| 1. |
- Brinson, Robert G., et al.
(författare)
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Enabling adoption of 2D-NMR for the higher order structure assessment of monoclonal antibody therapeutics
- 2019
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Ingår i: mAbs. - : Informa UK Limited. - 1942-0862 .- 1942-0870. ; 11:1, s. 94-105
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Tidskriftsartikel (refereegranskat)abstract
- The increased interest in using monoclonal antibodies (mAbs) as a platform for biopharmaceuticals has led to the need for new analytical techniques that can precisely assess physicochemical properties of these large and very complex drugs for the purpose of correctly identifying quality attributes (QA). One QA, higher order structure (HOS), is unique to biopharmaceuticals and essential for establishing consistency in biopharmaceutical manufacturing, detecting process-related variations from manufacturing changes and establishing comparability between biologic products. To address this measurement challenge, two-dimensional nuclear magnetic resonance spectroscopy (2D-NMR) methods were introduced that allow for the precise atomic-level comparison of the HOS between two proteins, including mAbs. Here, an inter-laboratory comparison involving 26 industrial, government and academic laboratories worldwide was performed as a benchmark using the NISTmAb, from the National Institute of Standards and Technology (NIST), to facilitate the translation of the 2D-NMR method into routine use for biopharmaceutical product development. Two-dimensional H-1,N-15 and H-1,C-13 NMR spectra were acquired with harmonized experimental protocols on the unlabeled Fab domain and a uniformly enriched-N-15, 20%-C-13-enriched system suitability sample derived from the NISTmAb. Chemometric analyses from over 400 spectral maps acquired on 39 different NMR spectrometers ranging from 500 MHz to 900 MHz demonstrate spectral fingerprints that are fit-for-purpose for the assessment of HOS. The 2D-NMR method is shown to provide the measurement reliability needed to move the technique from an emerging technology to a harmonized, routine measurement that can be generally applied with great confidence to high precision assessments of the HOS of mAb-based biotherapeutics.
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| 2. |
- Malm, Magdalena, et al.
(författare)
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Targeting HER3 using mono- and bispecific antibodies or alternative scaffolds
- 2016
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Ingår i: mAbs. - : Taylor & Francis. - 1942-0862 .- 1942-0870. ; 8:7, s. 1195-1209
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Tidskriftsartikel (refereegranskat)abstract
- The human epidermal growth factor receptor 3 (HER3) has in recent years been recognized as a key node in the complex signaling network of many different cancers. It is implicated in de novo and acquired resistance against therapies targeting other growth factor receptors, e.g., EGFR, HER2, and it is a major activator of the PI3K/Akt signaling pathway. Consequently, HER3 has attracted substantial attention, and is today a key target for drugs in clinical development. Sophisticated protein engineering approaches have enabled the generation of a range of different affinity proteins targeting this receptor, including antibodies and alternative scaffolds that are either mono- or bispecific. Here, we describe HER3 and its role as a key tumor target, and give a comprehensive review of HER3-targeted proteins currently in development, including discussions on the opportunities and challenges of targeting this receptor.
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| 3. |
- Scheffel, Julia, et al.
(författare)
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Optimization of a calcium-dependent Protein A-derived domain for mild antibody purification
- 2019
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Ingår i: mAbs. - : TAYLOR & FRANCIS INC. - 1942-0862 .- 1942-0870.
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Tidskriftsartikel (refereegranskat)abstract
- As reported here, we developed and optimized a purification matrix based on a Protein A-derived domain, Z(Ca), displaying calcium-dependent antibody binding. It provides an alternative to the acidic elution conditions of conventional Protein A affinity chromatography for purification of sensitive antibodies and other Fc-based molecules. We describe the multimerization of Z(Ca) to generate a chromatography resin with higher binding capacity. The highest order multimeric variant, Z(Ca)TetraCys, demonstrated a considerably high dynamic binding capacity (35 mg IgG/ml resin) while preserving the specificity for IgG. High recovery was obtained and host cell protein and DNA content in purified fractions proved to be comparable to commercial MabSelect SuRe and MabSelect PrismA. Various elution conditions for use of this domain in antibody purification were investigated. The purification data presented here revealed variations in the interaction of different subclasses of human IgG with Z(Ca)TetraCys. This resulted in diverse elution properties for the different IgGs, where complete elution of all captured antibody for IgG2 and IgG4 was possible at neutral pH. This optimized protein ligand and the proposed purification method offer a unique strategy for effective and mild purification of antibodies and Fc-fusion proteins that cannot be purified under conventional acidic elution conditions due to aggregation formation or loss of function.
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| 4. |
- Sustmann, Claudio, et al.
(författare)
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DuoMab : a novel CrossMab-based IgG-derived antibody format for enhanced antibody-dependent cell-mediated cytotoxicity
- 2019
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Ingår i: mAbs. - : Informa UK Limited. - 1942-0862 .- 1942-0870. ; 11:8, s. 1402-1414
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Tidskriftsartikel (refereegranskat)abstract
- High specificity accompanied with the ability to recruit immune cells has made recombinant therapeutic antibodies an integral part of drug development. Here we present a generic approach to generate two novel IgG-derived antibody formats that are based on a modification of the CrossMab technology. MoAbs harbor two heavy chains (HCs) resulting in one binding entity and one fragment crystallizable region (Fc), whereas DuoMabs are composed of four HCs harboring two binding entities and two Fc regions linked at a disulfide-bridged hinge. The latter bivalent format is characterized by avidity-enhanced target cell binding while simultaneously increasing the ‘Fc-load’ on the surface. DuoMabs were shown to be producible in high yield and purity and bind to surface cells with affinities comparable to IgGs. The increased Fc load directed at the surface of target cells by DuoMabs modulates their antibody-dependent cell-mediated cytotoxicity competency toward target cells, making them attractive for applications that require or are modulated by FcR interactions.
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