Human-like antibodies neutralizing Western equine encephalitis virus

Hülseweh, Birgit; Rülker, Torsten; Pelat, Thibaut; Langermann, Claudia; Frenzel, André; Schirrmann, Thomas; Dübel, Stefan; Thullier, Philippe; Hust, Michael

doi:10.4161/mabs.28170

Cited by 30 publications

(31 citation statements)

References 72 publications

(100 reference statements)

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“…In medical research, immune libraries are mostly used to discover antibodies against targets from infectious pathogens, e.g., antibodies neutralizing the human immunodeficiency virus (HIV) from long-term non-progressor patients, 48 West Nile virus 49 or immunized cancer patients. 50 Immune libraries can also be generated from immunized animals, 51-55 including macaques which have antibodies that are very similar to those of humans. 56 Use of transgenic animals containing human antibody gene repertoire allows the discovery of human antibody from such immune libraries.…”

Section: The Antibody Libraries Are “Gold Mines” For Therapeutic Antimentioning

confidence: 99%

Phage display-derived human antibodies in clinical development and therapy

Frenzel¹,

Schirrmann²,

Hust

2016

mAbs

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299

224

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Over the last 3 decades, monoclonal antibodies have become the most important class of therapeutic biologicals on the market. Development of therapeutic antibodies was accelerated by recombinant DNA technologies, which allowed the humanization of murine monoclonal antibodies to make them more similar to those of the human body and suitable for a broad range of chronic diseases like cancer and autoimmune diseases. In the early 1990s in vitro antibody selection technologies were developed that enabled the discovery of “fully” human antibodies with potentially superior clinical efficacy and lowest immunogenicity.Antibody phage display is the first and most widely used of the in vitro selection technologies. It has proven to be a robust, versatile platform technology for the discovery of human antibodies and a powerful engineering tool to improve antibody properties. As of the beginning of 2016, 6 human antibodies discovered or further developed by phage display were approved for therapy. In 2002, adalimumab (Humira®) became the first phage display-derived antibody granted a marketing approval. Humira® was also the first approved human antibody, and it is currently the best-selling antibody drug on the market. Numerous phage display-derived antibodies are currently under advanced clinical investigation, and, despite the availability of other technologies such as human antibody-producing transgenic mice, phage display has not lost its importance for the discovery and engineering of therapeutic antibodies.Here, we provide a comprehensive overview about phage display-derived antibodies that are approved for therapy or in clinical development. A selection of these antibodies is described in more detail to demonstrate different aspects of the phage display technology and its development over the last 25 years.

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Section: The Antibody Libraries Are “Gold Mines” For Therapeutic Antimentioning

confidence: 99%

Phage display-derived human antibodies in clinical development and therapy

Frenzel¹,

Schirrmann²,

Hust

2016

mAbs

Self Cite

299

224

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“…Due to the phylogenetic proximity between non-human primates (NPHs), such as macaques ( Macaca fascicularis ), and humans, the use of NHPs makes it possible to isolate human-like antibody fragments for therapeutic applications [ 30 ]. This strategy has been successfully used for the isolation of several antibody fragments with nanomolar or picomolar affinities against different antigens of clinical interest, such as Crf2, a surface antigen of Aspergillus fumigatus [ 31 ] , viruses like Western equine encephalitis virus (WEEV) and Venezuelan equine encephalitis virus (VEEV) [ 32 , 33 ] and several toxins, such as tetanus toxin [ 34 ], ricin [ 35 ] and the two sub-units of anthrax lethal toxin [ 36 , 37 ]. In a previous study we isolated several recombinant antibodies targeting the light chain of BoNT/A, using a macaque immune library, which neutralize the toxicity of BoNT/A in an ex vivo mouse phrenic nerve diaphragm assay [ 38 ].…”

