Evidence and role of autoantibodies in chronic rhinosinusitis with nasal polyps.

In this study, we review our current knowledge of the autoimmune etiopathogenesis of chronic rhinosinusitis with nasal polyps including bacterial infections, viral infections and immunomediated mechanisms and to discuss pathogenesis with relevance for pharmacotherapy. Relevant publications on the etiopathogenesis and treatment of chronic rhinosinusitis with nasal polyps (CRSwNP) from 1977 to 2013 were analyzed. The characteristic signs and symptoms include appearance of relapsing nasal polyps, with typical symptoms such as nasal obstruction, nasal discharge and, usually, loss of the sense of smell. The etiology and pathogenesis remain unknown. Proposed theories of causation include bacterial or viral infections and immunomediated mechanisms. The autoimmune aetiology of of unknown origin or failure to respond to classic pharmacological treatments with nasal and oral steroids is now suspected. At present, the nature of the antigen trigger, the exact role played by B/T cells and anti-dsDNA autoantibodies in the pathogenesis of nasal polyposis remains unclear. Corticosteroids and surgery are the first line of treatment in CRSwNP. In the case of corticosteroid treatment failure, other drugs can be used such as rituximab, belimumab or omalizumab which have demonstrated clinical efficacy in the treatment of nasal polyposis with comorbid asthma. Immunosuppressive drugs such as methotrexate, and cyclophosphamide have also been used with varying degrees of success

Soluble HIV-1 Env trimers in adjuvant elicit potent and diverse functional B cell responses in primates

Broadly neutralizing antibodies (bNAbs) against the HIV-1 envelope glycoproteins (Envs) have proven difficult to elicit by immunization. Therefore, to identify effective Env neutralization targets, efforts are underway to define the specificities of bNAbs in chronically infected individuals. For a prophylactic vaccine, it is equally important to define the immunogenic properties of the heavily glycosylated Env in healthy primates devoid of confounding HIV-induced pathogenic factors. We used rhesus macaques to investigate the magnitude and kinetics of B cell responses stimulated by Env trimers in adjuvant. Robust Env-specific memory B cell responses and high titers of circulating antibodies developed after trimer inoculation. Subsequent immunizations resulted in significant expansion of Env-specific IgG-producing plasma cell populations and circulating Abs that displayed increasing avidity and neutralization capacity. The neutralizing activity elicited with the regimen used was, in most aspects, superior to that elicited by a regimen based on monomeric Env immunization in humans. Despite the potency and breadth of the trimer-elicited response, protection against heterologous rectal simian-HIV (SHIV) challenge was modest, illustrating the challenge of eliciting sufficient titers of cross-reactive protective NAbs in mucosal sites. These data provide important information for the design and evaluation of vaccines aimed at stimulating protective HIV-1 immune responses in humans

Antibodies directed against influenza

Antibodies that specifically bind influenza virus hemagglutinin A (HA), and antigen binding fragments thereof are disclosed herein. In several embodiments, these antibodies are broadly neutralizing. Nucleic acids encoding these monoclonal antibodies, vectors including these nucleic acids, and host cells transformed with these vectors are also disclosed. Compositions are disclosed that include these antibodies, antigen binding fragments, nucleic acids, vectors and host cells. Method of using these antibodies, and antigen binding fragments, nucleic acids, vectors and host cells, such as for diagnosis and treatment of an influenza virus infection are also provided

