Extended plasma half-life of albumin-binding domain fused human IgA upon pH-dependent albumin engagement of human FcRn in vitro and in vivo

AbstractGranulocyte colony stimulating factor (GCSF) can decrease mortality of patients undergo chemotherapy through increasing neutrophil counts. Many strategies have been developed to improve its blood circulating time. Albumin binding domain (ABD) was genetically fused to N-terminal end of GCSF encoding sequence and expressed as cytoplasmic inclusion bodies within Escherichia coli. Biological activity of ABD-GCSF protein was assessed by proliferation assay on NFS-60 cells. Physicochemical properties were analyzed through size exclusion chromatography, circular dichroism, intrinsic fluorescence spectroscopy and dynamic light scattering. Pharmacodynamics and pharmacokinetic properties were also investigated in a neutropenic rat model. CD and IFS spectra revealed that ABD fusion to GCSF did not significantly affect the secondary and tertiary structures of the molecule. DLS and SEC results indicated the absence of aggregation formation. EC50 value of the ABD-GCSF in proliferation of NFS-60 cells was 75.76 pg/ml after 72 h in comparison with control GCSF molecules (Filgrastim: 73.1 pg/ml and PEG-Filgrastim: 44.6 pg/ml). Animal studies of ABD-GCSF represented improved serum half-life (9.3 ± 0.7 h) and consequently reduced renal clearance (16.1 ± 1.4 ml/h.kg) in comparison with Filgrastim (1.7 ± 0.1 h). Enhanced neutrophils count following administration of ABD-GCSF was comparable with Filgrastim and weaker than PEG-Filgrastim treated rats. In vitro and in vivo results suggested the ABD fusion as a potential approach for improving GCSF properties.

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The Influence of Domain Permutations of an Albumin-Binding Domain-Fused HER2-Targeting Affibody-Based Drug Conjugate on Tumor Cell Proliferation and Therapy Efficacy

Pharmaceutics ◽

10.3390/pharmaceutics13111974 ◽

2021 ◽

Vol 13 (11) ◽

pp. 1974

Author(s):

Wen Yin ◽

Tianqi Xu ◽

Mohamed Altai ◽

Maryam Orougeni ◽

Jie Zhang ◽

...

Keyword(s):

Cell Proliferation ◽

Tumor Cell ◽

Targeted Delivery ◽

Binding Domain ◽

Albumin Binding ◽

Tumor Cell Proliferation ◽

Drug Conjugate ◽

Therapy Efficacy

Human epidermal growth factor receptor 2 (HER2) is a clinically validated target for breast cancer therapy. Previously, a drug-fused HER2-targeting affinity protein construct successfully extended the survival of mice bearing HER2-expressing xenografts. The aim of this study was to evaluate the influence of the number and positioning of the protein domains in the drug conjugate. Seven HER2-targeting affibody-based constructs, including one or two affibody molecules (Z) with or without an albumin-binding domain (ABD), namely Z, Z-ABD, ABD-Z, Z-Z, Z-Z-ABD, Z-ABD-Z, and ABD-Z-Z, were evaluated on their effects on cell growth, in vivo targeting, and biodistribution. The biodistribution study demonstrated that the monomeric constructs had longer blood retention and lower hepatic uptake than the dimeric ones. A dimeric construct, specifically ABD-Z-Z, could stimulate the proliferation of HER2 expressing SKOV-3 cells in vitro and the growth of tumors in vivo, whereas the monomeric construct Z-ABD could not. These two constructs demonstrated a therapeutic effect when coupled to mcDM1; however, the effect was more pronounced for the non-stimulating Z-ABD. The median survival of the mice treated with Z-ABD-mcDM1 was 63 days compared to the 37 days for those treated with ABD-Z-Z-mcDM1 or for the control animals. Domain permutation of an ABD-fused HER2-targeting affibody-based drug conjugate significantly influences tumor cell proliferation and therapy efficacy. The monomeric conjugate Z-ABD is the most promising format for targeted delivery of the cytotoxic drug DM1.

