Stefan N.,University of Zurich |
Stefan N.,University of Bern |
Stefan N.,NBE-Therapeutics |
Zimmermann M.,University of Zurich |
And 6 more authors.
Bioconjugate Chemistry | Year: 2014
Highly potent biotoxins like Pseudomonas exotoxin A (ETA) are attractive payloads for tumor targeting. However, despite replacement of the natural cell-binding domain of ETA by tumor-selective antibodies or alternative binding proteins like designed ankyrin repeat proteins (DARPins) the therapeutic window of such fusion toxins is still limited by target-independent cellular uptake, resulting in toxicity in normal tissues. Furthermore, the strong immunogenicity of the bacterial toxin precludes repeated administration in most patients. Site-specific modification to convert ETA into a prodrug-like toxin which is reactivated specifically in the tumor, and at the same time has a longer circulation half-life and is less immunogenic, is therefore appealing. To engineer a prodrug-like fusion toxin consisting of the anti-EpCAM DARPin Ec1 and a domain I-deleted variant of ETA (ETA″), we used strain-promoted azide alkyne cycloaddition for bioorthogonal conjugation of linear or branched polyethylene glycol (PEG) polymers at defined positions within the toxin moiety. Reversibility of the shielding was provided by a designed peptide linker containing the cleavage site for the rhinovirus 3C model protease. We identified two distinct sites, one within the catalytic domain and one close to the C-terminal KDEL sequence of Ec1-ETA″, simultaneous PEGylation of which resulted in up to 1000-fold lower cytotoxicity in EpCAM-positive tumor cells. Importantly, the potency of the fusion toxin was fully restored by proteolytic unveiling. Upon systemic administration in mice, PEGylated Ec1-ETA″ was much better tolerated than Ec1-ETA″; it showed a longer circulation half-life and an almost 10-fold increased area under the curve (AUC). Our strategy of engineering prodrug-like fusion toxins by bioorthogonal veiling opens new possibilities for targeting tumors with more specificity and efficacy. (Figure Presented). © 2014 American Chemical Society. Source
Breous-Nystrom E.,4 Antibody AG |
Schultze K.,4 Antibody AG |
Meier M.,4 Antibody AG |
Flueck L.,4 Antibody AG |
And 9 more authors.
Methods | Year: 2014
Over the last nearly three decades in vitro display technologies have played an important role in the discovery and optimization of antibodies and other proteins for therapeutic applications. Here we describe the use of retroviral expression technology for the display of full-length IgG on B lineage cells in vitro with a hallmark of a tight and stable genotype to phenotype coupling. We describe the creation of a high-diversity (>1.0E09 different heavy- and light-chain combinations) cell displayed fully human antibody library from healthy donor-derived heavy- and light-chain gene libraries, and demonstrate the recovery of high affinity target-specific antibodies from this library by staining of cells with a labeled target antigen and their magnetic- and flow cytometry-based cell sorting. The present technology represents a further evolution in the discovery of full-length, fully human antibodies using mammalian display, and is termed Retrocyte Display® (Retroviral B lympho. cyte Display). © 2013 Elsevier Inc. Source
NBE-Therapeutics | Entity website
NBE-Therapeutics Transpo-mAb Technology for fully human antibody development In addition to NBE-Therapeutics SMACTM-Technology for specific conjugation of toxic payloads to antibodies, NBE-Therapeutics has also developed a novel, patent-pending in vitro mammalian cell based expression and screening technology, called Transpo-mAb Display. This technology is based in the expression of complex libraries of full-length, fully human antibodies on the surface of precursor B cells mediated by stable DNA-Transposition ...
NBE-Therapeutics | Entity website
Additional infos NBE Therapeutics Technology Parc Basel Hochbergerstrasse 60C CH-4057 Basel Switzerland Phone: +41-61-633-2230 FAX: +41-61-633-2231 E-Mail: info@NBE-Therapeutics.com
NBE-Therapeutics | Date: 2013-07-18
The method disclosed herein describes a novel technology offering unparalleled efficiency, flexibility, utility and speed for the discovery and optimization of polypeptides having desired binding specificity and/or functionality, including antigen-binding molecules such as antibodies and fragments thereof, for desired functional and/or binding phenotypes. The novel method is based on transposable constructs and diverse DNA libraries cloned into transposable vectors and their transfection into host cells by concomitant transient expression of a functional transposase enzyme. This ensures an efficient, stable introduction of the transposon-based expression vectors into vertebrate host cells in one step, which can then be screened for a desired functional or binding phenotype of the expressed proteins, after which the relevant coding sequences for the expressed proteins, including antibodies and fragments thereof, can be identified by standard cloning and DNA sequencing techniques.