Malaria Transmission Blocking Vaccines

Malaria affects 198 million people and kills 584,000 each year, predominantly in Sub-Saharan Africa (WHO). The most severe form of malaria is caused by the protozoan parasite Plasmodium falciparum. Development of a vaccine against P. falciparum has been hindered by its complex life cycle with multiple antigenically distinct human and mosquito stages. To effectively prevent disease and reduce the parasite burden in populations, a vaccine will need to target multiple stages, including blocking transmission at the mosquito stage. Antibodies generated against P. falciparum mosquito stage antigen Pfs25 can prevent parasite transmission from humans to mosquitoes. However, Pfs25 is poorly immunogenic and immunization with the protein with alum as an adjuvant does not provide adequate transmission blocking activity. In this study I used adenovirus vectors (Ad) modified to express Pfs25 as a transgene or with capsid displayed Pfs25 epitopes to elicit a stronger anti-Pfs25 antibody response specifically directed at epitopes highly associated with transmission blocking. Although antibody titers are closely associated with transmission blocking activity, it is unlikely that all antibodies generated by immunization with whole protein are transmission blocking. Based on molecular models, EGF-like domains 2 and 3 appear to be most surface accessible on the parasite surface. However, antibodies raised against soluble Pfs25 equally recognize all 4 EGF-like domains. Thus, a vaccine involving in situ expression of membrane anchored Pfs25 may better target antibody responses to domains 2 and 3. Additionally, using secondary immunization with capsid displayed Pfs25, I expected to further direct the antibody response to predicted B cell epitopes. Serum from mice immunized with these vectors can provide transmission blocking activity with lower antibody titers. Innate immune responses to Ad are an important component of generating adaptive immunity and may provide a means to further augment antibody responses to poorly immunogenic protein antigens. Molecules such as Pseudomonas aeruginosa flagellin (FliC) potently activate the innate immune system and could be used to modify vectors to induce greater adaptive immunity. We have generated Ad vectors with FliC attached to protein IX. Using these vectors with capsid displayed Pfs25 epitopes I assessed whether capsid displayed FliC increases antibody titers to Pfs25

VH1-69 Utilizing Antibodies Are Capable of Mediating Non-neutralizing Fc-Mediated Effector Functions Against the Transmitted/Founder gp120

Multiple antibody effector functions arise in HIV-1 infection that could be harnessed to protect against infection or clear the persistent reservoir. Here, we have investigated the genetic and functional memory B cell and antibody landscape present during early infection in six individuals infected with either subtype A, C, or an A/C recombinant HIV-1. These individuals demonstrated varying levels of plasma autologous neutralization (nAb) against the transmitted/founder envelope (T/F Env) pseudovirus and non-neutralizing Fc-mediated effector function (nnFc) antibody-dependent cell-mediated cytotoxicity (ADCC) against the T/F Env gp120 protein at ~7 months after infection. Genetic analysis of the immunoglobulin heavy (VH) and light (VL) chain variable domain gene segments from 352 autologous T/F Env gp120-specific single B cells recovered at this same 7-month time-point revealed an over-representation of the VH1-69 germline in five of six individuals. A defining feature of the VH1-69 utilizing gp120-specific antibodies was their significantly more hydrophobic complementarity-determining region-2 (CDRH2) regions compared to other VH CDRH2 sequences from each individual. While none of the VH1-69 antibodies possessed strong neutralizing activity against virions pseudotyped with the autologous T/F Env, almost a third were capable of mediating high ADCC activity, as assayed by intracellular granzyme B activity in CEM.NKr.CCR5 target cells coated with autologous T/F Env gp120. High ADCC mediating VH1-69 antibodies exhibited shorter complementarity-determining region-3 (CDRH3) lengths and a more neutral isoelectric point than antibodies lacking this function. In the individual that developed the highest autologous ADCC responses, the high granzyme B producing antibodies bound to surface expressed envelope in the absence of CD4 and were not enhanced by the addition of soluble CD4. Overall, VH1-69 utilizing antibodies are commonly induced against gp120 in diverse HIV-1 infections and a subset of these antibodies can mediate ADCC functions, serving as a bridge between the innate and adaptive immune response to HIV-1

Visualisation and analysis of patterns in serological data using "antibody landscapes"

In this thesis I develop and implement “antibody landscapes”, a method to profile immunity against a pathogen as a function of antigenic differences between a range of strains. Theoretically applicable to any antigenically variable pathogen and measurement of immunity, the work here focusses on antibody-mediated immunity against the A/H3N2 influenza subtype. Applying the methodology to study annual serum samples from individuals monitored for influenza infection over a period of six years, patterns of influenza immunity were found to be remarkably distinct and maintained almost unchanged over time in the absence of influenza exposure. Upon infection, the initial response is strikingly antigenically broad, including responses against viruses far beyond the extent of cross-reactivity observed after a primary infection. Analysis of two vaccination cohorts, one receiving an antigenically advanced vaccine strain and one a more typical vaccine strain choice, revealed many of the same patterns of response as seen with infection. Antigenically advanced vaccination generated greater responses against later strains but surprisingly, due to equivalent boosting of prior immunity, this came at no cost to responses generated against contemporary or older strains. Exploring in more detail the development of immunity over time, analysis of a cohort of children demonstrated that - in contrast to adults with diverse exposure histories - antibody responses to a first infections were remarkably similar in pattern and magnitude. Interestingly, for second infections, although post-infection antibody titres against circulating strains were comparable to those after first infections, overall cross-reactivity of the response against future antigenic variants appeared to be diminished. The findings here underline the significant role prior-immunity plays in affecting the response to new exposures and the importance of understanding it. An important conclusion is that by failing to account for it, current approaches to influenza vaccine strain selection may be suboptimal and pre-emptive vaccine strain updates may improve overall vaccine efficacy where immunity to current strains already exists in the population. Building on the work presented here should help to optimise strain choice and vaccine efficacy even further.Part funded by an MRC studentshi

Immunological monitoring of the B-cell compartment in renal transplant recipients.

