| 1. |
- Altai, Mohamed, et al.
(författare)
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Influence of Molecular Design on the Targeting Properties of ABD-Fused Mono- and Bi-Valent Anti-HER3 Affibody Therapeutic Constructs
- 2018
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Ingår i: Cells. - : MDPI AG. - 2073-4409. ; 7:10
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Tidskriftsartikel (refereegranskat)abstract
- Overexpression of human epidermal growth factor receptor type 3 (HER3) is associated with tumour cell resistance to HER-targeted therapies. Monoclonal antibodies (mAbs) targeting HER3 are currently being investigated for treatment of various types of cancers. Cumulative evidence suggests that affibody molecules may be appropriate alternatives to mAbs. We previously reported a fusion construct (3A3) containing two HER3-targeting affibody molecules flanking an engineered albumin-binding domain (ABD 035) included for the extension of half-life in circulation. The 3A3 fusion protein (19.7 kDa) was shown to delay tumour growth in mice bearing HER3-expressing xenografts and was equipotent to the mAb seribantumab. Here, we have designed and explored a series of novel formats of anti-HER3 affibody molecules fused to the ABD in different orientations. All constructs inhibited heregulin-induced phosphorylation in HER3-expressing BxPC-3 and DU-145 cell lines. Biodistribution studies demonstrated extended the half-life of all ABD-fused constructs, although at different levels. The capacity of our ABD-fused proteins to accumulate in HER3-expressing tumours was demonstrated in nude mice bearing BxPC-3 xenografts. Formats where the ABD was located on the C-terminus of affibody binding domains (3A, 33A, and 3A3) provided the best tumour targeting properties in vivo. Further development of these promising candidates for treatment of HER3-overexpressing tumours is therefore justified.
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| 2. |
- Fei, Chen, et al.
(författare)
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Detection of the human endogenous retrovirus ERV3-encoded Env-protein in human tissues using antibody-based proteomics
- 2014
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Ingår i: Journal of the Royal Society of Medicine. - : SAGE Publications. - 0141-0768 .- 1758-1095. ; 107:1, s. 22-29
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Tidskriftsartikel (refereegranskat)abstract
- Objectives: There is growing evidence to suggest that human endogenous retroviruses (HERVs) have contributed to human evolution, being expressed in development, normal physiology and disease. A key difficulty in the scientific evaluation of this potential viral contribution is the accurate demonstration of virally expressed protein in specific human cells and tissues. In this study, we have adopted the endogenous retrovirus, ERV3, as our test model in developing a reliable high-capacity methodology for the expression of such endogenous retrovirus-coded protein. Design: Two affinity-purified polyclonal antibodies to ERV3 Env-encoded protein were generated to detect the corresponding protein expression pattern in specific human cells, tissues and organs. Participants: Sampling included normal tissues from 144 individuals ranging from childhood to old age. This included more than forty different tissues and organs and some 216 different cancer tissues representing the twenty commonest forms of human cancer. Setting: The Rudbeck Laboratory, Uppsala University and Uppsala University Hospital, Uppsala, Sweden. Main Outcome Measures: The potential expression at likely physiological level of the ERV3Env encoded protein in a wide range of human cells, tissues and organs. Results: We found that ERV3 encoded Env protein is expressed at substantive levels in placenta, testis, adrenal gland, corpus luteum, Fallopian tubes, sebaceous glands, astrocytes, bronchial epithelium and the ducts of the salivary glands. Substantive expression was also seen in a variety of epithelial cells as well as cells known to undergo fusion in inflammation and in normal physiology, including fused macrophages, myocardium and striated muscle. This contrasted strongly with the low levels expressed in other tissues types. These findings suggest that this virus plays a significant role in human physiology and may also play a possible role in disease. Conclusion: This technique can now be extended to the study of other HERV genomes within the human chromosomes that may have contributed to human evolution, physiology and disease.
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| 3. |
- Fransson, Moa, et al.
(författare)
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T regulatory cells lacking CD25 are increased in MS during relapse
- 2010
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Ingår i: Autoimmunity. - : Informa UK Limited. - 0891-6934 .- 1607-842X. ; 43:8, s. 590-597
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Tidskriftsartikel (refereegranskat)abstract
- Dysregulation of inflammatory responses is considered to be a key element in autoreactive immune responses. T regulatory cells (Tregs) are important to maintain self-tolerance and the role of CD4(+)CD25(+)FoxP3(+) Tregs in autoimmunity has been extensively investigated. Recently, it was shown that Tregs in systemic lupus erythematosus lacked CD25 but were biologically functional. These data warrants for further investigation of CD25(- ) Tregs in human autoimmunity. We analyzed relapsing-remitting multiple sclerosis (MS) patients by multicolor flow cytometry for the expression of CD3, CD4, IL2R (CD25), FoxP3, and the IL7R (CD127). Further, the level of Tregs was compared in remitting and relapsing patients and correlated with disease duration. Patients in relapse exhibited higher levels of FoxP3-positive Tregs lacking CD25 compared to healthy controls (p < 0.05), indicating that Tregs attempt to restrain immune activity during relapse. The proportion of Tregs tended to be decreased with disease duration, while CD25(+)CD4(+) and CD25(+)CD8(+) effector T-cell proportions were elevated and positively correlated with overall disease duration (p < 0.05). In conclusion, while MS patients in remission have normal levels of Tregs of different phenotype, relapsing patients show an increased proportion of systemic CD25(- )FoxP3(+) Tregs. With time, the proportion of Tregs decrease while effector T cells expand.
