| 1. |
- Bondza, Sina, et al.
(författare)
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Bivalent binding on cells varies between anti-CD20 antibodies and is dose-dependent
- 2020
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Ingår i: mAbs. - : TAYLOR & FRANCIS INC. - 1942-0862 .- 1942-0870. ; 12:1
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Tidskriftsartikel (refereegranskat)abstract
- Based on their mechanism of action, two types of anti-CD20 antibodies are distinguished: Type I, which efficiently mediate complement-dependent cytotoxicity, and Type II, which instead are more efficient in inducing direct cell death. Several molecular characteristics of these antibodies have been suggested to underlie these different biological functions, one of these being the manner of binding to CD20 expressed on malignant B cells. However, the exact binding model on cells is unclear. In this study, the binding mechanism of the Type I therapeutic antibodies rituximab (RTX) and ofatumumab (OFA) and the Type II antibody obinutuzumab (OBI) were established by real-time interaction analysis on live cells. It was found that the degree of bivalent stabilization differed for the antibodies: OFA was stabilized the most, followed by RTX and then OBI, which had the least amount of bivalent stabilization. Bivalency inversely correlated with binding dynamics for the antibodies, with OBI displaying the most dynamic binding pattern, followed by RTX and OFA. For RTX and OBI, bivalency and binding dynamics were concentration dependent; at higher concentrations the interactions were more dynamic, whereas the percentage of antibodies that bound bivalent was less, resulting in concentration-dependent apparent affinities. This was barely noticeable for OFA, as almost all molecules bound bivalently at the tested concentrations. We conclude that the degree of bivalent binding positively correlates with the complement recruiting capacity of the investigated CD20 antibodies.
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| 2. |
- Klint, Susanne, et al.
(författare)
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Izokibep : Preclinical development and first-in-human study of a novel IL-17A neutralizing Affibody molecule in patients with plaque psoriasis
- 2023
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Ingår i: mAbs. - : Taylor & Francis. - 1942-0862 .- 1942-0870. ; 15:1
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Tidskriftsartikel (refereegranskat)abstract
- Psoriasis, an immune-mediated inflammatory disease, affects nearly 125 million people globally. The interleukin (IL)-17A homodimer is a key driver of psoriasis and other autoimmune diseases, including psoriatic arthritis, axial spondyloarthritis, hidradenitis suppurativa, and uveitis. Treatment with monoclonal antibodies (mAbs) against IL-17A provides an improvement in the Psoriasis Area and Severity Index compared to conventional systemic agents. In this study, the Affibody(CIRCLED LATIN CAPITAL LETTER R) technology was used to identify and optimize a novel, small, biological molecule comprising three triple helical affinity domains, izokibep (previously ABY-035), for the inhibition of IL-17A signaling. Preclinical studies show that izokibep, a small 18.6 kDa IL-17 ligand trap comprising two IL-17A-specific Affibody domains and one albumin-binding domain, selectively inhibits human IL-17A in vitro and in vivo with superior potency and efficacy relative to anti-IL-17A mAbs. A Phase 1 first-in-human study was conducted to establish the safety, pharmacokinetics, and preliminary efficacy of izokibep, when administered intravenously and subcutaneously as single doses to healthy subjects, and as single intravenous and multiple subcutaneous doses to patients with psoriasis (NCT02690142; EudraCT No: 2015-004531-13). Izokibep was well tolerated with no meaningful safety concerns identified in healthy volunteers and patients with psoriasis. Rapid efficacy was seen in all psoriasis patients after one dose which further improved in patients receiving multiple doses. A therapeutic decrease in joint pain was also observed in a single patient with concurrent psoriatic arthritis. The study suggests that izokibep has the potential to safely treat IL17A-associated diseases such as psoriasis, psoriatic arthritis, axial spondyloarthritis, hidradenitis suppurativa, and uveitis.
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| 3. |
- Mega, Alessandro, et al.
