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Gräslund, TorbjörnORCID iD iconorcid.org/0000-0002-5391-600X
Alternative names
Publications (10 of 65) Show all publications
Yu, S., Alkharusi, A., Norstedt, G. & Gräslund, T. (2019). An in vivo half-life extended prolactin receptor antagonist can prevent STAT5 phosphorylation. PLoS ONE, 14(5), Article ID e0215831.
Open this publication in new window or tab >>An in vivo half-life extended prolactin receptor antagonist can prevent STAT5 phosphorylation
2019 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 14, no 5, article id e0215831Article in journal (Refereed) Published
Abstract [en]

Increasing evidence suggests that signaling through the prolactin/prolactin receptor axis is important for stimulation the growth of many cancers including glioblastoma multiforme, breast and ovarian carcinoma. Efficient inhibitors of signaling have previously been developed but their applicability as cancer drugs is limited by the short in vivo half-life. In this study, we show that a fusion protein, consisting of the prolactin receptor antagonist PrlRA and an albumin binding domain for half-life extension can be expressed as inclusion bodies in Escherichia coli and efficiently refolded and purified to homogeneity. The fusion protein was found to have strong affinity for the two intended targets: the prolactin receptor (K-D = 2.3 +/- 0.2 nM) and mouse serum albumin (K-D = 0.38 +/- 0.01 nM). Further investigation showed that it could efficiently prevent prolactin mediated phosphorylation of STAT5 at 100 nM concentration and above, similar to the PrlRA itself, suggesting a potential as drug for cancer therapy in the future. Complexion with HSA weakened the affinity for the receptor to 21 +/- 3 nM, however the ability to prevent phosphorylation of STAT5 was still prominent. Injection into rats showed a 100-fold higher concentration in blood after 24 h compared to PrlRA itself.

Place, publisher, year, edition, pages
PUBLIC LIBRARY SCIENCE, 2019
National Category
Medical Biotechnology
Identifiers
urn:nbn:se:kth:diva-252381 (URN)10.1371/journal.pone.0215831 (DOI)000467148400008 ()31063493 (PubMedID)2-s2.0-85065793714 (Scopus ID)
Note

QC 20190718

Available from: 2019-07-18 Created: 2019-07-18 Last updated: 2019-08-29Bibliographically approved
Ding, H., Altai, M., Rinne, S. S., Vorobyeva, A., Tolmachev, V., Gräslund, T. & Orlova, A. (2019). Incorporation of a Hydrophilic Spacer Reduces Hepatic Uptake of HER2-Targeting Affibody-DM1 Drug Conjugates. Cancers, 11(8), Article ID 1168.
Open this publication in new window or tab >>Incorporation of a Hydrophilic Spacer Reduces Hepatic Uptake of HER2-Targeting Affibody-DM1 Drug Conjugates
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2019 (English)In: Cancers, ISSN 2072-6694, Vol. 11, no 8, article id 1168Article in journal (Refereed) Published
Abstract [en]

Affibody molecules are small affinity-engineered scaffold proteins which can be engineered to bind to desired targets. The therapeutic potential of using an affibody molecule targeting HER2, fused to an albumin-binding domain (ABD) and conjugated with the cytotoxic maytansine derivate MC-DM1 (AffiDC), has been validated. Biodistribution studies in mice revealed an elevated hepatic uptake of the AffiDC, but histopathological examination of livers showed no major signs of toxicity. However, previous clinical experience with antibody drug conjugates have revealed a moderateto high-grade hepatotoxicity in treated patients, which merits efforts to also minimize hepatic uptake of the AffiDCs. In this study, the aim was to reduce the hepatic uptake of AffiDCs and optimize their in vivo targeting properties. We have investigated if incorporation of hydrophilic glutamate-based spacers adjacent to MC-DM1 in the AffiDC, (Z(HER2:2891))(2) -ABD-MC-DM1, would counteract the hydrophobic nature of MC-DM1 and, hence, reduce hepatic uptake. Two new AffiDCs including either a triglutamate-spacer-, (Z(HER2:2891))(2)-ABD-E-3-MC-DM1, or a hexaglutamate-spacer-, (Z(HER2:2891))(2)-ABD-E-6-MC-DM1 next to the site of MC-DM1 conjugation were designed. We radiolabeled the hydrophilized AffiDCs and compared them, both in vitro and in vivo, with the previously investigated (Z(HER2:2891))(2)-ABD-MC-DM1 drug conjugate containing no glutamate spacer. All three AffiDCs demonstrated specific binding to HER2 and comparable in vitro cytotoxicity. A comparative biodistribution study of the three radiolabeled AffiDCs showed that the addition of glutamates reduced drug accumulation in the liver while preserving the tumor uptake. These results confirmed the relation between DM1 hydrophobicity and liver accumulation. We believe that the drug development approach described here may also be useful for other affinity protein-based drug conjugates to further improve their in vivo properties and facilitate their clinical translatability.

