<p>Circulating immune complexes play an important role in the modulation of antibody responses and in the pathogenesis of immune diseases. This thesis deals with the <i>in vivo </i>regulatory properties of antibodies and their specific Fc receptors.</p><p>The immunosuppressive function of IgG is used clinically, to prevent rhesus-negative women from becoming sensitized to rhesus-positive erythrocytes from the fetus. The mechanism behind this regulation is poorly understood but involvement of a receptor for IgG, FcγRII, has been suggested. It is shown in this thesis that IgG and also IgE induce immunosuppression against sheep erythrocytes to a similar extent both in mice lacking all the known Fc receptors as in wild-type animals. These findings imply that antibody-mediated suppression of humoral responses against particulate antigens is Fc-independent and that the major operating mechanism is masking of epitopes.</p><p>Immunization with soluble antigens in complex with specific IgG leads to an augmentation of antibody production. The cellular mechanism behind this control is examined here and it is found that the capture of IgG2a immune complexes by a bone marrow-derived cell expressing FcγRI (and FcγRIII) is essential. An analysis of the ability of IgG3 to mediate this regulation indicated that, in contrast, this subclass of IgG augments antibody responses independently of FcγRI (and FcγRIII). These findings suggest that distinct mechanisms mediate the enhancing effect of different subclasses of antibodies.</p><p>Finally, the contribution of FcγRIII was studied in the development of collagen-induced arthritis (CIA), an animal model for rheumatoid arthritis in humans. It was discovered that while DBA/1 wild-type control mice frequently developed severe CIA, with high incidence, FcγRIII-deficient mice were almost completely protected, indicating a crucial role for FcγRIII in CIA.</p><p>The results presented here help to understand how immune complexes regulate immune responses <i>in vivo</i> and show that Fc receptors for IgG, if involved, could be new targets for the treatment of immune complex-related disorders.</p>

Circulating immune complexes play an important role in the modulation of antibody responses and in the pathogenesis of immune diseases. This thesis deals with the in vivo regulatory properties of antibodies and their specific Fc receptors. The immunosuppressive function of IgG is used clinically, to prevent rhesus-negative women from becoming sensitized to rhesus-positive erythrocytes from the fetus. The mechanism behind this regulation is poorly understood but involvement of a receptor for IgG, FcγRII, has been suggested. It is shown in this thesis that IgG and also IgE induce immunosuppression against sheep erythrocytes to a similar extent both in mice lacking all the known Fc receptors as in wild-type animals. These findings imply that antibody-mediated suppression of humoral responses against particulate antigens is Fc-independent and that the major operating mechanism is masking of epitopes. Immunization with soluble antigens in complex with specific IgG leads to an augmentation of antibody production. The cellular mechanism behind this control is examined here and it is found that the capture of IgG2a immune complexes by a bone marrow-derived cell expressing FcγRI (and FcγRIII) is essential. An analysis of the ability of IgG3 to mediate this regulation indicated that, in contrast, this subclass of IgG augments antibody responses independently of FcγRI (and FcγRIII). These findings suggest that distinct mechanisms mediate the enhancing effect of different subclasses of antibodies. Finally, the contribution of FcγRIII was studied in the development of collagen-induced arthritis (CIA), an animal model for rheumatoid arthritis in humans. It was discovered that while DBA/1 wild-type control mice frequently developed severe CIA, with high incidence, FcγRIII-deficient mice were almost completely protected, indicating a crucial role for FcγRIII in CIA. The results presented here help to understand how immune complexes regulate immune responses in vivo and show that Fc receptors for IgG, if involved, could be new targets for the treatment of immune complex-related disorders.

