Multimerization of cRGD Peptides by Click Chemistry: Synthetic Strategies, Chemical Limitations, and Influence on Biological Properties

Abstract

Integrin α(ν)β(3) is overexpressed on endothelial cells of growing vessels as well as on several tumor types, and so integrin-binding radiolabeled cyclic RGD pentapeptides have attracted increasing interest for in vivo imaging of α(ν)β(3) integrin expression by positron emission tomography (PET). Of the cRGD derivatives available for imaging applications, systems comprising multiple cRGD moieties have recently been shown to exhibit highly favorable properties in relation to monomers. To assess the synthetic limits of the cRGD-multimerization approach and thus the maximum multimer size achievable by using different efficient conjugation reactions, we prepared a variety of multimers that were further investigated in vitro with regard to their avidities to integrin α(ν)β(3.) The synthesized peptide multimers containing increasing numbers of cRGD moieties on PAMAM dendrimer scaffolds were prepared by different click chemistry coupling strategies. A cRGD hexadecimer was the largest construct that could be synthesized under optimized reaction conditions, thus identifying the current synthetic limitations for cRGD multimerization. The obtained multimeric systems were conjugated to a new DOTA-based chelator developed for the derivatization of sterically demanding structures and successfully labeled with (68)Ga for a potential in vivo application. The evaluated multimers showed very high avidities-increasing with the number of cRGD moieties-in in vitro studies on immobilized α(ν)β(3) integrin and U87MG cells, of up to 131- and 124-fold, respectively, relative to the underivatized monomer.

Functionalization of a Triazine Dendrimer Presenting Four Maleimides on the Periphery and a DOTA Group at the Core

“…Critically, conjugation can be accomplished in a range of solvents-from organic solvents to aqueous solutions-to incorporate a range of thiol-containing species. The value of this conjugation strategy-maleimide and thiol-over alternatives has been reported: Wangler et al showed that thiol-maleimide couplings proceed more effectively than oxime formation or copper-catalyzed cycloadditions [26]. Finally, we note that, when the flexible dendritic domains are fully extended, 1 places ligands in the corners of a rectangular array measuring~2.5 nm by~4.2 nm.…”

Focal adhesion stabilization by enhanced integrin-cRGD binding affinity

“…Cyclization [28], which confers structural rigidity and thus chemical stability, and other non-natural peptide modifications such as D-amino acid incorporation [29] and N-methylation [30,31], are commonly employed to improve the selectivity and affinity of the RGD sequence for a specific integrin subtype [32]. Apart from strategies for restricting the conformational space of a ligand, multivalency can further enhance its affinity for target cells by displaying additional epitopes able to promote rebinding [33][34][35]. Notably, a minimum distance between the RGD peptides and the anchoring substrate is required, such that the binding motifs are adequately oriented to facilitate integrin binding [36,37].…”

Imagerie de la néoangiogenèse en médecine nucléaire

“…Ces dernières ont été marquées par des émetteurs bêta ou gamma et ont montré un ciblage prometteur chez le petit animal. D'autres équipes travaillent sur un nouveau type de VEGF, le single-chain VEGF (scVEGF), présentant dans sa distalité un motif Cys-tag permettant un marquage au 99m Tc, mais aussi avec des émetteurs bêta : 64 Cu, 68 Ga, 18 [11]. Au vu de ces premiers résultats prometteurs chez l'humain, différents essais cliniques avec pour cible le VEGF ou son récepteur sont en cours (www.clinicaltrials.org).…”

“…Alors que les premiers peptides cycliques étudiés étaient monomériques, certaines équipes ont développé des peptides dimériques ou tétramériques, cela afin d'accroître leur affinité aux intégrines [16][17][18]. Dijkgraaj et al ont ainsi synthétisé, à partir d'un peptide E-c(RGDfK), un peptide dimérique et tétramérique conjugué au DOTA, se traduisant par une augmentation significative de la captation du traceur avec le nombre de monomère conjugué [17].…”

“…La dose effective corps entier était estimée à 0,0396 mSv/MBq. Enfin, un essai clinique est en cours actuellement pour étudier la biodistribution de deux composants proches : le 18 [38].…”