Publications by authors named "Claudia Mendler"

6 Publications

  • Page 1 of 1

First In-Human Medical Imaging with a PASylated Zr-Labeled Anti-HER2 Fab-Fragment in a Patient with Metastatic Breast Cancer.

Nucl Med Mol Imaging 2020 Apr 20;54(2):114-119. Epub 2020 Apr 20.

5Klinikum rechts der Isar, Technische Universität München, 81675 Munich, Germany.

Purpose: PASylation® offers the ability to systematically tune and optimize the pharmacokinetics of protein tracers for molecular imaging. Here we report the first clinical translation of a PASylated Fab fragment (Zr∙Df-HER2-Fab-PAS) for the molecular imaging of tumor-related HER2 expression.

Methods: A patient with HER2-positive metastatic breast cancer received 37 MBq of Zr∙Df-HER2-Fab-PAS at a total mass dose of 70 μg. PET/CT was carried out 6, 24, and 45 h after injection, followed by image analysis of biodistribution, normal organ uptake, and lesion targeting.

Results: Images show a biodistribution typical for protein tracers, characterized by a prominent blood pool 6 h p.i., which decreased over time. Lesions were detectable as early as 24 h p.i. Zr∙Df-HER2-Fab-PAS was tolerated well.

Conclusion: This study demonstrates that a PASylated Fab tracer shows appropriate blood clearance to allow sensitive visualization of small tumor lesions in a clinical setting.
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http://dx.doi.org/10.1007/s13139-020-00638-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7198682PMC
April 2020

Selection of an Anticalin® against the membrane form of Hsp70 via bacterial surface display and its theranostic application in tumour models.

Biol Chem 2018 02;399(3):235-252

Munich Center for Integrated Protein Science, CIPS-M, and Lehrstuhl für Biologische Chemie, Technische Universität München, D-85354 Freising (Weihenstephan), Germany.

We describe the selection of Anticalins against a common tumour surface antigen, human Hsp70, using functional display on live Escherichia coli cells as fusion with a truncated EspP autotransporter. While found intracellularly in normal cells, Hsp70 is frequently exposed in a membrane-bound state on the surface of tumour cells and, even more pronounced, in metastases or after radiochemotherapy. Employing a recombinant Hsp70 fragment comprising residues 383-548 as the target, Anticalins were selected from a naïve bacterial library. The Anticalin with the highest affinity (KD=13 nm), as determined towards recombinant full-length Hsp70 by real-time surface plasmon resonance analysis, was improved to KD=510 pm by doped random mutagenesis and another cycle of E. coli surface display, followed by rational combination of mutations. This Anticalin, which recognises a linear peptide epitope located in the interdomain linker of Hsp70, was demonstrated to specifically bind Hsp70 in its membrane-associated form in immunofluorescence microscopy and via flow cytometry using the FaDu cell line, which is positive for surface Hsp70. The radiolabelled and PASylated Anticalin revealed specific tumour accumulation in xenograft mice using positron emission tomography (PET) imaging. Furthermore, after enzymatic coupling to the protein toxin gelonin, the Anticalin showed potent cytotoxicity on FaDu cells in vitro.
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http://dx.doi.org/10.1515/hsz-2017-0207DOI Listing
February 2018

Tumor Uptake of Anti-CD20 Fabs Depends on Tumor Perfusion.

J Nucl Med 2016 Dec 14;57(12):1971-1977. Epub 2016 Jul 14.

Nuklearmedizinische Klinik und Poliklinik, Klinikum rechts der Isar, Technische Universität München, München, Germany.

Antibodies have become an established treatment modality in cancer therapy during the last decade. However, these treatments often suffer from an insufficient and heterogeneous response despite validated antigen or target receptor expression in the tumor. In fact, therapeutic success depends on both the presence of the tumor antigen and its accessibility by the antibody. In search of a suitable preclinical animal model to evaluate the mechanisms of tumor heterogeneity and hemodynamics, we characterized two exemplary non-Hodgkin lymphoma subtypes with comparable CD20 expression and metabolism, SUDHL-4 and Granta-519, using multimodal imaging techniques.

Methods: To investigate in vivo biodistribution, two differently modified αCD20 antigen-binding fragments (Fab), prepared by PASylation with a 200-residue polypeptide tag comprising Pro, Ala, and Ser (PAS) and by fusion with an albumin-binding domain (ABD), were radiolabeled with I and intravenously injected into immunocompromised mice bearing corresponding xenografts.

Results: Validation with F-FDG revealed a similar distribution in vital tumor tissue 1 h after injection. However, large differences in tumor uptake were observed when the CD20-specific radiotracers I-Fab-ABD and I-Fab-PAS were applied (respective percentages injected dose per gram at 24 h after injection: 12.3 and 2.4 for Granta-519 vs. 5.8 and 1.2 for SUDHL-4). Three-dimensional light-sheet fluorescence microscopy with Cy5-Fab-PAS confirmed better tracer extravasation in the Granta-519 tumors. Moreover, dynamic contrast-enhanced (DCE) MRI revealed significantly reduced perfusion in the SUDHL-4 tumors.

Conclusion: Tracer uptake was highly dependent on local tumor perfusion and Fab permeation in the SUDHL-4 and Granta-519 tumors. Thus, the SUDHL-4 xenograft offers an excellent model for investigating the influence of therapies affecting tumor angiogenesis.
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http://dx.doi.org/10.2967/jnumed.116.176784DOI Listing
December 2016

α/β coiled coils.

Elife 2016 Jan 15;5. Epub 2016 Jan 15.

Department of Protein Evolution, Max Planck Institute for Developmental Biology, Tübingen, Germany.