Immunity to asexual blood-stage of Plasmodium falciparum malaria is believed to be associated with protective antibodies of certain immunoglobulin classes and subclasses. This thesis addressed the importance of antibodies in relation to malaria infection and their effective interactions with Fc gamma receptor IIa (FcyRIIa) polymorphisms. Our data indicate that the frequency of FcyRIIa-R/R131 genotype was statistically significantly higher in Sudanese patients with severe malaria, while the FcyRIIa-H/H131 genotype showed a significant association with mild malaria. The levels of IgG1 and IgG3 subclass antibodies were statistically higher in the mild malaria patients. The Fulani ethnic group in West Africa has been shown to be relatively resistant to malaria. We investigated the possible impact of FcyRIIa polymorphisms in the Fulani and non-Fulani in Mali and Sudan, and analysed their malaria-reactive IgG subclass profiles. The FcyRIIa-H/H131 genotype and H131-allele were found to be prevalent in the Fulani while R131-allele was prevalent in non-Fulani. The Fulani had higher serum levels of IgG1-3, with higher proportion of IgG2 than the non-Fulani. Most clinico-epidemiology studies have been in areas with holo- and hyper-malaria endemicity. We have analysed antibody responses to a panel of six blood-stage antigens in relation to clinical malaria outcome in mesoendemic Sudan. Our results revealed a linear association with anti-AMA-1 IgG1 antibodies and reduced risk of severe malaria while a non-linear relationship with IgG3 antibodies was observed for MSP-2, MSP-3 and GLURP. In the combined final model, the highest levels of IgG1 subclass antibodies to AMA-1, GLURP-R0, and the highest levels of IgG3 subclass antibodies reactive to 3D7 MSP-2 were independently associated with a reduced risk of clinical malaria. Taken together, these data suggest a possible association between FcyRIIa-R/H131 and anti-malarial antibody responses in the clinical outcome of malaria.

In regions where endemic measles virus circulation has been interrupted, laboratory confirmation of measles is like puzzle solving. The complexity of these puzzles depends on the available pieces of clinical, epidemiological and laboratory information. The main goal of this dissertation is to evaluate diagnostic laboratory tools to aid in suspected case confirmation in these settings. First, protocols to elute measles IgM and IgG antibodies from blood spots dried onto filter paper were compared to propose one that will permit the recovery of the maximum volume of eluted sample in the minimum time, effort and cost. An easy-to-implement protocol is proposed for the rapid extraction of serum for measles/rubella serology in outbreak situations for use in the World Health Organization Global Measles and Rubella Laboratory Network. Second, due to inherent limitations of measles specific IgM enzyme immunoassays and molecular methods used for measles confirmation, not all suspected cases can be resolved. For example, IgM and RNA may not be detected in vaccinated cases with waning immunity (secondary vaccine failures) and presenting with modified measles. The observation is made that serological parameters of elevated titers of high avidity neutralizing antibodies correlate with measles secondary vaccines failures and may be useful biomarkers for confirming secondary vaccine failures that cannot be confirmed otherwise. Third, a highly accurate measles IgG avidity enzyme immunoassay for vaccine failure classification is described. Detection of low avidity antibodies using this highly sensitive and specific avidity assay can complement existing measles diagnostic tools in confirming suspected cases when routine IgM testing may be inconclusive. Therefore, these diagnostic approaches can provide additional laboratory information to resolve suspected cases irrespective of vaccination status. Together, data presented in this dissertation may assist in enhancing measles control and surveillance in elimination settings. / En les regions on s'ha interromput la circulació del virus del xarampió endèmic, la confirmació del xarampió a nivell de laboratori és com resoldre trencaclosques. La complexitat d'aquests trencaclosques depèn de les peces disponibles d'informació clínica, epidemiològica i de laboratori. L'objectiu principal d'aquesta tesi doctoral és avaluar eines de diagnòstic de laboratori per a ajudar en la confirmació de casos sospitosos en regions on el xarampió està eliminat. En primer lloc, es van comparar protocols per a eluir les IgM i IgG anti-xarampionoses de mostres de taques de sang seca sobre paper de filtre i proposar un protocol que permetés la recuperació del volum màxim de mostra eluïda en el mínim temps, esforç i cost. Es proposa un protocol de fàcil implementació dins de la xarxa de laboratoris de rubèola i xarampió de l'Organització Mundial de la Salut per a ser usat en situacions de brot. En segon lloc, les IgM anti-xarampionoses i l’ARN del virus del xarampió poden no ser detectats en casos vacunats amb disminució de la immunitat (fallada vacunal secundària) i amb simptomatologia de xarampió modificada. L'observació de que paràmetres serològics de títols elevats d'anticossos anti-xarampionosos neutralizants i d'alta avidesa correlacionen amb fallades vacunals secundàries permet proposar aquests paràmetres com a biomarcadors útils per a confirmar aquestes fallades vacunals quan d'altra manera no podrien confirmar-se. En tercer lloc, es descriu un assaig immumoenzimàtic per a determinar l'avidesa de les IgG contra el virus del xarampió per a la classificació de fallades vacunals. Aquest assaig és altament sensible, específic i precís. La detecció d'anticossos de baixa avidesa mitjançant aquest assaig pot a més complementar les eines actuals de diagnòstic de xarampió en la confirmació de casos sospitosos quan les proves rutinàries de IgM no són concloents. Per tant, aquestes estratègies diagnòstiques poden proporcionar informació de laboratori addicionals per a resoldre casos sospitosos de xarampió independentment del seu estat de vacunació. Les dades presentades en aquesta tesi doctoral poden ajudar a millorar el control del xarampió i vigilància epidemiològica allà on el xarampió ja està eliminat.

