances in recent years and one of the leading vaccine candidates, RTS,S/AS01 is going to enter pilot deployments in Africa in 2018, there has been no vaccine that has demonstrated durable high level efficacy. This thesis describes four Phase I and Phase II clinical trials assessing novel pre-erythrocytic malaria vaccines in healthy adult UK volunteers. R21 has been developed at the Jenner Institute, University of Oxford. This is an improved RTS,S construct, an antigen derived from the pre-erythrocytic circumsporozoite protein, which is an abundant coat protein involved in sporozoite development and hepatocyte invasion. R21 comprises recombinant particles expressing the central repeat and the C-terminus of the circumsporozoite protein (CSP) fused to the Hepatitis B surface antigen (HBsAg), but without the excess of unfused HBsAg protein found in RTS,S. Both RTS,S and R21 predominantly function by inducing antibodies against the circumsporozoite protein. In addition to humoral immunity, it is widely recognised that T-cell mechanisms are likely to play an important role in malaria immunity and heterologous prime-boost regimes using viral-vectored vaccines have been shown to induce potent T-cell responses. ChAd63-MVA ME-TRAP is a regime that has been developed in the Jenner Institute and the first of the clinical trials described in Chapter 3 assesses the safety and immunogenicity of adjuvanting this regime with a novel saponin-based adjuvant, Matrix-MTM. This trial showed that this regime was safe, well tolerated and did not result in any immunological interference. R21 had never been administered to humans prior to the work described in this thesis. In Chapter 4, I describe two Phase I clinical trials assessing the safety and immunogenicity of R21 given at varying doses administered with two different adjuvants, Matrix-M (MM) and AS01B. Both these trials showed that R21 was safe and well tolerated when administered with either adjuvant and elicited comparable humoral immune responses to RTS,S/AS01B even at one-fifth of the dose (10μg). This formed the basis to test the efficacy of R21/MM, which had a favourable reactogenicity profile, using controlled human malaria infection in malaria-naïve UK volunteers. High level efficacy (&amp;GT; 80%) is reported in Chapter 5 with a three-dose schedule of 10/10/10μg R21/MM given 4 weeks apart. This provides the first evidence of proof-of-concept that this vaccine regime should be tested in malaria-endemic populations.</p

CXCR3+ T Follicular Helper Cells Induced by Co-Administration of RTS,S/AS01B and Viral-Vectored Vaccines Are Associated With Reduced Immunogenicity and Efficacy Against Malaria

A malaria vaccine strategy targeting multiple lifecycle stages may be required to achieve a high level of efficacy. In two Phase IIa clinical trials, we tested immunogenicity and efficacy of RTS,S/AS01B administered alone, in a staggered regimen with viral-vectored vaccines or co-administered with viral-vectored vaccines. RTS,S/AS01B induces high titers of antibody against sporozoites and viral-vectored vaccines ChAd63 ME-TRAP and MVA ME-TRAP induce potent T cell responses against infected hepatocytes. By combining these two strategies, we aimed to improve efficacy by inducing immune responses targeting multiple parasite antigens. Vaccination with RTS,S/AS01B alone or in a staggered regimen with viral vectors produced strong immune responses and demonstrated high levels of protection against controlled human malaria infection. However, concomitant administration of these vaccines significantly reduced humoral immunogenicity and protective efficacy. Strong Th1-biased cytokine responses induced by MVA ME-TRAP were associated with a skew in circulating T follicular helper cells toward a CXCR3+ phenotype and a reduction in antibody quantity and quality. This study illustrates that while a multistage-targeting vaccine strategy could provide high-level efficacy, the regimen design will require careful optimization

High levels of neurological involvement but low mortality in miliary tuberculosis: a six-year case-series from the United Kingdom

Tuberculosis (TB) remains one of the biggest global health challenges. Whilst the greatest burden of active disease is seen in Asia and Africa [1], TB remains a significant issue in the UK. Miliary TB is one of the severest manifestations of TB disease [2–4]. Up-to-date clinicopathological data on miliary TB from the developed world are lacking. We undertook a comprehensive 6-year review (2007–2012) of cases presenting to a single UK centre with an ethnically diverse population with high levels of population exchange with the Indian Subcontinent and Africa. Miliary TB was defined as the presence of miliary nodules on thoracic imaging in patients who presented with symptoms compatible with the diagnosis and either culture of Mycobacterium tuberculosis complex or culture-negative patients with clinical and/or histological features compatible with TB who were commenced on a course of antituberculous therapy (ATT)

Genotyping techniques to address diversity in tumors.

