Conformational Control of Integrin‐Subtype Selectivity in <i>iso</i>DGR Peptide Motifs: A Biological Switch

Frank, Albin; Otto, Elke; Mas‐Moruno, Carlos; Schiller, Herbert B.; Marinelli, Luciana; Cosconati, Sandro; Bochen, Alexander; Vossmeyer, Dörte; Zahn, Grit; Stragies, Roland; Novellino, Ettore; Kessler, Horst

doi:10.1002/anie.201004363

Cited by 77 publications

(78 citation statements)

References 32 publications

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“…The activity and selectivity of integrin ligands were determined by a solid-phase binding assay according to the previously reported protocol103 using coated extracellular matrix proteins and soluble integrins. The following compounds were used as internal standards: Cilengitide , c (RGDf( N Me)V) (αvβ3–0.54 nM, αvβ5–8 nM, α5β1–15.4 nM), linear peptide RTDLDSLRT 4 (αvβ6–33 nM; αvβ8–100 nM) and tirofiban 5 (αIIbβ3–1.2 nM).…”

Section: Methods Sectionmentioning

confidence: 99%

“…In the first case, the integrin is coated on the surface, followed by an incubation with a mixture of the native ECM protein and increasing concentrations of the ligand of interest49. In an alternative test system, the natural ECM protein (vitronectin for αvβ3 and αvβ5, LAP (TGFβ) for αvβ6 and αvβ8, fibronectin for α5β1 and fibrinogen for αIIbβ3) is immobilized onto the surface, and the soluble integrin, together with a serial dilution of the inhibitory ligand, is added afterwards57. The read-out in both procedures is usually done in an ELISA-like manner by using conjugated antibodies recognizing the integrin head groups.…”

Section: Development Of Integrin Subtype-selective Compoundsmentioning

confidence: 99%

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A Comprehensive Evaluation of the Activity and Selectivity Profile of Ligands for RGD-binding Integrins

Kapp

Rechenmacher

Neubauer

et al. 2017

Sci Rep

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487

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Integrins, a diverse class of heterodimeric cell surface receptors, are key regulators of cell structure and behaviour, affecting cell morphology, proliferation, survival and differentiation. Consequently, mutations in specific integrins, or their deregulated expression, are associated with a variety of diseases. In the last decades, many integrin-specific ligands have been developed and used for modulation of integrin function in medical as well as biophysical studies. The IC50-values reported for these ligands strongly vary and are measured using different cell-based and cell-free systems. A systematic comparison of these values is of high importance for selecting the optimal ligands for given applications. In this study, we evaluate a wide range of ligands for their binding affinity towards the RGD-binding integrins αvβ3, αvβ5, αvβ6, αvβ8, α5β1, αIIbβ3, using homogenous ELISA-like solid phase binding assay.

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Section: Methods Sectionmentioning

confidence: 99%

Section: Development Of Integrin Subtype-selective Compoundsmentioning

confidence: 99%

A Comprehensive Evaluation of the Activity and Selectivity Profile of Ligands for RGD-binding Integrins

Kapp

Rechenmacher

Neubauer

et al. 2017

Sci Rep

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487

513

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“…Affinities of nat Ga-aquibeprin and nat Ga-avebetrin for the integrins a 5 b 1 , a v b 3 , and a IIb b 3 were determined in a solid-phase binding assay according to a previously described method (34). Briefly, after coating the corresponding extracellular matrix protein (a 5 b 1 , fibronectin; a v b 3 , vitronectin; a IIb b 3 , fibrinogen) on a plate, a mixture of the ligand and soluble integrin (a 5 b 1 , a v b 3 , or a IIb b 3, respectively) was added.…”

Section: Integrin Affinitiesmentioning

confidence: 99%

“…Some of the applied experimental protocols have been fully described earlier, namely, 68 Ga labeling for rodent experiments (32), measurement of log D (33), determination of integrin affinities (34), culture of M21 human melanoma cells and generation of respective xenografts in mice (33), biodistribution experiments and determination of in vivo metabolization (33), and micro-PET imaging (32). Thus, only brief accounts are given here, highlighting alterations made to previously reported procedures.…”

Section: Generalmentioning

confidence: 99%

Complementary, Selective PET Imaging of Integrin Subtypes α₅β₁ and α_vβ₃ Using ⁶⁸Ga-Aquibeprin and ⁶⁸Ga-Avebetrin

