Aspergillus-specific antibodies – Targets and applications

Aspergillus is a fungal genus comprising several hundred species, many of which can damage the health of plants, animals and humans by direct infection and/or due to the production of toxic secondary metabolites known as mycotoxins. Aspergillus-specific antibodies have been generated against polypeptides, polysaccharides and secondary metabolites found in the cell wall or secretions, and these can be used to detect and monitor infections or to quantify mycotoxin contamination in food and feed. However, most Aspergillus-specific antibodies are generated against heterogeneous antigen preparations and the specific target remains unknown. Target identification is important because this can help to characterize fungal morphology, confirm host penetration by opportunistic pathogens, detect specific disease-related biomarkers, identify new candidate targets for antifungal drug design, and qualify antibodies for diagnostic and therapeutic applications. In this review, we discuss how antibodies are raised against heterogeneous Aspergillus antigen preparations and how they can be characterized, focusing on strategies to identify their specific antigens and epitopes. We also discuss the therapeutic, diagnostic and biotechnological applications of Aspergillus-specific antibodies.     

The “less-is-more” in therapeutic antibodies: Afucosylated anti-cancer antibodies with enhanced antibody-dependent cellular cytotoxicity

Therapeutic monoclonal antibodies are the fastest growing class of biological therapeutics for the treatment of various cancers and inflammatory disorders. In cancer immunotherapy, some IgG1 antibodies rely on the Fc-mediated immune effector function, antibody-dependent cellular cytotoxicity (ADCC), as the major mode of action to deplete tumor cells. It is well-known that this effector function is modulated by the N-linked glycosylation in the Fc region of the antibody. In particular, absence of core fucose on the Fc N-glycan has been shown to increase IgG1 Fc binding affinity to the FcγRIIIa present on immune effector cells such as natural killer cells and lead to enhanced ADCC activity. As such, various strategies have focused on producing afucosylated antibodies to improve therapeutic efficacy. This review discusses the relevance of antibody core fucosylation to ADCC, different strategies to produce afucosylated antibodies, and an update of afucosylated antibody drugs currently undergoing clinical trials as well as those that have been approved.     

Why recombinant antibodies — benefits and applications

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Xenoreactive natural antibodies and induced antibodies — their effects on beating cardiomyocytes as a model of a xenograft

Xenotransplantation has been complicated by hyperacute rejection reactions, which are supposedly triggered by preformed natural antibodies (PNAb) of the recipient organism, whereas the role of antibodies specifically induced by previous antigen contact (IAb) is less clear.Primary cultures of spontaneously beating neonatal rat cardiomyocytes were used as a model of the heart to elaborate the effects of both PNAb and IAb from xenogeneic species and to investigate into their mechanisms of action. An experimental setup allowing for rapid medium exchange under continuous observation was employed.Sera containing PNAb reproducibly bring about a stereotype pattern of altered contractility including an initial increase in beating frequency followed by a temporary cessation of beating within the first minutes after administration. After recovery of spontaneous contractions, the cells within the monolayer exhibited a dissociation of the synchronicity of the beating persisting for several hours. The temporary pause in beating was prevented by a very high extracellular calcium concentration, but not by extracellular electrical stimulation sufficient to trigger contractions in control cells. Electrophysiological measurements carried out in adult ventricular guinea pig heart muscle cells under the same experimental conditions revealed an increase of the excitation threshold of the cells after application of sera containing PNAb due to an enhanced input resistance. These results indicate that the effect of PNAb is the consequence rather of a generally reduced excitability of the cell than of the inhibition of a singular ionic conductance. After specific absorption of PNAb directed against rat antigens beating of neonatal rat cardiomyocytes ensued without interruption. Sera specimens devoid of complement produced similar effects on contractility, although the duration of the standstill period was significantly shorter. The increase in input resistance visualized in guinea pig myocytes was absent after removing PNAb against guinea pig antigens but not after absorbing PNAb directed against rat epitopes.Signs of a permanent cytotoxicity after the administration of PNAb were lacking in all experiments.IAb against rat heart tissue raised in rabbits stopped the contractions of neonatal rat cardiomyocytes within 30 min after administration irreversibly and lead concentration-dependently to a destruction of the cells. (Mol Cell Biochem 160/161: 315–324, 1996)     