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HER2-Specific Pseudomonas Exotoxin A PE25 Based Fusions: Influence of Targeting Domain on Target Binding, Toxicity, and In Vivo Biodistribution

Pharmaceutics ◽

10.3390/pharmaceutics12040391 ◽

2020 ◽

Vol 12 (4) ◽

pp. 391

Author(s):

Haozhong Ding ◽

Mohamed Altai ◽

Wen Yin ◽

Sarah Lindbo ◽

Hao Liu ◽

...

Keyword(s):

Half Life ◽

Binding Domain ◽

Protein Domain ◽

Specific Cell ◽

Albumin Binding ◽

Exotoxin A ◽

Pseudomonas Exotoxin ◽

Pseudomonas Exotoxin A ◽

Number Of Patients ◽

Plasma Half Life

The human epidermal growth factor receptor 2 (HER2) is a clinically validated target for cancer therapy, and targeted therapies are often used in regimens for patients with a high HER2 expression level. Despite the success of current drugs, a number of patients succumb to their disease, which motivates development of novel drugs with other modes of action. We have previously shown that an albumin binding domain-derived affinity protein with specific affinity for HER2, ADAPT6, can be used to deliver the highly cytotoxic protein domain PE25, a derivative of Pseudomonas exotoxin A, to HER2 overexpressing malignant cells, leading to potent and specific cell killing. In this study we expanded the investigation for an optimal targeting domain and constructed two fusion toxins where a HER2-binding affibody molecule, ZHER2:2891, or the dual-HER2-binding hybrid ZHER2:2891-ADAPT6 were used for cancer cell targeting. We found that both targeting domains conferred strong binding to HER2; both to the purified extracellular domain and to the HER2 overexpressing cell line SKOV3. This resulted in fusion toxins with high cytotoxic potency toward cell lines with high expression levels of HER2, with EC50 values between 10 and 100 pM. For extension of the plasma half-life, an albumin binding domain was also included. Intravenous injection of the fusion toxins into mice showed a profound influence of the targeting domain on biodistribution. Compared to previous results, with ADAPT6 as targeting domain, ZHER2:2891 gave rise to further extension of the plasma half-life and also shifted the clearance route of the fusion toxin from the liver to the kidneys. Collectively, the results show that the targeting domain has a major impact on uptake of PE25-based fusion toxins in different organs. The results also show that PE25-based fusion toxins with high affinity to HER2 do not necessarily increase the cytotoxicity beyond a certain point in affinity. In conclusion, ZHER2:2891 has the most favorable characteristics as targeting domain for PE25.

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Fusion to an albumin-binding domain with a high affinity for albumin extends the circulatory half-life and enhances the in vivo antitumor effects of human TRAIL

Journal of Controlled Release ◽

10.1016/j.jconrel.2016.03.004 ◽

2016 ◽

Vol 228 ◽

pp. 96-106 ◽

Cited By ~ 30

Author(s):

Rui Li ◽

Hao Yang ◽

Dianlong Jia ◽

Qianxue Nie ◽

Huawei Cai ◽

...

Keyword(s):

Half Life ◽

Binding Domain ◽

Albumin Binding ◽

Antitumor Effects ◽

High Affinity

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Targeting HER2 Expressing Tumors with a Potent Drug Conjugate Based on an Albumin Binding Domain-Derived Affinity Protein

Pharmaceutics ◽

10.3390/pharmaceutics13111847 ◽

2021 ◽

Vol 13 (11) ◽

pp. 1847

Author(s):

Javad Garousi ◽

Haozhong Ding ◽

Emma von Witting ◽

Tianqi Xu ◽

Anzhelika Vorobyeva ◽

...