(författare)
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A PDGFRB- and CD40-targeting bispecific AffiMab induces stroma-targeted immune cell activation
- 2023
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Ingår i: mAbs. - : Informa UK Limited. - 1942-0862 .- 1942-0870. ; 15:1
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Tidskriftsartikel (refereegranskat)abstract
- CD40 agonism by systemic administration of CD40 monoclonal antibodies has been explored in clinical trials for immunotherapy of cancer, uncovering enormous potential, but also dosing challenges in terms of systemic toxicity. CD40-dependent activation of antigen presenting cells is dependent on crosslinking of the CD40 receptor. Here we exploited this requisite by coupling crosslinking to cancer-receptor density by dual-targeting of CD40 and platelet-derived growth factor receptor beta (PDGFRB), which is highly expressed in the stroma of various types of tumors. A novel PDGFRBxCD40 Fc-silenced bispecific AffiMab was developed to this end to test whether it is possible to activate CD40 in a PDGFRB-targeted manner. A PDGFRB-binding Affibody molecule was fused to each heavy chain of an Fc-silenced CD40 agonistic monoclonal antibody to obtain a bispecific "AffiMab". Binding of the AffiMab to both PDGFRB and CD40 was confirmed by surface plasmon resonance, bio-layer interferometry and flow cytometry, through analysis of cells expressing respective target. In a reporter assay, the AffiMab displayed increased CD40 potency in the presence of PDGFRB-conjugated beads, in a manner dependent on PDGFRB amount/bead. To test the concept in immunologically relevant systems with physiological levels of CD40 expression, the AffiMab was tested in human monocyte-derived dendritic cells (moDCs) and B cells. Expression of activation markers was increased in moDCs specifically in the presence of PDGFRB-conjugated beads upon AffiMab treatment, while the Fc-silenced CD40 mAb did not stimulate CD40 activation. As expected, the AffiMab did not activate moDCs in the presence of unconjugated beads. Finally, in a co-culture experiment, the AffiMab activated moDCs and B cells in the presence of PDGFRB-expressing cells, but not in co-cultures with PDGFRB-negative cells. Collectively, these results suggest the possibility to activate CD40 in a PDGFRB-targeted manner in vitro. This encourages further investigation and the development of such an approach for the treatment of solid cancers.
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| 4. |
- Nyesiga, Barnabas, et al.
(författare)
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RUBY® - a tetravalent (2+2) bispecific antibody format with excellent functionality and IgG-like stability, pharmacology and developability properties
- 2024
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Ingår i: mAbs. - : Taylor & Francis. - 1942-0862 .- 1942-0870. ; 16:1
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Tidskriftsartikel (refereegranskat)abstract
- Despite the large number of existing bispecific antibody (bsAb) formats, the generation of novel bsAbs is still associated with development and bioprocessing challenges. Here, we present RUBY, a novel bispecific antibody format that allows rapid generation of bsAbs that fulfill key development criteria. The RUBYTM format has a 2 + 2 geometry, where two Fab fragments are linked via their light chains to the C-termini of an IgG, and carries mutations for optimal chain pairing. The unique design enables generation of bsAbs with mAb-like attributes. Our data demonstrate that RUBY bsAbs are compatible with small-scale production systems for screening purposes and can be produced at high yields (>3 g/L) from stable cell lines. The bsAbs produced are shown to, in general, contain low amounts of aggregates and display favorable solubility and stress endurance profiles. Further, compatibility with various IgG isotypes is shown and tailored Fc gamma receptor binding confirmed. Also, retained interaction with FcRn is demonstrated to translate into a pharmacokinetic profile in mice and non-human primates that is comparable to mAb controls. Functionality of conditional active RUBY bsAbs is confirmed in vitro. Anti-tumor effects in vivo have previously been demonstrated, and shown to be superior to a comparable mAb, and here it is further shown that RUBY bsAbs penetrate and localize to tumor tissue in vivo. In all, the RUBY format has attractive mAb-like attributes and offers the possibility to mitigate many of the development challenges linked to other bsAb formats, facilitating both high functionality and developability.
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| 5. |
- Petersen, Inga, et al.