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| 4. |
- Orlova, Anna, 1960-, et al.
(författare)
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Evaluation of the Therapeutic Potential of a HER3-Binding Affibody Construct TAM-HER3 in Comparison with a Monoclonal Antibody, Seribantumab
- 2018
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Ingår i: Molecular Pharmaceutics. - : AMER CHEMICAL SOC. - 1543-8384 .- 1543-8392. ; 15:8, s. 3394-3403
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Tidskriftsartikel (refereegranskat)abstract
- Human epidermal growth factor receptor type 3 (HER3) is recognized to be involved in resistance to HER targeting therapies. A number of HER3-targeting monoclonal antibodies are under clinical investigation as potential cancer therapeutics. Smaller high-affinity scaffold proteins are attractive non-Fc containing alternatives to antibodies. A previous study indicated that anti-HER3 affibody molecules could delay the growth of xenografted HER3-positive tumors. Here, we designed a second-generation HER3-targeting construct (TAM-HER3), containing two HER3-specific affibody molecules bridged by an albumin-binding domain (ABD) for extension of blood circulation. Receptor blocking activity was demonstrated in vitro. In mice bearing BxPC-3 xenografts, the therapeutic efficacy of TAM-HER3 was compared to the HER3-specific monoclonal antibody seribantumab (MM-121). TAM-HER3 inhibited heregulin-induced phosphorylation in a panel of HER3-expressing cancer cells and was found to be equally as potent as seribantumab in terms of therapeutic efficacy in vivo and with a similar safety profile. Median survival times were 60 days for TAM-HER3, 54 days for seribantumab, and 41 days for the control group. No pathological changes were observed in cytopathological examination. The multimeric HER3-binding affibody molecule in fusion to ABD seems promising for further evaluation as candidate therapeutics for treatment of HER3-overexpressing tumors.
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| 5. |
- Westerlund, Kristina, et al.
(författare)
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Radionuclide Therapy of HER2-Expressing Human Xenografts Using Affibody-Based Peptide Nucleic Acid-Mediated Pretargeting : In Vivo Proof of Principle
- 2018
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Ingår i: Journal of Nuclear Medicine. - : Society of Nuclear Medicine. - 0161-5505 .- 1535-5667 .- 2159-662X. ; 59:7, s. 1092-1098
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Tidskriftsartikel (refereegranskat)abstract
- Affibody molecules are small proteins engineered using a nonanti-body scaffold. Radiolabeled Affibody molecules are excellent imaging probes, but their application to radionuclide therapy has been prevented by high renal reabsorption. The aim of this study was to test the hypothesis that Affibody-based peptide nucleic acid (PNA)-mediated pretargeted therapy of human epidermal growth factor receptor 2 (HER2)-expressing cancer extends survival without accompanying renal toxicity.Methods: A HER2-targeting Affibody molecule ligated with an AGTCGTGATGTAGTC PNA hybridization probe (Z(HER2:342)-SR-HP1) was used as the primary pretargeting agent. A complementary AGTCGTGATGTAGTC PNA conjugated to the chelator DOTA and labeled with the radionuclide Lu-177 (Lu-177-HP2) was used as the secondary agent. The influence of different factors on pretargeting was investigated. Experimental radionuclide therapy in mice bearing SKOV-3 xenografts was performed in 6 cycles separated by 7 d.Results: Optimal tumor targeting was achieved when 16 MBq/3.5 mu g (0.65 nmol) of Lu-177-HP2 was injected 16 h after injection of 100 mu g (7.7 nmol) of Z(HER2:342)-SR-HP1. The calculated absorbed dose to tumors was 1,075 mGy/MBq, whereas the absorbed dose to kidneys was 206 mGy/MBq and the absorbed dose to blood (surrogate of bone marrow) was 4 mGy/MBq. Survival of mice was significantly longer (P < 0.05) in the treatment group (66 d) than in the control groups treated with the same amount of Z(HER2:342)-SR-HP1 only (37 d), the same amount and activity of Lu-177-HP2 only (32 d), or phosphate-buffered saline (37 d).Conclusion: The studied pretargeting system can deliver an absorbed dose to tumors appreciably exceeding absorbed doses to critical organs, making Affibody-based PNA-mediated pretargeted radionuclide therapy highly attractive.
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