Place, publisher, year, edition, pages
MDPI, 2019
Keywords
affibody, drug conjugates, hepatic uptake, DM1
National Category
Medical Biotechnology
Identifiers
urn:nbn:se:kth:diva-261042 (URN)10.3390/cancers11081168 (DOI)000484438000128 ()31416167 (PubMedID)2-s2.0-85071755584 (Scopus ID)
Note

QC 20191002

Available from: 2019-10-02 Created: 2019-10-02 Last updated: 2019-10-02Bibliographically approved
Liu, H., Lindbo, S., Ding, H., Altai, M., Garousi, J., Orlova, A., . . . Gräslund, T. (2019). Potent and specific fusion toxins consisting of a HER2‑binding, ABD‑derived affinity protein, fused to truncated versions of Pseudomonas exotoxin A. International Journal of Oncology, 55(1), 309-319
Open this publication in new window or tab >>Potent and specific fusion toxins consisting of a HER2‑binding, ABD‑derived affinity protein, fused to truncated versions of Pseudomonas exotoxin A
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2019 (English)In: International Journal of Oncology, ISSN 1019-6439, Vol. 55, no 1, p. 309-319Article in journal (Refereed) Published
Abstract [en]

Fusion toxins consisting of an affinity protein fused to toxic polypeptides derived from Pseudomonas exotoxin A (ETA) are promising agents for targeted cancer therapy. In this study, we examined whether fusion toxins consisting of an albumin binding domain-derived affinity protein (ADAPT) interacting with human epidermal growth factor receptor 2 (HER2), coupled to the ETA-derived polypeptides PE38X8 or PE25, with or without an albumin binding domain (ABD) for half-life extension, can be used for specific killing of HER2-expressing cells. The fusion toxins could easily be expressed in a soluble form in Escherichia coli and purified to homogeneity. All constructs had strong affinity for HER2 (K-D 10 to 26 nM) and no tendency for aggregation could be detected. The fusion toxins including the ABD showed strong interaction with human and mouse serum albumin [equilibrium dissociation constant (K-D) 1 to 3 nM and 2 to 10 nM, respectively]. The in vitro investigation of the cytotoxic potential revealed IC50-values in the picomolar range for cells expressing high levels of HER2. The specificity was also demonstrated, by showing that free HER2 receptors on the target cells are required for fusion toxin activity. In mice, the fusion toxins containing the ABD exhibited an appreciably longer time in circulation. The uptake was highest in liver and kidney. Fusion with PE25 was associated with the highest hepatic uptake. Collectively, the results suggest that fusion toxins consisting of ADAPTs and ETA-derivatives are promising agents for targeted cancer therapy.

Place, publisher, year, edition, pages
Spandidos Publications, 2019
Keywords
exotoxin A, Pseudomonas, ABD-derived affinity protein, half-life extension, cancer, human epidermal growth factor receptor 2
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:kth:diva-255762 (URN)10.3892/ijo.2019.4814 (DOI)000476557600026 ()31180549 (PubMedID)2-s2.0-85068411992 (Scopus ID)
Note

QC 20190812

Available from: 2019-08-12 Created: 2019-08-12 Last updated: 2019-08-12Bibliographically approved
Altai, M., Liu, H., Ding, H., Mitran, B., Edqvist, P.-H. -., Tolmachev, V., . . . Gräslund, T. (2018). Affibody-derived drug conjugates: Potent cytotoxic molecules for treatment of HER2 over-expressing tumors. Journal of Controlled Release, 288, 84-95
Open this publication in new window or tab >>Affibody-derived drug conjugates: Potent cytotoxic molecules for treatment of HER2 over-expressing tumors
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2018 (English)In: Journal of Controlled Release, ISSN 0168-3659, E-ISSN 1873-4995, Vol. 288, p. 84-95Article in journal (Refereed) Published
Abstract [en]