Array-based genotyping platforms have during recent years been established as a valuable tool for the characterization of genomic alterations in cancer. The analysis of tumor samples, however, presents challenges for data analysis and interpretation. For example, tumor samples are often admixed with nonaberrant cells that define the tumor microenvironment, such as infiltrating lymphocytes and fibroblasts, or vasculature. Furthermore, tumors often comprise subclones harboring divergent aberrations that are acquired subsequent to the tumor-initiating event. The combined analysis of both genotype and copy number status obtained by array-based genotyping platforms provide opportunities to address these challenges. In this chapter, we present the basic principles for current array-based genotyping platforms and how they can be used to infer genotype and copy number for acquired genomic alterations. We describe how these techniques can be used to resolve tumor ploidy, normal cell admixture, and subclonality. We also exemplify how genotyping techniques can be applied in tumor studies to elucidate the hierarchy among tumor clones, and thus, provide means to study clonal expansion and tumor evolution

Image_1_CXCR3+ T Follicular Helper Cells Induced by Co-Administration of RTS,S/AS01B and Viral-Vectored Vaccines Are Associated With Reduced Immunogenicity and Efficacy Against Malaria.JPEG

<p>A malaria vaccine strategy targeting multiple lifecycle stages may be required to achieve a high level of efficacy. In two Phase IIa clinical trials, we tested immunogenicity and efficacy of RTS,S/AS01B administered alone, in a staggered regimen with viral-vectored vaccines or co-administered with viral-vectored vaccines. RTS,S/AS01B induces high titers of antibody against sporozoites and viral-vectored vaccines ChAd63 ME-TRAP and MVA ME-TRAP induce potent T cell responses against infected hepatocytes. By combining these two strategies, we aimed to improve efficacy by inducing immune responses targeting multiple parasite antigens. Vaccination with RTS,S/AS01B alone or in a staggered regimen with viral vectors produced strong immune responses and demonstrated high levels of protection against controlled human malaria infection. However, concomitant administration of these vaccines significantly reduced humoral immunogenicity and protective efficacy. Strong Th1-biased cytokine responses induced by MVA ME-TRAP were associated with a skew in circulating T follicular helper cells toward a CXCR3⁺ phenotype and a reduction in antibody quantity and quality. This study illustrates that while a multistage-targeting vaccine strategy could provide high-level efficacy, the regimen design will require careful optimization.</p

Assessment of novel vaccination regimens using viral vectored liver stage malaria vaccines encoding ME-TRAP

Heterologous prime-boost vaccination with viral vectors simian adenovirus 63 (ChAd63) and Modified Vaccinia Ankara (MVA) induces potent T cell and antibody responses in humans. The 8-week regimen demonstrates significant efficacy against malaria when expressing the pre-erythrocytic malaria antigen Thrombospondin-Related Adhesion Protein fused to a multiple epitope string (ME-TRAP). We tested these vaccines in 7 new 4- and 8- week interval schedules to evaluate safety and immunogenicity of multiple ChAd63 ME-TRAP priming vaccinations (denoted A), multiple MVA ME-TRAP boosts (denoted M) and alternating vectors. All regimens exhibited acceptable reactogenicity and CD8+ T cell immunogenicity was enhanced with a 4-week interval (AM) and with incorporation of additional ChAd63 ME-TRAP vaccination at 4- or 8-weeks (AAM or A-A-M). Induction of TRAP antibodies was comparable between schedules. T cell immunity against the ChAd63 hexon did not affect T cell responses to the vaccine insert, however pre-vaccination ChAd63-specific T cells correlated with reduced TRAP antibodies. Vaccine-induced antibodies against MVA did not affect TRAP antibody induction, and correlated positively with ME-TRAP-specific T cells. This study identifies potentially more effective immunisation regimens to assess in Phase IIa trials and demonstrates a degree of flexibility with the timing of vectored vaccine administration, aiding incorporation into existing vaccination programmes.</p

Integrated genome and transcriptome sequencing of the same cell

Single-cell genomics and single-cell transcriptomics have emerged as powerful tools to study the biology of single cells at a genome-wide scale. However, a major challenge is to sequence both genomic DNA and mRNA from the same cell, which would allow direct comparison of genomic variation and transcriptome heterogeneity. We describe a quasilinear amplification strategy to quantify genomic DNA and mRNA from the same cell without physically separating the nucleic acids before amplification. We show that the efficiency of our integrated approach is similar to existing methods for single-cell sequencing of either genomic DNA or mRNA. Further, we find that genes with high cell-to-cell variability in transcript numbers generally have lower genomic copy numbers, and vice versa, suggesting that copy number variations may drive variability in gene expression among individual cells. Applications of our integrated sequencing approach could range from gaining insights into cancer evolution and heterogeneity to understanding the transcriptional consequences of copy number variations in healthy and diseased tissues