et al. 2015

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Despite in vivo mapping of integrin α v β 3 expression being thoroughly investigated in recent years, its clinical value is still not well defined. For imaging of angiogenesis, the integrin subtype α 5 β 1 appears to be a promising target, for which purpose we designed the PET radiopharmaceutical 68 Ga-aquibeprin. Methods: 68 Ga-aquibeprin was obtained by click-chemistry (CuAAC) trimerization of a α 5 β 1 integrin-binding pseudopeptide on the triazacyclononane-triphosphinate (TRAP) chelator, followed by automated 68 Ga labeling. Integrin α 5 β 1 and α v β 3 affinities were determined in enzyme linked immune sorbent assay on immobilized integrins, using fibronectin and vitronectin, respectively, as competitors. M21 (human melanoma)-bearing severe combined immunodeficient mice were used for biodistribution, PET imaging, and determination of in vivo metabolization. The expression of α 5 and β 3 subunits was determined by immunohistochemistry on paraffin sections of M21 tumors. Results: 68 Ga-aquibeprin shows high selectivity for integrin α 5 β 1 (50% inhibition concentration [IC 50 ] 5 0.088 nM) over α v β 3 (IC 50 5 620 nM) and a pronounced hydrophilicity (log D 5 -4.2). Severe combined immunodeficient mice xenografted with M21 human melanoma were found suitable for in vivo evaluation, as M21 immunohistochemistry showed not only an endothelial and strong cytoplasmatic expression of the β 3 integrin subunit but also an intense expression of the α 5 integrin subunit particularly in the endothelial cells of intratumoral small vessels. Ex vivo biodistribution (90 min after injection) showed high uptake in M21 tumor (2.42 ± 0.21 percentage injected dose per gram), fast renal excretion, and low background; tumor-to-blood and tumor-to-muscle ratios were 10.6 ± 2.5 and 20.9 ± 2.4, respectively. 68 Ga-aquibeprin is stable in vivo; no metabolites were detected in mouse urine, blood serum, kidney, and liver homogenates 30 min after injection. PET imaging was performed for 68 Ga-aquibeprin and the previously described, structurally related c(RGDfK) trimer 68 Ga-avebetrin, which shows an inverse selectivity for integrin α v β 3 (IC 50 5 0.22 nM) over α 5 β 1 (IC 50 5 39 nM). In vivo target specificity was proven by cross-competition studies; tumor uptake of either tracer was not affected by the coadministration of 40 nmol (∼5 mg/kg) of the respective other compound. Conclusion: 68 Ga-aquibeprin and 68 Ga-avebetrin are recommendable for complementary mapping of integrins α 5 β 1 and α v β 3 by PET, allowing for future studies on the role of these integrins in angiogenesis, tumor progression, metastasis, and myocardial infarct healing.

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“…As a novel class of peptidic integrin ligands it paves the way to drug-design studies focusing on the synthesis and characterization of a new generation of isoDGR-based cyclopeptides. [125][126][127]. IsoAspartic acid is a "-aminoacid, which induces high flexibility in isoDGRcontaining macrocycles, thus augmenting the range of accessible inter-converting conformations which are difficult to be characterized by the common spectroscopic techniques.…”

Section: The Combination Of Metadynamics and Docking Calculations: A mentioning

confidence: 99%

NMR and Computational Methods in the Structural and Dynamic Characterization of Ligand-Receptor Interactions

Ghitti

Musco

Spitaleri

2013

Advances in Experimental Medicine and Biology

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The recurrent failures in drug discovery campaigns, the asymmetry between the enormous financial investments and the relatively scarce results have fostered the development of strategies based on complementary methods. In this context in recent years the rigid lock-and-key binding concept had to be revisited in favour of a dynamic model of molecular recognition accounting for conformational changes of both the ligand and the receptor. The high level of complexity required by a dynamic description of the processes underlying molecular recognition requires a multidisciplinary investigation approach. In this perspective, the combination of nuclear magnetic resonance spectroscopy with molecular docking, conformational searches along with molecular dynamics simulations has given new insights into the dynamic mechanisms governing ligand receptor interactions, thus giving an enormous contribution to the identification and design of new and effective drugs. Herein a succinct overview on the applications of both NMR and computational methods to the structural and dynamic characterization of ligand-receptor interactions will be presented.

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Conformational Control of Integrin‐Subtype Selectivity in isoDGR Peptide Motifs: A Biological Switch

Cited by 77 publications

References 32 publications

A Comprehensive Evaluation of the Activity and Selectivity Profile of Ligands for RGD-binding Integrins

A Comprehensive Evaluation of the Activity and Selectivity Profile of Ligands for RGD-binding Integrins

Complementary, Selective PET Imaging of Integrin Subtypes α₅β₁ and α_vβ₃ Using ⁶⁸Ga-Aquibeprin and ⁶⁸Ga-Avebetrin

NMR and Computational Methods in the Structural and Dynamic Characterization of Ligand-Receptor Interactions

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Conformational Control of Integrin‐Subtype Selectivity in isoDGR Peptide Motifs: A Biological Switch

Cited by 77 publications

References 32 publications

A Comprehensive Evaluation of the Activity and Selectivity Profile of Ligands for RGD-binding Integrins

A Comprehensive Evaluation of the Activity and Selectivity Profile of Ligands for RGD-binding Integrins

Complementary, Selective PET Imaging of Integrin Subtypes α5β1 and αvβ3 Using 68Ga-Aquibeprin and 68Ga-Avebetrin

NMR and Computational Methods in the Structural and Dynamic Characterization of Ligand-Receptor Interactions

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Complementary, Selective PET Imaging of Integrin Subtypes α₅β₁ and α_vβ₃ Using ⁶⁸Ga-Aquibeprin and ⁶⁸Ga-Avebetrin