Keyword(s):

Half Life ◽

Binding Domain ◽

Great Promise ◽

Albumin Binding ◽

Her2 Expression ◽

Serum Albumins ◽

Her2 Receptor ◽

Drug Conjugate ◽

Affinity Proteins

Albumin binding domain derived affinity proteins (ADAPTs) are a class of small and folded engineered scaffold proteins that holds great promise for targeting cancer tumors. Here, we have extended the in vivo half-life of an ADAPT, targeting the human epidermal growth factor receptor 2 (HER2) by fusion with an albumin binding domain (ABD), and armed it with the highly cytotoxic payload mertansine (DM1) for an investigation of its properties in vitro and in vivo. The resulting drug conjugate, ADAPT6-ABD-mcDM1, retained binding to its intended targets, namely HER2 and serum albumins. Further, it was able to specifically bind to cells with high HER2 expression, get internalized, and showed potent toxicity, with IC50 values ranging from 5 to 80 nM. Conversely, no toxic effect was found for cells with low HER2 expression. In vivo, ADAPT6-ABD-mcDM1, radiolabeled with 99mTc, was characterized by low uptake in most normal organs, and the main excretion route was shown to be through the kidneys. The tumor uptake was 5.5% ID/g after 24 h, which was higher than the uptake in all normal organs at this time point except for the kidneys. The uptake in the tumors was blockable by pre-injection of an excess of the monoclonal antibody trastuzumab (having an overlapping epitope on the HER2 receptor). In conclusion, half-life extended drug conjugates based on the ADAPT platform of affinity proteins holds promise for further development towards targeted cancer therapy.

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Preclinical Evaluation of the GRPR-Targeting Antagonist RM26 Conjugated to the Albumin-Binding Domain for GRPR-Targeting Therapy of Cancer

Pharmaceutics ◽

10.3390/pharmaceutics12100977 ◽

2020 ◽

Vol 12 (10) ◽

pp. 977

Author(s):

Ayman Abouzayed ◽

Hanna Tano ◽

Ábel Nagy ◽

Sara S. Rinne ◽

Fadya Wadeea ◽

...

Keyword(s):

Targeted Therapy ◽

Molecular Design ◽

Binding Domain ◽

Albumin Binding ◽

Targeting Therapy ◽

Indium 111 ◽

Rm26 Peptide ◽

Grpr Antagonist

The targeting of gastrin-releasing peptide receptors (GRPR) was recently proposed for targeted therapy, e.g., radiotherapy. Multiple and frequent injections of peptide-based therapeutic agents would be required due to rapid blood clearance. By conjugation of the GRPR antagonist RM26 (D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2) to an ABD (albumin-binding domain), we aimed to extend the blood circulation of peptides. The synthesized conjugate DOTA-ABD-RM26 was labelled with indium-111 and evaluated in vitro and in vivo. The labelled conjugate was stable in PBS and retained specificity and its antagonistic function against GRPR. The half-maximal inhibitory concentration (IC50) of natIn-DOTA-ABD-RM26 in the presence of human serum albumin was 49 ± 5 nM. [111In]In-DOTA-ABD-RM26 had a significantly longer residence time in blood and in tumors (without a significant decrease of up to 144 h pi) than the parental RM26 peptide. We conclude that the ABD-RM26 conjugate can be used for GRPR-targeted therapy and delivery of cytotoxic drugs. However, the undesirable elevated activity uptake in kidneys abolishes its use for radionuclide therapy. This proof-of-principle study justified further optimization of the molecular design of the ABD-RM26 conjugate.

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Controlling aminosilane layer thickness to extend the plasma half-life of stealth persistent luminescence nanoparticles in vivo

Journal of Materials Chemistry B ◽

10.1039/c5tb00146c ◽

2015 ◽

Vol 3 (19) ◽

pp. 4009-4016 ◽

Cited By ~ 13

Author(s):

Thomas Maldiney ◽

Maxime Rémond ◽

Michel Bessodes ◽

Daniel Scherman ◽

Cyrille Richard

Keyword(s):

Protein Adsorption ◽

Layer Thickness ◽

Half Life ◽

Persistent Luminescence ◽

Plasma Half Life

In this article, we suggest the use of persistent luminescence to study the influence of aminosilane layer thickness, on both in vitro protein adsorption and in vivo biodistribution of stealth persistent nanophosphors.