Previous studies have demonstrated that chitosan and its derivative, N-trimethyl chitosan chloride (TMC) are effective and safe absorption enhancers to improve mucosal delivery of macromolecular drugs including vaccines. Furthermore, chitosan and TMC can easily form microparticles and nanoparticles, which have the ability to encapsulate large amounts of antigens. Emzaloid™ technology has proven in the past to be an effective delivery system for numerous drugs. Emzaloids can entrap, transport and deliver large amounts of drugs including vaccines. In this study, the ability of chitosan microparticles and nanoparticles, TMC microparticles as well as micrometer and nanometer range Emzaloids to enhance both the systemic and mucosal (local) immune response against diphtheria toxoid (DT) after nasal administration in mice was investigated. The above mentioned formulations were prepared and characterised according to size and morphology. DT was then associated to the chitosan microparticles and nanoparticles as well as TMC microparticles to determine the antigen loading and release. It was found that the loading efficacy of the formulations was 88.9 %, 27.74 % and 63.1 % respectively, and the loading capacity of the formulations was 25.7 %, 8.03 % and 18.3 %. DT loaded and unloaded (empty) chitosan microparticles and nanoparticles, TMC microparticles, micrometer and nanometer range Emzaloids as well as DT in phosphate buffered saline (PBS) were administered nasally to mice. Mice were also vaccinated subcutaneous with DT associated to alum as a positive control. All mice were vaccinated on three consecutive days in week 1 and boosted in week 3. Sera was analysed for anti- DT IgG and nasal lavages were analysed for anti-DT IgA using an enzyme linked imrnunosorbent assay (ELISA). In the study conducted to determine the systemic (IgG) and local (IgA) immune responses it was seen that DT associated to all the experimental formulations produced a systemic immune response. The said formulations produced a significantly higher systemic immune response when compared to the formulation of DT in PBS. Furthermore, the mice vaccinated with DT associated to the TMC formulations showed a much higher systemic immune response than the mice that were vaccinated subcutaneously with DT associated to alum, whereas the other formulations produced systemic immune responses that were comparable to that of DT associated to alum. It was also found that DT associated to the experimental formulations produced a local immune response, however only DT associated to TMC microparticles produced a consistent local immune response. It can be concluded from the in vivo experiments that the TMC formulations, moreover, the TMC microparticles is the most effective and promising formulation for the nasal delivery of vaccines. / Thesis (M.Sc. (Pharmaceutics))--North-West University, Potchefstroom Campus, 2005.