Section: Introductionmentioning

confidence: 99%

Development of Human-Like scFv-Fc Neutralizing Botulinum Neurotoxin E

Miethe

Rasetti-Escargueil

Avril³

et al. 2015

PLoS ONE

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BackgroundBotulinum neurotoxins (BoNTs) are considered to be the most toxic substances known on earth and are responsible for human botulism, a life-threatening disease characterized by flaccid muscle paralysis that occurs naturally by food-poisoning or colonization of the gastrointestinal tract by BoNT-producing clostridia. BoNTs have been classified as category A agent by the Centers of Disease Control and Prevention (CDC) and are listed among the six agents with the highest risk to be used as bioweapons. Neutralizing antibodies are required for the development of effective anti-botulism therapies to deal with the potential risk of exposure.ResultsIn this study, a macaque (Macaca fascicularis) was immunized with recombinant light chain of BoNT/E3 and an immune phage display library was constructed. After a multi-step panning, several antibody fragments (scFv, single chain fragment variable) with nanomolar affinities were isolated, that inhibited the endopeptidase activity of pure BoNT/E3 in vitro by targeting its light chain. Furthermore, three scFv were confirmed to neutralize BoNT/E3 induced paralysis in an ex vivo mouse phrenic nerve-hemidiaphragm assay. The most effective neutralization (20LD50/mL, BoNT/E3) was observed with scFv ELC18, with a minimum neutralizing concentration at 0.3 nM. Furthermore, ELC18 was highly effective in vivo when administered as an scFv-Fc construct. Complete protection of 1LD50 BoNT/E3 was observed with 1.6 ng/dose in the mouse flaccid paralysis assay.ConclusionThese scFv-Fcs antibodies are the first recombinant antibodies neutralizing BoNT/E by targeting its light chain. The human-like nature of the isolated antibodies is predicting a good tolerance for further clinical development.

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“…Since macaque VH and VL sequences have an extremely high homology to their human counterparts [34], they are a more obvious choice for the development of therapeutic antibodies than rodents. They have successfully been used to generate antibodies against the most dangerous toxins, like ricin [35], anthrax lethal factor [36], or various botulinum toxins [37,38] but also against fungi [39], encephalitis viruses [40], or human proteins [41]. The last example also demonstrates the very high specificity achievable from phage display by design, since the discovered antibody is able to discriminate between very subtle conformation differences of Alzheimer peptide aggregates.…”

Section: Phage Display Is a Robust And Reliable Source For Human Monomentioning

confidence: 97%

Designing Human Antibodies by Phage Display

Frenzel

Kügler²,

Helmsing

et al. 2017

Transfus Med Hemother

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ever, are recognized as foreign proteins by humans and therefore induce immune reactions against the therapeutic reagent (human anti-mouse antibody; HAMA) [3]. A solution to this problem was only brought by recombinant DNA technology. Using genetic engineering, the murine sequences were exchanged in parts of the molecule by the homologous human sequences. The result were 'chimeric' (with murine constant domains replaced by human constant domains) or 'humanized' antibodies (only the antigen-binding loops / complementarity determining regions (CDRs) were transferred to human variable region frameworks) [4,5]. These engineered mouse antibodies dominated the first decade of therapeutic antibody approvals, while today it is preferred to use completely human antibodies from the start, thanks to a number of novel technologies allowing to identify them without the necessity to immunize humans. These are on the one hand transgenic animals, typically mice or rats, which carry genetically introduced human immunoglobulin loci while their own antibody genes were knocked out [6][7][8]. Consequently, after immunization they utilize this sequence repertoire to generate IgG from human germ-line sequences, thereby allowing to generate monoclonals by means of classical hybridoma technology. An even more radical way was introduced by the development of antibody phage display, which allowed for the first time to generate human antibodies completely in the test tube and without immunization. Inspired by the work of G.P. Smith on peptide display using filamentous phage [9], antibody phage display was development independently in Heidelberg by Breitling and Dübel [10], in Cambridge by McCafferty et al. [11], and in California by Barbas et al. [12]. The method is based on a physical link between function (antigen binding) and information (antibody gene) in a nanoparticle (the phage virus particle). To achieve this, the antigen binding parts of the antibodies, either as Fab (fragment antigen binding) [13,14] or scFv (single chain fragment variable) [15][16][17], are genetically linked to the surface protein III (pIII) of the M13 phage and thus expressed on the surface of the virus particle. Mixtures of such phage particles, each enKeywords Human monoclonal antibodies · Phage display · Immunotherapy · In vitro evolution · Panning · scFv Summary With six approved products and more than 60 candidates in clinical testing, human monoclonal antibody discovery by phage display is well established as a robust and reliable source for the generation of therapeutic antibodies. While a vast diversity of library generation philosophies and selection strategies have been conceived, the power of molecular design offered by controlling the in vitro selection step is still to be recognized by a broader audience outside of the antibody engineering community. Here, we summarize some opportunities and achievements, e.g., the generation of antibodies which could not be generated otherwise, and the design of antibody properties by different panning strategie...

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Human-like antibodies neutralizing Western equine encephalitis virus

Cited by 30 publications

References 72 publications

Phage display-derived human antibodies in clinical development and therapy

Phage display-derived human antibodies in clinical development and therapy

Development of Human-Like scFv-Fc Neutralizing Botulinum Neurotoxin E

Designing Human Antibodies by Phage Display

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