(författare)
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Multivalent design of the monoclonal SynO2 antibody improves binding strength to soluble α-Synuclein aggregates
- 2023
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Ingår i: mAbs. - : Taylor & Francis. - 1942-0862 .- 1942-0870. ; 15:1
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Tidskriftsartikel (refereegranskat)abstract
- Soluble aggregates are reported to be the most neurotoxic species of alpha-Synuclein (alpha Syn) in Parkinson's disease (PD) and hence are a promising target for diagnosis and treatment of PD. However, the predominantly intracellular location of alpha Syn limits its accessibility, especially for antibody-based molecules and prompts the need for exceptionally strong soluble alpha Syn aggregate binders to enhance their sensitivity and efficacy for targeting the extracellular alpha Syn pool. In this study, we have created the multivalent antibodies TetraSynO2 and HexaSynO2, derived from the alpha Syn oligomer-specific antibody SynO2, to increase avidity binding to soluble alpha Syn aggregate species through more binding sites in close proximity. The multivalency was achieved through recombinant fusion of single-chain variable fragments of SynO2 to the antibodies' original N-termini. Our ELISA results indicated a 20-fold increased binding strength of the multivalent formats to alpha Syn aggregates, while binding to alpha Syn monomers and unspecific binding to amyloid beta protofibrils remained low. Kinetic analysis using LigandTracer revealed that only 80% of SynO2 bound bivalently to soluble aSyn aggregates, whereas the proportion of TetraSynO2 and HexaSynO2 binding bi- or multivalently to soluble alpha Syn aggregates was increased to similar to 95% and 100%, respectively. The overall improved binding strength of TetraSynO2 and HexaSynO2 implies great potential for immunotherapeutic and diagnostic applications with targets of limited accessibility, like extra-cellular alpha Syn aggregates. The ability of the multivalent antibodies to bind a wider range of alpha Syn aggregate species, which are not targetable by conventional bivalent antibodies, thus could allow for an earlier and more effective intervention in the progression of PD.
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| 6. |
- Schlein, Eva, et al.
(författare)
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Reducing neonatal Fc receptor binding enhances clearance and brain-to-blood ratio of TfR-delivered bispecific amyloid-β antibody
- 2024
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Ingår i: mAbs. - : Taylor & Francis. - 1942-0862 .- 1942-0870. ; 16:1
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Tidskriftsartikel (refereegranskat)abstract
- Recent development of amyloid-β (Aβ)-targeted immunotherapies for Alzheimer’s disease (AD) have highlighted the need for accurate diagnostic methods. Antibody-based positron emission tomography (PET) ligands are well suited for this purpose as they can be directed toward the same target as the therapeutic antibody. Bispecific, brain-penetrating antibodies can achieve sufficient brain concentrations, but their slow blood clearance remains a challenge, since it prolongs the time required to achieve a target-specific PET signal. Here, two antibodies were designed based on the Aβ antibody bapineuzumab (Bapi) – one monospecific IgG (Bapi) and one bispecific antibody with an antigen binding fragment (Fab) of the transferrin receptor (TfR) antibody 8D3 fused to one of the heavy chains (Bapi-Fab8D3) for active, TfR-mediated transport into the brain. A variant of each antibody was designed to harbor a mutation to the neonatal Fc receptor (FcRn) binding domain, to increase clearance. Blood and brain pharmacokinetics of radiolabeled antibodies were studied in wildtype (WT) and AD mice (AppNL-G-F). The FcRn mutation substantially reduced blood half-life of both Bapi and Bapi-Fab8D3. Bapi-Fab8D3 showed high brain uptake and the brain-to-blood ratio of its FcRn mutated form was significantly higher in AppNL-G-F mice than in WT mice 12 h after injection and increased further up to 168 h. Ex vivo autoradiography showed specific antibody retention in areas with abundant Aβ pathology. Taken together, these results suggest that reducing FcRn binding of a full-sized bispecific antibody increases the systemic elimination and could thereby drastically reduce the time from injection to in vivo imaging.
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| 7. |
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| 8. |
- Voskuil, Jan L. A., et al.
(författare)
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The Antibody Society's antibody validation webinar series
- 2020
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Ingår i: mAbs. - : Informa UK Limited. - 1942-0862 .- 1942-0870. ; 12:1
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Tidskriftsartikel (refereegranskat)abstract
- In the wake of the reproducibility crisis and numerous discussions on how commercially available antibodies as research tool contribute to it, The Antibody Society developed a series of 10 webinars to address the issues involved. The webinars were delivered by speakers with both academic and commercial backgrounds. This report highlights the problems, and offers solutions to help the scientific community appropriately identify the right antibodies and to validate them for their research and development projects. Despite the various solutions proposed here, they must be applied on a case-by-case basis. Each antibody must be verified based on the content of the product sheet, and subsequently through experimentation to confirm integrity, specificity and selectivity. Verification needs to focus on the precise application and tissue/cell type for which the antibody will be used, and all verification data must be reported openly. The various approaches discussed here all have caveats, so a combination of solutions must be considered.
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