Abstract
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Objectives Glutamate-urea-based PSMA-binding peptides are a promising approach for imaging of prostate carcinoma. In these tracer peptides the pharmacophoric group is connected via a linker or spacer to the imaging component in form of a chelator or a fluorescence group. Modifications of the various components can improve the target interaction and provide new fields of application. To analyze the effect of various chelators on the binding affinity the equilibrium dissociation constant KD and also the kinetics of association and dissociation are relevant values that are measured and compared using surface plasmon resonance spectroscopy.
Methods The PSMA (prostate-specific membrane antigen) protein was covalently bound by NHS-ester coupling to the surface of a CM5 chip using a Biacore X100 (GE Healthcare). Various PCa tracers have been tested in single cycle kinetic mode to repetitively determine the association and dissociation rate and subsequently calculate the binding coefficient. For comparison the KD values were also determined using a cell binding assay.
Results The binding affinity constant for DUPA-Pep peptides coupled with the chelators DTPA (0.5 nM), DOTA (5.0 nM) and deferrioxamine (0.13 nM) or the fluorescence dye AlexaFluor®488 (0.25 nM) were determined. Measurements were performed using the same chip by regenerating the surface as well as using various chips to test for reproducibility and reliability. For various chelators an effect on the peptide binding could be observed. Furthermore there was also a detectable influence of the bound radionuclide.
Conclusions To use diverse chelators or fluorescence groups provide the option to label peptide tracer for various imaging and therapeutic applications. The results demonstrate that modification of an imaging group or radionuclide can also affect the binding properties of the entire peptide. Comparable and reproducible determination of the binding coefficient of new designed PCa tracers is necessary and can be performed by surface plasmon resonance spectroscopy.