Autoreactive-Aβ antibodies promote APP β-secretase processing

Several prior investigations of Alzheimer's disease (AD) patients have indicated naturally occurring autoantibodies against amyloid-β (Aβ) species are produced. Although many studies have focused on the relative concentrations or binding affinities of autoantibodies against Aβ-related proteins in AD and aging, data regarding their functional properties are limited. It is generally believed that these antibodies act to aid in clearance of Aβ. However, as antibodies which bind to Aβ also typically bind to the parent amyloid precursor protein (APP), we reasoned that certain Aβ-targeting autoantibodies may bind to APP thereby altering its conformation and processing. Here we show for the first time, that naturally occurring Aβ-reactive autoantibodies isolated from AD patients, but not from healthy controls, promote β-secretase activity in cultured cells. Furthermore, using monoclonal antibodies to various regions of Aβ, we found that antibodies generated against the N-terminal region, especially Aβ(1-17) , dose dependently promoted amyloidogenic processing of APP viaβ-secretase activation. Thus, this property of certain autoantibodies in driving Aβ generation could be of etiological importance in the development of sporadic forms of AD. Furthermore, future passive or active anti-Aβ immunotherapies must consider potential off-target effects resulting from antibodies targeting the N-terminus of Aβ, as co-binding to the corresponding region of APP may actually enhance Aβ generation.     

Antibacterial monoclonal antibodies: the next generation?

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Monoclonal antibodies against plasma protease inhibitors: I. Production and characterization of 23 monoclonal antibodies against human α2

23 hybridomas secreting monoclonal antibodies against human ₂ , the fast-acting inhibitor of plasmin present in plasma, have been produced by the cell-fusion technique. Isotyping of the monoclonal antibodies has revealed that 14 monoclonal antibodies belong to the class IgG1, 6 to the class IgG2a, and 3 to the class tgG2b. All light chains belong to the group. The specificity and relative avidity of these monoclonals have been determined Using an indirect enzyme-linked immunosorbent assay. 13 monoclonals exhibit a relatively high avidity for ₂ , 5 are of intermediate avidity, and 5 of low avidity. The epitope specificity of these 23 rnonoclonal antibodies, originating from a single mouse, have been examined in inhibition experiments. A group of 10 monoclonal antibodies exhibit a very similar inhibition pattern. Partial inhibition effects displayed by 10 other antibodies define partially overlapping antigenic regions. The binding of these antibodies seems to produce a conformational change in the ₂ molecule, reducing the binding of two other antibodies. The last antibody defines an independent epitope.     

Anti-bacterial monoclonal antibodies: Back to the future?

Today's medicine has to deal with the emergence of multi-drug resistant bacteria, and is beginning to be confronted with pan-resistant microbes. This worsening inadequacy of the antibiotics concept, which has ruled infectious medicine in the last six decades creates an increasing unmet medical need that can be addressed by passive immunization. While past experience from the pre-antibiotic era with serum therapy was in many cases encouraging, antibacterial monoclonal antibodies have so far suffered high attrition rates in the clinic, generally from lack of efficacy. Yet, we believe that recent developments in a number of areas such as infectious disease pathogenesis research, translational medicine, mAb engineering, mAb manufacturing and rapid bedside diagnostics are converging to make the medium-term future permissive for antibacterial mAb development. Here, we review antibacterial mAb-based approaches that are or were in clinical development, and may potentially act as paradigms with regards to molecular targets, antibody formats and mode-of-action, pre-clinical validation and selection of most relevant patient populations, in order to increase the likelihood of successful product development in this field.     

Potential therapeutic use of antibodies directed towards HuIFN-γ

IFN-γ is an important regulator of immune responses and inflammation. Studies in animal models of inflammation, autoimmunity, cancer, transplant rejection and delayed-type hypersensitivity have indicated that administration of antibodies against IFN-γ can prevent the occurrence of diseases or alleviate disease manifestations. Therefore, it is speculated that such antibodies may have therapeutical efficacy in human diseases. Since animal-derived antibodies are immunogenic in patients several strategies are being developed in order to reduce or abolish this human anti-mouse antibody (HAMA) response. In our laboratory, we have constructed a single-chain variable fragment (scFv) derived from a mouse antibody with neutralizing potential for human IFN-γ. A scFv consists of only variable domains tethered together by a flexible linker. The scFv was demonstrated to neutralize the antiviral activity of HuIFN-γin vitro and therefore might be considered as a candidate for human therapy.     

The problem of natural antibodies, 1894–1905

As we have seen, natural antibodies first emerged as an experimental phenomenon without a plausible theoretical explanation. They were originally denied the status of antibody; then, adjustments to the side-chain theory transformed them from a curiosity into a foundation of the theory. However, in accommodating natural antibodies, Ehrlich had opened several holes in his mechanism of antibody formation.Thus, by 1905, natural antibodies were clearly established as problematic. From the practical standpoint, it seemed unwise to maintain an identity between normal and immune antibodies, given the therapeutic differences in their avidity. With the decline of Ehrlich's theory of antibody formation and the spread of Landsteiner's hapten technique for the production of antibodies against artificial antigens after World War I, the theoretical possibility of their existence as other than anomaly seemed more remote than ever. However, outside the theory and despite clinical considerations, natural antibodies remained a perplexing experimental phenomenon.⁴⁹     