Previous studies have demonstrated that chitosan and its derivative, N-trimethyl chitosan chloride (TMC) are effective and safe absorption enhancers to improve mucosal delivery of macromolecular drugs including vaccines. Furthermore, chitosan and TMC can easily form microparticles and nanoparticles, which have the ability to encapsulate large amounts of antigens. Emzaloid™ technology has proven in the past to be an effective delivery system for numerous drugs. Emzaloids can entrap, transport and deliver large amounts of drugs including vaccines. In this study, the ability of chitosan microparticles and nanoparticles, TMC microparticles as well as micrometer and nanometer range Emzaloids to enhance both the systemic and mucosal (local) immune response against diphtheria toxoid (DT) after nasal administration in mice was investigated. The above mentioned formulations were prepared and characterised according to size and morphology. DT was then associated to the chitosan microparticles and nanoparticles as well as TMC microparticles to determine the antigen loading and release. It was found that the loading efficacy of the formulations was 88.9 %, 27.74 % and 63.1 % respectively, and the loading capacity of the formulations was 25.7 %, 8.03 % and 18.3 %. DT loaded and unloaded (empty) chitosan microparticles and nanoparticles, TMC microparticles, micrometer and nanometer range Emzaloids as well as DT in phosphate buffered saline (PBS) were administered nasally to mice. Mice were also vaccinated subcutaneous with DT associated to alum as a positive control. All mice were vaccinated on three consecutive days in week 1 and boosted in week 3. Sera was analysed for anti- DT IgG and nasal lavages were analysed for anti-DT IgA using an enzyme linked imrnunosorbent assay (ELISA). In the study conducted to determine the systemic (IgG) and local (IgA) immune responses it was seen that DT associated to all the experimental formulations produced a systemic immune response. The said formulations produced a significantly higher systemic immune response when compared to the formulation of DT in PBS. Furthermore, the mice vaccinated with DT associated to the TMC formulations showed a much higher systemic immune response than the mice that were vaccinated subcutaneously with DT associated to alum, whereas the other formulations produced systemic immune responses that were comparable to that of DT associated to alum. It was also found that DT associated to the experimental formulations produced a local immune response, however only DT associated to TMC microparticles produced a consistent local immune response. It can be concluded from the in vivo experiments that the TMC formulations, moreover, the TMC microparticles is the most effective and promising formulation for the nasal delivery of vaccines. / Thesis (M.Sc. (Pharmaceutics))--North-West University, Potchefstroom Campus, 2005.

Hantaviruses are the causative agents of hemorrhagic fever with renal syndrome (HFRS) in Eurasia, and of hantavirus cardiopulmonary syndrome (HCPS) in the Americas. Transmission to humans usually occurs by inhalation of aerosolized virus-contaminated rodent excreta. To date, human-to-human transmission has only been described for the Andes hantavirus. The mode of transmission of Andes hantavirus is not yet known, but transmission through saliva has been suggested. In Sweden, we have one hantavirus that is pathogenic to humans, Puumala virus (PUUV), which is endemic in Central and Northern Europe. It induces a relatively mild form of HFRS, also called nephropathia epidemica (NE). The rodent reservoir is the bank vole (Myodes glareolus). The mechanism behind the pathogenesis of hantavirus is complex and probably involves both virus-mediated and host-mediated mechanisms. The aim of this project was to investigate the transmission mechanisms and pathogenesis of hantavirus disease in humans. In our first study, we described the largest outbreak of PUUV so far in Sweden. We investigated factors that might be important for causing the outbreak, and suggested that a peak in the bank vole population together with concurrent extreme weather conditions most probably contributed to the outbreak. Our next studies concentrated on human-to-human transmission of hantaviruses. We found PUUV RNA in saliva from PUUV-infected patients, suggesting that there is PUUV in the saliva of infected humans, although no person-to person transmission appears to occur with PUUV.  In the studies that followed, we showed that human saliva and human salivary components could inhibit hantavirus replication. We also found PUUV-specific IgA in the saliva of PUUV-infected patients, which might prevent person-to-person transmission of the virus.  In the final study, we focused on the pathogenesis of NE. One hundred five patients were included in a prospective study.  