Patients with HER2-positive tumors often suffer resistance to therapy, warranting development of novel treatment modalities. Affibody molecules are small affinity proteins which can be engineered to bind to desired targets. They have in recent years been found to allow precise targeting of cancer specific molecular signatures such as the HER2 receptor. In this study, we have investigated the potential of an affibody molecule targeting HER2, ZHER2:2891, conjugated with the cytotoxic maytansine derivate MC-DM1, for targeted cancer therapy. ZHER2:2891 was expressed as a monomer (ZHER2:2891), dimer ((ZHER2:2891)2) and dimer with an albumin binding domain (ABD) for half-life extension ((ZHER2:2891)2-ABD). All proteins had a unique C-terminal cysteine that could be used for efficient and site-specific conjugation with MC-DM1. The resulting affibody drug conjugates were potent cytotoxic molecules for human cells over-expressing HER2, with sub-nanomolar IC50-values similar to trastuzumab emtansine, and did not affect cells with low HER2 expression. A biodistribution study of a radiolabeled version of (ZHER2:2891)2-ABD-MC-DM1, showed that it was taken up by the tumor. The major site of off-target uptake was the kidneys and to some extent the liver. (ZHER2:2891)2-ABD-MC-DM1 was found to have a half-life in circulation of 14 h. The compound was tolerated well by mice at 8.5 mg/kg and was shown to extend survival of mice bearing HER2 over-expressing tumors. The findings in this study show that affibody molecules are a promising class of engineered affinity proteins to specifically deliver small molecular drugs to cancer cells and that such conjugates are potential candidates for clinical evaluation on HER2-overexpressing cancers. 

Place, publisher, year, edition, pages
Elsevier B.V., 2018
Keywords
ADC, Affibody molecule, DM1, HER2, Maytansine, Amino acids, Diseases, Mammals, Molecules, Monoclonal antibodies, Patient treatment, Tumors, Affibody molecules, Clinical evaluation, Molecular signatures, Targeted cancer therapy, Terminal cysteine, Treatment modality, Dimers, (z human epidermal growth factor receptor 2 2891) 2 albumin binding domain iaa conjugate, (z human epidermal growth factor receptor 2 2891) 2 albumin binding domain mc dm1 conjugate, (z human epidermal growth factor receptor 2 2891) 2 mc dm1 conjugate, (z human epidermal growth factor receptor 2 2891) mc dm1 conjugate, (z taq) 2 albumin binding domain mc dm1 conjugate, cysteine, cytotoxic agent, dimer, epidermal growth factor receptor 2, epidermal growth factor receptor kinase inhibitor, monomer, serum albumin, technetium 99m, trastuzumab emtansine, unclassified drug, albumin binding domain, animal experiment, animal model, animal tissue, Article, AU565 cell line, binding affinity, cancer survival, carboxy terminal sequence, controlled study, drug conjugation, drug cytotoxicity, drug dose comparison, drug efficacy, drug half life, drug potency, drug specificity, drug targeting, drug tolerability, female, human, human tissue, IC50, in vitro study, isotope labeling, kidney tissue, liver tissue, MCF-7 cell line, molecularly targeted therapy, mouse, nonhuman, ovary carcinoma, priority journal, protein expression, protein structure, SK-BR-3 cell line, SK-OV-3 cell line, tissue distribution
National Category
Chemical Sciences
Identifiers
urn:nbn:se:kth:diva-236627 (URN)10.1016/j.jconrel.2018.08.040 (DOI)000446237700007 ()2-s2.0-85053511226 (Scopus ID)
Funder
Swedish Cancer Society, 14-0298; 14/474; 15/350; 15/746Swedish Research Council, 2015-02509; 2015-02353VINNOVA, 2015-02509; 2015-02353Swedish Society for Medical Research (SSMF)
Note

Export Date: 22 October 2018; Article; CODEN: JCREE; Correspondence Address: Gräslund, T.; Department of Protein Sciecne, KTH Royal Institute of Technology, Roslagstullsbacken 21, Sweden; email: torbjorn@kth.se; Funding details: 14/474, Cancerfonden; Funding details: 14-0298, Cancerfonden; Funding details: 15/746, Cancerfonden; Funding details: 15/350, Cancerfonden; Funding details: SSMF, Svenska Sällskapet för Medicinsk Forskning; Funding details: 2016-04060; Funding details: 2015-02353, VR, Vetenskapsrådet; Funding details: 2015-02509, VR, Vetenskapsrådet; Funding text: This work was supported by The Swedish Cancer Society (Cancerfonden), grants 14-0298, 14/474, 15/350, 15/746 , the Swedish Research Council ; grants 2015-02509 and 2015-02353 , and the Swedish Agency for Innovation VINNOVA (grant 2016-04060 . Mohamed Altai's research activities are kindly supported by the Swedish Society for Medical Research (SSMF). Appendix A. QC 20181114