PhDB cells contribute to chronic allograft deterioration, negatively impacting graft survival, and curtailing the lifespan of a resource already in short supply. Given this, identifying alloreactive B cells could generate an important target in the battle against rejection. This study described an IgG-detecting ELISPOT used to determine if the risk of developing antibody-mediated rejection (AMR) could be predicted pretransplantation by in vitro analysis of allospecific B cells. This method failed to discriminate accurately B-cell responses to donor antigen. An alternative approach used was to detect peripheral HLA-specific B cells. Circulating HLA–A*0201 and – DQB1*0301 B cells were identified at higher frequency in sensitised patients, and this correlated with the level of serum alloantibody. Expression of HLA-DQB1*0301 B cells were at a higher frequency than HLA-A*0201 B cells in those with serum de novo donor-specific antibody (dnDSA). Next, levels of B-cell activating factor (BAFF) were investigated. Excess BAFF has been related to rejection and the development of DSA. Here elevated serum BAFF, low BAFF-receptor and DSA were all associated with deteriorating graft function. In addition intrarenal CD19+ cells, BAFF and BAFFreceptor identified with acute AMR. In contrast to a pathogenic role of B cells, a small population may be protective. The presence of regulatory B cells, defined by IL-10 production were higher in those with stable graft function, and identified with naïve B cells rather than memory B cells when compared to those with deteriorating grafts. The CD19+CD24highCD38high subset was also elevated in stable patients, and the ability to supress T-cell activation and secretion of the Th1 cell pro-inflammatory cytokine, IFN-γ was altered as a function of allograft stability. These data demonstrated characteristics within the B-cell compartment associated with stable graft function. The ability to monitor these cells may have clinical implications for predicating the risk of rejection, to dictate immunosuppressive therapy and promote allograft survival.Central London Research Ethics Committee (REC1: 07/H0707/10) Funding: NHS Blood and Transplant (UKT06-4

Travel, infection and immunity

Preface: The content of this thesis is based on research that was conducted at the travel and vaccination clinic at Leiden University Medical Centre (LUMC). This clinic provides pre-travel care to the general population, and to special groups of travellers, such as patients who use immunosuppressants or who have chronic diseases. The clinic is closely connected to the department of Infectious Diseases at LUMC. The setting of a travel clinic within an academic medical hospital, provides unique circumstances for medical research, like an experienced team of nurses, expertise regarding immunization, a constant flux of travellers and the knowledge and infrastructure that is required for research into microbiology, virology and parasitology. Examples of research that stem from this clinic are projects on immunization against malaria, yellow fever, travellers' diarrhea, poliomyelitis and hepatitis B, vaccination of immunocompromised patients, and projects on travel related acquisition of extended spectrum ß-lactamase producing Enterobacteriacae and on the utility of post-travel screening of asymptomatic travellers for parasites.UBL - phd migration 201

Antibody Levels Poorly Reflect on the Frequency of Memory B Cells Generated following SARS-CoV-2, Seasonal Influenza, or EBV Infection

The scope of immune monitoring is to define the existence, magnitude, and quality of immune mechanisms operational in a host. In clinical trials and praxis, the assessment of humoral immunity is commonly confined to measurements of serum antibody reactivity without accounting for the memory B cell potential. Relying on fundamentally different mechanisms, however, passive immunity conveyed by pre-existing antibodies needs to be distinguished from active B cell memory. Here, we tested whether, in healthy human individuals, the antibody titers to SARS-CoV-2, seasonal influenza, or Epstein–Barr virus antigens correlated with the frequency of recirculating memory B cells reactive with the respective antigens. Weak correlations were found. The data suggest that the assessment of humoral immunity by measurement of antibody levels does not reflect on memory B cell frequencies and thus an individual’s potential to engage in an anamnestic antibody response against the same or an antigenically related virus. Direct monitoring of the antigen-reactive memory B cell compartment is both required and feasible towards that goal