They were divided into a group with mild disease and a group with moderate or severe disease. We found that the immune response had a dual role in disease development. It was partly responsible for development of severe disease, with significantly higher amounts of neutrophils in severely ill patients, but it was also protective against severe disease, because patients with mild disease had higher levels of PUUV-specific IgG. In conclusion, a peak in the bank vole population in combination with extreme weather will increase the risk of human infection, PUUV RNA is present in saliva, PUUV-specific IgA and salivary components inhibit person-to-person transmission of PUUV, and the immune response is important for the pathogenesis of PUUV and the severity of the disease. / Hantavirus är en grupp av virus som finns hos gnagare som bär på viruset utan att själva bli märkbart sjuka. Varje hantavirus har anpassat sig till sin egen art av gnagare som de infekterar (kallas virusets reservoar). Hantaviruset kan överföras till människor från gnagare och kallas då för en zoonos eftersom detsprids från djur till människa. I människa orsakar hantavirus blödarfeber med njurpåverkan i Eurasien och blödarfeber med med hjärt och lungpåverkan i Nord- och Sydamerika. I Sverige har vi bara ett hantavirus som är sjukdomsframkallande hos människor, Puumala-viruset som även finns i delar av övriga Europa. Det framkallar en relativt mild form av blödarfeber, som kallas sorkfeber eller Nephropathia epidemica. Puumala-virusets reservoar är skogssorken (Myodes glareolus). Människor smittas oftast av hantavirus när de andas in infekterat damm som innehåller utsöndringar (avföring, urin eller saliv) från gnagare som har torkat in och sedan blivit luftburet. Vad man vet hittills så finns det bara ett hantavirus som smittar från person till person, för övriga hantavirus är människan en ”dead end”. Det virus som kan smitta från person till person heter Andes hantavirus och finns i Sydamerika. Andes hantavirus har en mus som reservoar från vilken människor kan smittas, sedan har smittan i vissa fall förts vidare från människa till människa, som tur är har dessa utbrott gått att stoppa. Fastän utbrotten har varit små har många personer dött, eftersom dödligheten är så hög, ungefär 30-40% av de diagnostiserade fallen dör. Hur Andes hantavirus överförs från människa till människa är inte känt men överföring genom saliv har föreslagits. Hur viruset ger upphov till sjukdom hos människa är inte klarlagt. Studier talar för att mekanismen bakom sjukdomsutvecklingen (den så kallade patogenesen) hos hantavirusorsakade blödarfebrar är komplex. Sannolikt beror patogenesen både på egenskaper hos viruset och värden d.v.s. människan som är smittad av viruset. Vårt mål med detta projekt var att undersöka vad som hindrar överföring av Puumala hantavirus från människa till människa och att undersöka hur virusinfektionen påverkar sjukdomsutvecklingen hos människan. I vår första studie beskrev vi det största utbrottet av sorkfeber hittills i Sverige och vi undersökte faktorer som kan ha orsakat utbrottet. Vi föreslog att en topp i skogssorkpopulationen samtidigt med extremt varmt väder troligen bidrog till utbrottet. Utbrottet skedde i december och det extremt varma vädret medförde att snön smälte bort. Sorkarna bor vanligtvis under snön på vintern, vi tror att frånvaro av snötäcke fick sorkarna att söka sig till byggnader för att söka skydd och där kom i kontakt med människor. Våra efterföljande studier fokuserade på överföring av hantavirus från människa till människa. Vi hittade Puumala-virusets arvsmassa (RNA) i saliv från sorkfeberpatienter, vilket tyder på att det finns Puumala-virus i saliven hos infekterade människor, även om ingen överföring från person till person verkar inträffa. I efterföljande studier visade vi att mänsklig saliv och mänskliga salivkomponenter minskar hantavirus smittsamhet. Vi fann också Puumala-virusspecifika IgA-antikroppar i saliven från sorkfeberpatienter, vilket kan förhindra överföring från person till person. I den sista studien fokuserade vi på patogenesen hos människor efter hantavirusinfektion. 105 patienter ingick i en prospektiv studie och delades in i en grupp med mild sjukdom och en grupp med måttlig/svår sjukdom. Vi hittade en dubbel roll hos immunsvaret för sjukdomsutvecklingen. Immunsvaret var delvis ansvarig för utveckling av svår sjukdom med betydligt högre mängd neutrofiler hos svårt sjuka patienter, men det var också skyddande mot allvarlig sjukdom, eftersom patienter med en mild sjukdom hade högre nivåer av Puumalavirusspecifika IgG-antikroppar. Detta talar för att behandling med IgG-antikroppar specifikt riktade mot hantavirus skulle kunna vara effektiv hos hantavirusinfekterade patienter. Sammanfattningsvis; en topp i skogssorkspopulationen i kombination med extremt väder ökar risken för infektion hos människor; Puumala-virus arvsmassa (RNA) finns i saliv; Puumala-virusspecifika IgA-antikroppar och salivkomponenter hämmar överföring av Puumalavirus från person till person; immunsvaret är viktigt för Puumala-virus patogenes och sjukdomens svårighetsgrad.