Immunocytochemical study of seminal γ-glutamyltransferase using monoclonal antibodies

We produced three monoclonal antibodies, SG1, SG2 and SG3, specific for human seminal -glutamyltransferase when characterized by enzyme-linked immunosorbent assay and immunoblotting. Seminal -glutamyltransferase was localized, by immunostaining, to the epithelial cells of the ductus epididymidis, seminal vesicle and prostate gland with SG1, those of the prostate gland with SG2, and those of the seminal vesicle with SG3. Rabbit polyclonal anti-seminal -glutamyltransferase serum reacted with the proximal convolution of the kidney and the bile capillaries of the liver, and with the epithelial cells of the reproductive organs. However, immunoreactivity was not observed in the kidney or liver with the monoclonal antibodies. Thus, these monoclonal antibodies are probably all specific to seminal -glutamyltransferase but recognize different epitopes.     

Alternative affinity tools: more attractive than antibodies?

Antibodies are the most successful affinity tools used today, in both fundamental and applied research (diagnostics, purification and therapeutics). Nonetheless, antibodies do have their limitations, including high production costs and low stability. Alternative affinity tools based on nucleic acids (aptamers), polypeptides (engineered binding proteins) and inorganic matrices (molecular imprinted polymers) have received considerable attention. A major advantage of these alternatives concerns the efficient (microbial) production and in vitro selection procedures. The latter approach allows for the high-throughput optimization of aptamers and engineered binding proteins, e.g. aiming at enhanced chemical and physical stability. This has resulted in a rapid development of the fields of nucleic acid- and protein-based affinity tools and, although they are certainly not as widely used as antibodies, the number of their applications has steadily increased in recent years. In the present review, we compare the properties of the more conventional antibodies with these innovative affinity tools. Recent advances of affinity tool developments are described, both in a medical setting (e.g. diagnostics, therapeutics and drug delivery) and in several niche areas for which antibodies appear to be less attractive. Furthermore, an outlook is provided on anticipated future developments.     

Is there an association between antiphospholipid antibodies and psoriasis?

Ever increasing evidence supports the association between psoriasis and cardiovascular risk. Antiphospholipid antibodies (APAs), which can occur in many autoimmune diseases, are considered prothrombotic and have been associated with atherosclerosis. The aim of this study is to evaluate the prevalence and levels of APAs in psoriasis patients. Fifty patients with moderate to severe plaque psoriasis and 48 healthy subjects were investigated for lupus anticoagulant (LAC) by screening and confirmatory coagulation tests, as well as for antibodies against cardiolipin or beta2-glycoprotein I. Levels of APAs and LAC-related parameters were similar for patients with psoriasis and normal controls (p>0.05). APAs were found in only one psoriatic patient (2%) and in none of the controls. LAC was detected in 2 patients (4%) and in one subject of the control group (2.1%). These results suggest that the prevalence of APAs is not increased in plaque psoriasis as compared to the control group. The increased cardiovascular risk observed in psoriatic patients is therefore likely to be correlated to factors different from APAs.     

“Relatedness” between antibodies —Implications on immunogenetic systems

Summary A serologic model, a mathematical treatment of this model as well as experimental serologic evidence have all made it highly probable that a new type of relatedness between antibodies, (at least assumed to react with antigens within the same immunogenetic system) is a very common phenomenon.It is shown that conventional models assuming monospecific and unrelated base reagents give rise to erroneous assumptions regarding the serologic and genetic aspects of immunogenetic systems if the base reagents are instead related. Thus a relation between antibodies could falsely be interpreted as being instead a relation between antigens on a serologic; genetic or biochemical level.

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Zusammenfassung Die statistische Behandlung eines serologischen Modells und experimentelle serologische Ergebnisse führen zu der Annahme einer häufig vorkommenden neuartigen Beziehung zwischen Antikörpern.Voraussagen über serologische und genetische Aspekte immunogenetischer Systeme können unter der Voraussetzung der hier postulierten Beziehung deshalb zu Irrtümern führen, weil die konventionellen Modelle monospezifische und nicht miteinander in Beziehung stehende Reagentien voraussetzen.

     

Are antibodies important in mice infected with Plasmodium yoelii?

It is unwise to extrapolate results, even from one mouse strain to another, when attempting to define the mechanisms that control and effect anti-malaria immunity. It is important to better characterize the broad range o f possible responses that are likely to occur when individuals in an outbred population are infected. Here, Peter Sayles and Donald Wossom discuss briefly their views on the role of antibody in murine and human malaria infections, based on their work on mice infected with Plasmodium yoelii.