Multimerisation of c(RGDfK) resulted in enhanced affinity for alpha(v)beta(3) as determined in vitro. Tumour uptake of a tetrameric RGD peptide was significantly higher than that of the monomeric and dimeric analogues, presumably owing to the enhanced statistical likelihood for rebinding to alpha(v)beta(3).

The overexpression of the chemokine receptor CXCR4 plays an important role in oncology, since together with its endogenous ligand, the stromal cell-derived factor (SDF1-a), CXCR4 is involved in tumor development, growth, and organ-specific metastasis. As part of our ongoing efforts to develop highly specific CXCR4-targeted imaging probes and with the aim to assess the suitability of this ligand for first proof-of-concept studies in humans, we further evaluated the new 68 Ga-labeled high-affinity cyclic CXCR4 ligand, 68 Ga-CPCR4-2 (cyclo(D- 68 Ga-DOTA]-Arg 3 -2-Nal 4 -Gly 5 )). Methods: Additional biodistribution and competitions studies in vivo, dynamic PET studies, and investigations on the metabolic stability and plasma protein binding were performed in nude mice bearing metastasizing OH1 human small cell lung cancer xenografts. CXCR4 expression on OH1 tumor sections was determined by immunohistochemical staining. Results: nat Ga-CPCR4-2 exhibits high CXCR4 affinity with a half maximum inhibitory concentration of 4.99 6 0.72 nM. 68 Ga-CPCR4-2 showed high in vivo stability and high and specific tumor accumulation, which was reduced by approximately 80% in competition studies with AMD3100. High CXCR4 expression in tumors was confirmed by immunohistochemical staining. 68 Ga-CPCR4-2 showed low uptake in nontumor tissue and particularly low kidney accumulation despite predominant renal excretion, leading to high-contrast delineation of tumors in small-animal PET studies. Conclusion: The small and optimized cyclic peptide CPCR4-2 labeled with 68 Ga is a suitable tracer for targeting and imaging of human CXCR4 receptor expression in vivo. The high affinity for CXCR4, its in vivo stability, and the excellent pharmacokinetics recommend the further evaluation of 68 Ga-CPCR4-2 in a proof-of-concept study in humans.

Chemokines orchestrate leukocyte trafficking and function in health and disease. Heterophilic interactions between chemokines in a given microenvironment may amplify, inhibit, or modulate their activity; however, a systematic evaluation of the chemokine interactome has not been performed. We used immunoligand blotting and surface plasmon resonance to obtain a comprehensive map of chemokine-chemokine interactions and to confirm their specificity. Structure-function analyses revealed that chemokine activity can be enhanced by CC-type heterodimers but inhibited by CXC-type heterodimers. Functional synergism was achieved through receptor heteromerization induced by CCL5-CCL17 or receptor retention at the cell surface via auxiliary proteoglycan binding of CCL5-CXCL4. In contrast, inhibitory activity relied on conformational changes (in CXCL12), affecting receptor signaling. Obligate CC-type heterodimers showed high efficacy and potency and drove acute lung injury and atherosclerosis, processes abrogated by specific CCL5-derived peptide inhibitors or knock-in of an interaction-deficient CXCL4 variant. Atheroprotective effects of CCL17 deficiency were phenocopied by a CCL5-derived peptide disrupting CCL5-CCL17 heterodimers, whereas a CCL5 α-helix peptide mimicked inhibitory effects on CXCL12-driven platelet aggregation. Thus, formation of specific chemokine heterodimers differentially dictates functional activity and can be exploited for therapeutic targeting.

This report describes the design and synthesis of a series of a V b 3 integrin-directed monomeric, dimeric and tetrameric cyclo [Arg-Gly-Asp-D-Phe-Lys] dendrimers using "click chemistry". It was found that the unprotected N-e-azido derivative of cyclo[Arg-Gly-Asp-D-Phe-Lys] underwent a highly chemoselective conjugation to amino acid-based dendrimers bearing terminal alkynes using a microwave-assisted Cu(I)-catalyzed 1,3-dipolar cycloaddition. The a V b 3 binding characteristics of the dendrimers were determined in vitro and their in vivo a V b 3 targeting properties were assessed in nude mice with subcutaneously growing human SK-RC-52 tumors. The multivalent RGD-dendrimers were found to have enhanced affinity toward the a V b 3 integrin receptor as compared to the monomeric derivative as determined in an in vitro binding assay. In case of the DOTA-conjugated 111 In-labeled RGD-dendrimers, it was found that the radiolabeled multimeric dendrimers showed specifically enhanced uptake in a V b 3 integrin expressing tumors in vivo. These studies showed that the tetrameric RGD-dendrimer had better tumor targeting properties than its dimeric and monomeric congeners.