Available from: 2018-11-14 Created: 2018-11-14 Last updated: 2018-11-14Bibliographically approved
Seijsing, J., Yu, S., Frejd, F. Y., Hoiden-Guthenberg, I. & Gräslund, T. (2018). In vivo depletion of serum IgG by an affibody molecule binding the neonatal Fc receptor. Scientific Reports, 8, Article ID 5141.
Open this publication in new window or tab >>In vivo depletion of serum IgG by an affibody molecule binding the neonatal Fc receptor
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2018 (English)In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 8, article id 5141Article in journal (Refereed) Published
Abstract [en]

Lowering the total level of Immunoglobulin G (IgG) in circulation is a promising general treatment option for many autoimmune diseases driven by pathogenic autoantibodies. The half-life of IgG in circulation is unusually long as a consequence of its interaction with the neonatal Fc receptor (FcRn), which protects it from lysosomal degradation by cells in contact with blood. Blocking the IgG/FcRn interaction prevents FcRn-mediated rescue, which may lead to increased catabolism and a lowering of the total IgG level. Here, we find that an engineered alternative scaffold protein, an affibody molecule, interacting specifically with FcRn, is able to block the IgG/FcRn interaction in vitro. The affibody molecule (Z(FcRn)) was expressed alone or as a fusion to an albumin binding domain (ABD), to extend its half-life in circulation, in both cases with retained affinity and blocking potential. Repeated i.v. injections in mice of Z(FcRn) and Z(FcRn)-ABD were found to result in an up to 40% reduction of the IgG serum-level after 5 days. Potential applications of Z(FcRn) as a general treatment modality for autoimmune diseases are discussed.

Place, publisher, year, edition, pages
NATURE PUBLISHING GROUP, 2018
National Category
Biological Sciences
Identifiers
urn:nbn:se:kth:diva-225708 (URN)10.1038/s41598-018-23481-5 (DOI)000428163600012 ()29572538 (PubMedID)2-s2.0-85044363795 (Scopus ID)
Note

QC 20180411

Available from: 2018-04-11 Created: 2018-04-11 Last updated: 2019-08-29Bibliographically approved
Ståhl, S., Gräslund, T., Eriksson Karlström, A., Frejd, F. Y., Nygren, P.-Å. & Löfblom, J. (2017). Affibody Molecules in Biotechnological and Medical Applications. Trends in Biotechnology, 35(8), 691-712
Open this publication in new window or tab >>Affibody Molecules in Biotechnological and Medical Applications
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2017 (English)In: Trends in Biotechnology, ISSN 0167-7799, E-ISSN 1879-3096, Vol. 35, no 8, p. 691-712Article, review/survey (Refereed) Published
Abstract [en]

Affibody molecules are small (6.5-kDa) affinity proteins based on a three-helix bundle domain framework. Since their introduction 20 years ago as an alternative to antibodies for biotechnological applications, the first therapeutic affibody molecules have now entered clinical development and more than 400 studies have been published in which affibody molecules have been developed and used in a variety of contexts. In this review, we focus primarily on efforts over the past 5 years to explore the potential of affibody molecules for medical applications in oncology, neurodegenerative, and inflammation disorders, including molecular imaging, receptor signal blocking, and delivery of toxic payloads. In addition, we describe recent examples of biotechnological applications, in which affibody molecules have been exploited as modular affinity fusion partners.

Place, publisher, year, edition, pages
Elsevier, 2017
National Category
Biological Sciences
Identifiers
urn:nbn:se:kth:diva-211597 (URN)10.1016/j.tibtech.2017.04.007 (DOI)000405847200005 ()
Note

QC 20170815

Available from: 2017-08-15 Created: 2017-08-15 Last updated: 2017-08-15Bibliographically approved
Gräslund, T. (2016). Affibody molecules: therapy and in vivo diagnostic applications. New Biotechnology, 33, S48-S48
Open this publication in new window or tab >>Affibody molecules: therapy and in vivo diagnostic applications
2016 (English)In: New Biotechnology, ISSN 1871-6784, E-ISSN 1876-4347, Vol. 33, p. S48-S48Article in journal (Refereed) Published
Place, publisher, year, edition, pages
Elsevier, 2016
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:kth:diva-202794 (URN)10.1016/j.nbt.2016.06.890 (DOI)000393400600150 ()
Note