Diss. (sammanfattning) Stockholm : Stockholms universitet, 2008. / Härtill 5 uppsatser.

O lúpus eritematoso sistêmico (LES) é uma doença autoimune e a suscetibilidade às infecções está associada as suas anormalidades imunológicas e a sua terapia imunossupressora e citotóxica. Dentre as alterações que predispõem às infecções no LES estão anormalidades moleculares e funcionais dos neutrófilos: clearance e fagocitose ineficientes de imunocomplexos (IC) e bactérias, neutropenia, defeitos na quimiotaxia, redução do burst oxidativo e redução da expressão de receptores para IgG (Fc?R) e para complemento (CR). Além disso, os polimorfismos genéticos dos Fc?R têm sido associados com as disfunções imunes do LES. Os Fc?R são importantes mediadores das funções efetoras do neutrófilo e atuam em sinergismo com os CR (CR1 e CR3). O polimorfismo dos genes Fc?RIIA e Fc?RIIIB determina a expressão de variantes alélicas com diferenças funcionais, as quais podem influenciar as respostas biológicas e a suscetibilidade e o prognóstico das doenças infecciosas. Em particular, o alótipo Fc?RIIa-R131, tem menor afinidade para a IgG2, o que resulta em prejuízo na fagocitose mediada por esta imunoglobulina. A IgG2 é essencial contra bactérias encapsuladas e os pneumococos são responsáveis por 6-18% das infecções bacterianas no LES. O objetivo deste estudo foi investigar a influência dos polimorfismos genéticos dos Fc?R na fagocitose e desgranulação dos neutrófilos, associados ao polimorfismo do CR3, e à ocorrência de infecções bacterianas no LES. Os genótipos foram determinados por reações da polimerase em cadeia para os polimorfismos das variantes alélicas; a fagocitose e desgranulação foram estimuladas por IC contendo IgG (IC-IgG) e por IC-IgG e complemento (IC-IgG/SHN); a fagocitose de IC por neutrófilos e a expressão dos Fc?R e CR3 foram avaliadas por citometria de fluxo; e a desgranulação dos neutrófilos estimulada por IC foi medida pela liberação de elastase e lisozima. Os resultados mostraram: maior frequência para o genótipo R-131 no LES; associação do genótipo HNA-4a negativo para o CR3 com a suscetibilidade para o LES e associação do HNA-4a positivo com proteção; fagocitose menor em neutrófilos de pacientes com LES com genótipo HR-131 comparados aos neutrófilos do grupo controle com genótipo R-131 (IC-IgG e IC-IgG/SHN); no polimorfismo do Fc?RIIIb, a fagocitose e a lisozima foram menores em neutrófilos de pacientes com LES com genótipo HNA-1b e maior para HNA-1a/1b comparados aos controles com os respectivos genótipos (IC-IgG/SHN); a ocorrência de infecções foi mais frequentemente associada à presença do alelo R-131 e HNA-1b; nenhuma diferença foi observada para o polimorfismo HNA-4a do CR3, bem como para e elastase. Este estudo contribui para o entendimento das anormalidades nas funções dos neutrófilos no LES e para a identificação de indivíduos, cujo polimorfismo dos Fc?R e CR3 possa conferir suscetibilidade ou proteção às infecções. / Systemic lupus erythematosus (SLE) is an autoimmune disease in which disease-related and genetic factors and immunosuppressive and cytotoxic therapies all contribute to an increased susceptibility to infections. Factors predisposing to infection include defects in chemotaxis, abnormalities in neutrophil phagocytic activity, decreased immune complex (IC) and bacteria clearance, neutropenia, reduced oxidative burst, abnormalities in the expression of Fc?R (Fc?R) and complement (CR) receptors. Recent data have provided evidence that genetic polymorphism of Fc?R is associated with immune abnormalities and risk to development of SLE. Fc?R can mediate neutrophil effector functions and play a synergistic action with CR. Fc?RIIa and Fc?RIIIb display functionally relevant genetic polymorphisms, which allelic variants can influence the biological responses and the susceptibility to and course of infectious diseases. In particular, the presence of the Fc?RIIa-R131 allotype results in lower affinity binding to IgG2, a subclass of IgG specific for encapsulated bacteria. Since pneumococci accounts for 6-18% of all bacterial infections in