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Engineering bispecificity into a single albumin-binding domain aimed for drug targeting and in vivo half-life extension

Current Opinion in Biotechnology ◽

10.1016/j.copbio.2013.05.065 ◽

2013 ◽

Vol 24 ◽

pp. S35

Author(s):

Johan Nilvebrant ◽

Mikael Åstrand ◽

John Löfblom ◽

Sophia Hober

Keyword(s):

Half Life ◽

Drug Targeting ◽

Binding Domain ◽

Life Extension ◽

Albumin Binding

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The reversible binding of vinblastine to platelets: implications for therapy

Blood ◽

10.1182/blood.v57.3.431.431 ◽

1981 ◽

Vol 57 (3) ◽

pp. 431-438 ◽

Cited By ~ 18

Author(s):

JG Kelton ◽

JW McDonald ◽

RM Barr ◽

I Walker ◽

W Nicholson ◽

...

Keyword(s):

Half Life ◽

Immune Thrombocytopenia ◽

Suitable Model ◽

Reversible Binding ◽

In Vivo Binding ◽

Rabbit Platelets ◽

Kinetics Of ◽

Plasma Half Life

Abstract The ability of platelets to adsorb vinblastine has been used to treat patients with immune thrombocytopenia. It is hypothesized that the drug- platelet complex is coated with antibody, taken up by macrophages which are then destroyed by the drug. We gave 16 courses of vinblastine- platelets to six patients with immune thrombocytopenia. Only one patient responded, and therefore we examined possible reasons for the lack of benefit. Using 3H-vinblastine, the kinetics of vinblastine binding to platelets was studied in vitro. The binding of vinblastine to both human and rabbit platelets was identical with maximal binding occurring within 10 min at 600 microgram/ml vinblastine. Similarly, the plasma half-life of vinblastine in rabbits was close to that reported for man, and therefore, in vivo binding of vinblastine to platelets in rabbits was considered a suitable model for man. Homologous donor rabbit platelets were labeled with 51Cr alone, 51Cr plus vinblastine, or 3H-vinblastine and infused into recipient rabbits. Vinblastine had no effect on 51Cr survival, but all measureable vinblastine had left the platelets within 2 hr of the infusion. These observations suggest that delivery of the vinblastine to the macrophages depends on the platelets being phagtocytized before the drug leaves the platelets. This would be likely to occur only in those patients with severe immune thrombocytopenia. Further investigations into this treatment should be directed at methods to maintain the drug within the platelet.

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An Albumin-Binding Domain Peptide Confers Enhanced Immunoprotection Against Viral Myocarditis by CVB3 VP1 Vaccine

Frontiers in Immunology ◽

10.3389/fimmu.2021.666594 ◽

2021 ◽

Vol 12 ◽

Author(s):

Yue Gao ◽

Yan Yue ◽

Sidong Xiong

Keyword(s):

Cytotoxic T Lymphocyte ◽

Viral Myocarditis ◽

Body Weight Loss ◽

Binding Domain ◽

Albumin Binding ◽

Protein Vaccine ◽

Histomorphological Changes ◽

Domain Peptide

Coxsackievirus B3 (CVB3)-induced viral myocarditis is a common clinical cardiovascular disease without effective available vaccine. In this study, we tried to potentiate the immunoprotection efficacy of our previous CVB3-specific VP1 protein vaccine by introducing a streptococcal protein G-derived, draining lymph nodes (dLNs)-targeting albumin-binding domain (ABD) peptide. We found that compared with the original VP1 vaccine, ABD-fused VP1 (ABD-VP1) vaccine gained the new ability to efficiently bind murine albumin both in vitro and in vivo, possessed a much longer serum half-life in serum and exhibited more abundance in the dLNs after immunization. Accordingly, ABD-VP1 immunization not only significantly facilitated the enrichment and maturation of dendritic cells (DCs), induced higher percentages of IFN-γ+ CD8+ cells in the dLNs, but also robustly promoted VP1-induced T cell proliferation and cytotoxic T lymphocyte (CTL) responses in the spleens. More importantly, ABD-VP1 also elicited higher percentages of protective CD44hi CD62Lhi memory T cells in dLNs and spleens. Consequently, obvious protective effect against viral myocarditis was conferred by ABD-VP1 vaccine compared to the VP1 vaccine, reflected by the less body weight loss, improved cardiac function, alleviated cardiac histomorphological changes and an increased 28-day survival rate. Our results indicated that the ABD might be a promising immune-enhancing regime for vaccine design and development.

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