Coiled coils are the best-understood protein fold, as their backbone structure can uniquely be described by parametric equations. This level of understanding has allowed their manipulation in unprecedented detail. They do not seem a likely source of surprises, yet we describe here the unexpected formation of a new type of fiber by the simple insertion of two or six residues into the underlying heptad repeat of a parallel, trimeric coiled coil. These insertions strain the supercoil to the breaking point, causing the local formation of short β-strands, which move the path of the chain by 120° around the trimer axis. The result is an α/β coiled coil, which retains only one backbone hydrogen bond per repeat unit from the parent coiled coil. Our results show that a substantially novel backbone structure is possible within the allowed regions of the Ramachandran space with only minor mutations to a known fold.
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http://dx.doi.org/10.7554/eLife.11861DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4786415PMC
January 2016

⁸⁹Zr-Labeled Versus ¹²⁴I-Labeled αHER2 Fab with Optimized Plasma Half-Life for High-Contrast Tumor Imaging In Vivo.

J Nucl Med 2015 Jul 21;56(7):1112-8. Epub 2015 May 21.

Munich Center for Integrated Protein Science (CIPS-M) and Lehrstuhl für Biologische Chemie, Technische Universität München, Freising-Weihenstephan, Germany

Unlabelled: Immuno-PET imaging of the tumor antigen HER2/neu allows for the noninvasive detection and monitoring of oncogene expression; such detection and monitoring are of prognostic value in patients with breast cancer. Compared with the full-size antibody trastuzumab, smaller protein tracers with more rapid blood clearance permit higher imaging contrast at earlier time points. Antigen-binding fragments (Fabs) of antibodies with moderately prolonged circulation achieved through the genetic fusion with a long, conformationally disordered chain of the natural amino acids Pro, Ala, and Ser (PASylation)-a biologic alternative to chemical conjugation with polyethylene glycol, PEG-offer a promising tracer format with improved pharmacokinetics for in vivo imaging. Recently, the transition metal radionuclide (89)Zr has attracted increasing interest for immuno-PET studies, complementing the conventional halogen radionuclide (124)I.

Methods: To allow direct comparison of these 2 radioactive labels for the same protein tracer, the recombinant αHER2 Fab fused with 200 Pro, Ala, and Ser (PAS200) residues was either conjugated with (124)I via an iodination reagent or coupled with deferoxamine (Df) and complexed with (89)Zr. After confirmation of the stability of both radioconjugates and quality control in vitro, immuno-PET and biodistribution studies were performed with CD1-Foxn1(nu) mice bearing HER2-positive human tumor xenografts.

Results: (89)Zr⋅Df-Fab-PAS200 and (124)I-Fab-PAS200 showed specific tumor uptake of 11 and 2.3 percentage injected dose per gram 24 h after injection, respectively; both led to high tumor-to-blood (3.6 and 4.4, respectively) and tumor-to-muscle (20 and 43, respectively) ratios. With regard to off-target accumulation, overt (124)I activity was seen in the thyroid, as expected, whereas high kidney uptake was evident for (89)Zr; the latter was probably due to glomerular filtration and reabsorption of the protein tracer in proximal tubular cells.

Conclusion: Both (89)Zr- and (124)I-labeled versions of αHER2 Fab-PAS200 allowed PET tumor imaging with high contrast. With its residualizing radiometal, the tracer (89)Zr⋅Df-Fab-PAS200 showed better in vivo stability and higher tumor uptake.
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http://dx.doi.org/10.2967/jnumed.114.149690DOI Listing
July 2015

High contrast tumor imaging with radio-labeled antibody Fab fragments tailored for optimized pharmacokinetics via PASylation.

MAbs 2015 ;7(1):96-109

a Munich Center for Integrated Protein Science (CIPS-M) and Lehrstuhl für Biologische Chemie ; Technische Universität München ; Freising-Weihenstephan , Germany.

Although antigen-binding fragments (Fabs) of antibodies constitute established tracers for in vivo radiodiagnostics, their functionality is hampered by a very short circulation half-life. PASylation, the genetic fusion with a long, conformationally disordered amino acid chain comprising Pro, Ala and Ser, provides a convenient way to expand protein size and, consequently, retard renal filtration. Humanized αHER2 and αCD20 Fabs were systematically fused with 100 to 600 PAS residues and produced in E. coli. Cytofluorimetric titration analysis on tumor cell lines confirmed that antigen-binding activities of the parental antibodies were retained. The radio-iodinated PASylated Fabs were studied by positron emission tomography (PET) imaging and biodistribution analysis in mouse tumor xenograft models. While the unmodified αHER2 and αCD20 Fabs showed weak tumor uptake (0.8% and 0.2% ID/g, respectively; 24 h p.i.) tumor-associated radioactivity was boosted with increasing PAS length (up to 9 and 26-fold, respectively), approaching an optimum for Fab-PAS400. Remarkably, 6- and 5-fold higher tumor-to-blood ratios compared with the unmodified Fabs were measured in the biodistribution analysis (48 h p.i.) for αHER2 Fab-PAS100 and Fab-PAS200, respectively. These findings were confirmed by PET studies, showing high imaging contrast in line with tumor-to-blood ratios of 12.2 and 5.7 (24 h p.i.) for αHER2 Fab-PAS100 and Fab-PAS200. Even stronger tumor signals were obtained with the corresponding αCD20 Fabs, both in PET imaging and biodistribution analysis, with an uptake of 2.8% ID/g for Fab-PAS100 vs. 0.24% ID/g for the unmodified Fab. Hence, by engineering Fabs via PASylation, plasma half-life can be tailored to significantly improve tracer uptake and tumor contrast, thus optimally matching reagent/target interactions.
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http://dx.doi.org/10.4161/19420862.2014.985522DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4622060PMC
September 2015
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