| 1. |
- Bonvicini, Gillian, et al.
(författare)
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ImmunoPET imaging of amyloid-beta in a rat model of Alzheimer's disease with a bispecific, brain-penetrating fusion protein
- 2022
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Ingår i: Translational Neurodegeneration. - : BioMed Central (BMC). - 2047-9158. ; 11
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Tidskriftsartikel (refereegranskat)abstract
- Background: Hijacking the transferrin receptor (TfR) is an effective strategy to transport amyloid-beta (A beta) immuno-positron emission tomography (immunoPET) ligands across the blood-brain barrier (BBB). Such ligands are more sensitive and specific than small-molecule ligands at detecting A beta pathology in mouse models of Alzheimer's disease (AD). This study aimed to determine if this strategy would be as sensitive in rats and to assess how TfR affinity affects BBB transport of bispecific immunoPET radioligands.Methods: Two affinity variants of the rat TfR antibody, OX26, were chemically conjugated to a F(ab')(2) fragment of the anti-A beta antibody, bapineuzumab (Bapi), to generate two bispecific fusion proteins: OX26(5)-F(ab')(2)-Bapi and OX26(76)-F(ab')(2)-Bapi. Pharmacokinetic analyses were performed 4 h and 70 h post-injection of radioiodinated fusion proteins in wild-type (WT) rats. [I-124]I-OX26(5)-F(ab')(2)-Bapi was administered to TgF344-AD and WT rats for in vivo PET imaging. Ex vivo distribution of injected [I-124]I-OX26(5)-F(ab')(2)-Bapi and A beta pathology were assessed.Results: More [I-125]I-OX26(5)-F(ab')(2)-Bapi was taken up into the brain 4 h post-administration than [I-124]I-OX26(76)-F(ab')(2)-Bapi. [I-124]I-OX26(5)-F(ab')(2)-Bapi PET visualized A beta pathology with significantly higher signals in the TgF344-AD rats than in the WT littermates without A beta pathology. The PET signals significantly correlated with A beta levels in AD animals.Conclusion: Affinity to TfR affects how efficiently a TfR-targeting bispecific fusion protein will cross the BBB, such that the higher-affinity bispecific fusion protein crossed the BBB more efficiently. Furthermore, bispecific immunoPET imaging of brain A beta pathology using TfR-mediated transport provides good imaging contrast between TgF344-AD and WT rats, suggesting that this immunoPET strategy has the potential to be translated to higher species.
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| 2. |
- Gustavsson, Tobias, et al.
(författare)
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Long-term effects of immunotherapy with a brain penetrating Aβ antibody in a mouse model of Alzheimer's disease
- 2023
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Ingår i: Alzheimer's Research & Therapy. - : BioMed Central (BMC). - 1758-9193. ; 15:1
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundBrain-directed immunotherapy is a promising strategy to target amyloid-β (Aβ) deposits in Alzheimer’s disease (AD). In the present study, we compared the therapeutic efficacy of the Aβ protofibril targeting antibody RmAb158 with its bispecific variant RmAb158-scFv8D3, which enters the brain by transferrin receptor-mediated transcytosis.MethodsAppNL−G−F knock-in mice received RmAb158, RmAb158-scFv8D3, or PBS in three treatment regimens. First, to assess the acute therapeutic effect, a single antibody dose was given to 5 months old AppNL−G−F mice, with evaluation after 3 days. Second, to assess the antibodies’ ability to halt the progression of Aβ pathology, 3 months old AppNL−G−F mice received three doses during a week, with evaluation after 2 months. Reduction of RmAb158-scFv8D3 immunogenicity was explored by introducing mutations in the antibody or by depletion of CD4+ T cells. Third, to study the effects of chronic treatment, 7-month-old AppNL−G−F mice were CD4+ T cell depleted and treated with weekly antibody injections for 8 weeks, including a final diagnostic dose of [125I]RmAb158-scFv8D3, to determine its brain uptake ex vivo. Soluble Aβ aggregates and total Aβ42 were quantified with ELISA and immunostaining.ResultsNeither RmAb158-scFv8D3 nor RmAb158 reduced soluble Aβ protofibrils or insoluble Aβ1-42 after a single injection treatment. After three successive injections, Aβ1-42 was reduced in mice treated with RmAb158, with a similar trend in RmAb158-scFv8D3-treated mice. Bispecific antibody immunogenicity was somewhat reduced by directed mutations, but CD4+ T cell depletion was used for long-term therapy. CD4+ T cell-depleted mice, chronically treated with RmAb158-scFv8D3, showed a dose-dependent increase in blood concentration of the diagnostic [125I]RmAb158-scFv8D3, while concentration was low in plasma and brain. Chronic treatment did not affect soluble Aβ aggregates, but a reduction in total Aβ42 was seen in the cortex of mice treated with both antibodies.ConclusionsBoth RmAb158 and its bispecific variant RmAb158-scFv8D3 achieved positive effects of long-term treatment. Despite its ability to efficiently enter the brain, the benefit of using the bispecific antibody in chronic treatment was limited by its reduced plasma exposure, which may be a result of interactions with TfR or the immune system. Future research will focus in new antibody formats to further improve Aβ immunotherapy.