QC 20170307

Available from: 2017-03-07 Created: 2017-03-07 Last updated: 2017-06-29Bibliographically approved
Altai, M., Liu, H., Orlova, A., Tolmachev, V. & Gräslund, T. (2016). Improving of molecular design of a novel Affibody-fused HER2-recognising anticancer toxin using radionuclide-based techniques. Paper presented at Annual Congress of the European-Association-of-Nuclear-Medicine (EANM), OCT 15-19, 2016, Barcelona, SPAIN. European Journal of Nuclear Medicine and Molecular Imaging, 43, S178-S178
Open this publication in new window or tab >>Improving of molecular design of a novel Affibody-fused HER2-recognising anticancer toxin using radionuclide-based techniques
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2016 (English)In: European Journal of Nuclear Medicine and Molecular Imaging, ISSN 1619-7070, E-ISSN 1619-7089, Vol. 43, p. S178-S178Article in journal, Meeting abstract (Other academic) Published
Place, publisher, year, edition, pages
SPRINGER, 2016
National Category
Radiology, Nuclear Medicine and Medical Imaging
Identifiers
urn:nbn:se:kth:diva-242613 (URN)000391801600438 ()
Conference
Annual Congress of the European-Association-of-Nuclear-Medicine (EANM), OCT 15-19, 2016, Barcelona, SPAIN
Note

QC 20190226

Available from: 2019-02-26 Created: 2019-02-26 Last updated: 2019-02-26Bibliographically approved
Alkharusi, A., Yu, S., Landazuri, N., Zadjali, F., Davodi, B., Nystrom, T., . . . Norstedt, G. (2016). Stimulation of prolactin receptor induces STAT-5 phosphorylation and cellular invasion in glioblastoma multiforme. OncoTarget, 7(48), 79558-79569
Open this publication in new window or tab >>Stimulation of prolactin receptor induces STAT-5 phosphorylation and cellular invasion in glioblastoma multiforme
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2016 (English)In: OncoTarget, ISSN 1949-2553, E-ISSN 1949-2553, Vol. 7, no 48, p. 79558-79569Article in journal (Refereed) Published
Abstract [en]

Glioblastoma multiforme (GBM) is the most common and aggressive primary brain tumor in humans and is characterized with poor outcome. In this study, we investigated components of prolactin (Prl) system in cell models of GBM and in histological tissue sections obtained from GBM patients. Expression of Prolactin receptor (PrlR) was detected at high levels in U251-MG, at low levels in U87-MG and barely detectable in U373 cell lines and in 66% of brain tumor tissues from 32 GBM patients by immunohistochemical technique. In addition, stimulation of U251-MG and U87-MG cells but not U373 with Prl resulted in increased STAT5 phosphorylation and only in U251-MG cells with increased cellular invasion. Furthermore, STAT5 phosphorylation and cellular invasion induced in Prl stimulated cells were significantly reduced by using a Prl receptor antagonist that consists of Prl with four amino acid replacements. We conclude that Prl receptor is expressed at different levels in the majority of GBM tumors and that blocking of PrlR in U251-MG cells significantly reduce cellular invasion.

Place, publisher, year, edition, pages
Impact Journals LLC, 2016
Keywords
prolactin, prolactin receptor, prolactin receptor antagonist, STAT5, GBM
National Category
Cancer and Oncology Cell Biology
Identifiers
urn:nbn:se:kth:diva-199535 (URN)10.18632/oncotarget.12840 (DOI)000389636000109 ()2-s2.0-84998678038 (Scopus ID)
Note

QC 20170116

Available from: 2017-01-16 Created: 2017-01-09 Last updated: 2019-08-29Bibliographically approved
Gräslund, T. (2015). Affibody molecules: Therapy and in vivo diagnostic applications. International Journal of Molecular Medicine, 36, S98-S98
Open this publication in new window or tab >>Affibody molecules: Therapy and in vivo diagnostic applications
2015 (English)In: International Journal of Molecular Medicine, ISSN 1107-3756, E-ISSN 1791-244X, Vol. 36, p. S98-S98Article in journal, Meeting abstract (Other academic) Published
National Category
Biological Sciences
Identifiers
urn:nbn:se:kth:diva-175951 (URN)000361863000375 ()
Note

QC 20151029

Available from: 2015-10-29 Created: 2015-10-26 Last updated: 2017-12-01Bibliographically approved
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Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0002-5391-600X

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