SLE, the R131 allele can be relevant as a risk factor for infections. The aim of this study was to investigate the influence of the Fc?R polymorphisms on the phagocytosis and degranulation of neutrophils, associated with CR3 polymorphism, and bacterial infections in SLE. Genotypes were determined by polymerase chain reactions for the polymorphisms of the allelic variants; the phagocytosis and the degranulation were stimulated with IC-containing IgG (IC-IgG) and IC-IgG and complement (IC-IgG/NHS); the phagocytosis of IC by neutrophils and expression of Fc?R and CR3 were evaluated by flow cytometry, and degranulation of neutrophils stimulated with IC was measured by the release of lysozyme and elastase. The results showed a higher frequency for the genotype R-131 in SLE and association of genotype HNA-4a negative for the CR3 with susceptibility to SLE and association of HNA-4a with protection; the phagocytosis was lower in neutrophils from patients with SLE with HR-131 genotype than those in neutrophils from control group with genotype R-131 (IC-IgG and IC-IgG/NHS); in the Fc?RIIIb polymorphism, the phagocytosis and lysozyme were lower in neutrophils from patients with SLE with genotype HNA-1b and higher for HNA-1a/1b than those controls with the respective genotypes (IC-IgG/NHS); the occurrence of infections was more frequently associated with the presence of the allele R-131 and HNA-1b; no difference was observed for polymorphism HNA-4a of the CR3, neither for elastase. This study can contribute for the understanding of neutrophil abnormalities in SLE and identifying genetic markers that would predict patients Who are at high risk for infections.

La pharmacocinétique (PK) des anticorps monoclonaux (mAbs) est sujette à d’importantes variations interindividuelles. Le récepteur néonatal au Fc des IgG (FcRn) et le statut immun à l’encontre de ces mAbs sont des déterminants de cette PK. La bioconjugaison des mAbs à des cytotoxiques entraîne une altération de leur PK. Nous montrons que le taux de couplage modifie l’affinité de ces espèces pour le FcRn à pH6. La proportion d’agrégats au sein des solutions d’anticorps armés augmente avec le taux de couplage et pourrait entraîner une altération de leur PK. Par ailleurs, cette agrégation est impliquée dans l’immunogénicité des mAbs, et nous avons donc cherché à identifier des acides aminés impliqués dans l’agrégation de mAbs indiqués en clinique. Il apparait que la nature biochimique de résidus des paratopes pourrait augmenter cette agrégation. Les anti-TNF- présentent très peu d'agrégats et figurent pourtant parmi les plus immunogènes chez l’Homme. Nous avons donc exploré le rôle des complexes immuns dans leur immunogénicité chez la souris. Il apparait que la présence du FcRn n’est pas à l’origine de l’immunisation contre ces mAbs, contrairement à celle des complexes immuns. Ces résultats donnent des pistes pour la production de mAbs plus efficients et mieux tolérés. / The pharmacokinetic (PK) profile of monoclonal antibodies (mAbs) shows interindividudal variability. The neonatal Fc receptor (FcRn) and the immounogenicity of these mAbs are determinative factors of mAb PK. Generation of antibody-drug-conjugates alters their PK profile. We show that the the affinity for FcRn at pH6 increases with the drug-to-mAb ratio, as does the amount of aggregates inside the mAb-drug-conjugate. The amount of aggregates could be responsible for an avidity effect towards FcRn. These aggregates are known to cause immunogenicity, so we studied biochemical determinants inside the aminoacid sequence of marketed mAbs. We show that the biochemical nature of some aminoacids inside the paratope has an impact on the amount of aggregation. Anti-TNF- mAbs show very little aggregation but are very immunogenic in humans. We studied the role of the formation of immune complexes in the immunization against anti-TNF- mAbs in mice, and showed that immune complexes, but not FcRn are essential in the immunization process against anti- TNF- mAbs. These results give leads towards the generation of more efficient, better tolerated mAbs.