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| 3. |
- Bonvicini, Gillian, et al.
(författare)
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Comparing in vitro affinity measurements of antibodies to TfR1 : Surface plasmon resonance versus on-cell affinity
- 2024
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Ingår i: Analytical Biochemistry. - : Elsevier. - 0003-2697 .- 1096-0309. ; 686
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Tidskriftsartikel (refereegranskat)abstract
- Despite years of utilizing the transferrin receptor 1 (TfR1) to transport large biomolecules into the brain, there is no consensus on how to optimally measure affinity to it. The aim of this study was to compare different methods for measuring the affinities of anti-TfR1 antibodies.Antibodies 15G11, OX26 and 8D3 are known to successfully carry large biologics across the blood-brain barrier in humans, rats, and mice, respectively. The affinity to their respective species of TfR1 was measured with different surface plasmon resonance setups in Biacore and an on-cell assay.When the antibody was captured and TfR1 was the analyte, the dissociation in Biacore was very slow. The dissociation was faster when the antibody was the analyte and TfR1 was the ligand. The Biacore setup with capture of N-terminal FLAG-tag TfR1 yielded the most similar apparent affinities as the cell assay.In conclusion, it is important to evaluate assay parameters including assay orientation, surface capture method, and antibody format when comparing binding kinetics for TfR1 antibodies. Although it seems possible to determine relative affinities of TfR1 antibodies using the methods described here, both the FLAG-tag TfR1 capture setup and cell assays likely yield apparent affinities that are most translatable in vivo.
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| 4. |
- Oroujeni, Maryam, PhD, 1982-, et al.
(författare)
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The Use of a Non-Conventional Long-Lived Gallium Radioisotope Ga-66 Improves Imaging Contrast of EGFR Expression in Malignant Tumours Using DFO-ZEGFR:2377 Affibody Molecule
- 2021
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Ingår i: Pharmaceutics. - : MDPI AG. - 1999-4923. ; 13:2
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Tidskriftsartikel (refereegranskat)abstract
- Epidermal growth factor receptor (EGFR) is overexpressed in many malignancies. EGFR-targeted therapy extends survival of patients with disseminated cancers. Radionuclide molecular imaging of EGFR expression would make EGFR-directed treatment more personalized and therefore more efficient. A previous study demonstrated that affibody molecule [Ga-68]Ga-DFO-ZEGFR:2377 permits specific positron-emission tomography (PET) imaging of EGFR expression in xenografts at 3 h after injection. We anticipated that imaging at 24 h after injection would provide higher contrast, but this is prevented by the short half-life of Ga-68 (67.6 min). Here, we therefore tested the hypothesis that the use of the non-conventional long-lived positron emitter Ga-66 (T-1/2 = 9.49 h, beta(+) = 56.5%) would permit imaging with higher contrast. Ga-66 was produced by the Zn-66(p,n)Ga-66 nuclear reaction and DFO-ZEGFR:2377 was efficiently labelled with Ga-66 with preserved binding specificity in vitro and in vivo. At 24 h after injection, [Ga-66]Ga-DFO-ZEGFR:2377 provided 3.9-fold higher tumor-to-blood ratio and 2.3-fold higher tumor-to-liver ratio than [Ga-68]Ga-DFO-ZEGFR:2377 at 3 h after injection. At the same time point, [Ga-66]Ga-DFO-ZEGFR:2377 provided 1.8-fold higher tumor-to-blood ratio, 3-fold higher tumor-to-liver ratio, 1.9-fold higher tumor-to-muscle ratio and 2.3-fold higher tumor-to-bone ratio than [Zr-89]Zr-DFO-ZEGFR:2377. Biodistribution data were confirmed by whole body PET combined with magnetic resonance imaging (PET/MRI). The use of the positron emitter Ga-66 for labelling of DFO-ZEGFR:2377 permits PET imaging of EGFR expression at 24 h after injection and improves imaging contrast.
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| 5. |
- 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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