CPCR4-2

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-α), 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-Tyr(1)-[NMe]-D-Orn(2)-[4-(aminomethyl) benzoic acid,(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 ± 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.

Fostered by the clinical success of sst-ligands, the development and evaluation of (68)Ga-labeled peptides have become a very active field in radiopharmaceutical chemistry. Consequently, various new peptide tracers have been developed, e.g. [(68)Ga]CPCR4.2 for in vivo imaging of solid and haematological tumors or [(68)Ga]TRAP(RGD)₃ for imaging of α(v)β₃ integrin expression. As a consequence of different matrices (TiO₂, SnO₂, polymers) exploited in commercial (68)Ge/(68)Ga-generators, HCl of different concentrations (0.05...1.0 M) is used to obtain (68)Ga as starting material for automated syntheses. We have developed a purification method which reduces the eluate volume and adjusts the HCl concentration. The method may potentially allow standardization of the eluate composition of different commercial generators prior to labeling. Recently, a cationic purification process has been reported which allows the pre-fixation of (68)Ga on a Varian SCX cation exchange cartridge and subsequent elution of (68)Ga with acidified NaCl solutions. As part of the development of ready-to-use cassettes for the automated production of (68)Ga-CPCR4.2 using a SCINTOMICS GRP module and an iThemba Labs generator that is eluted with 0.6...1.0 M HCl, we tested and compared the (68)Ga-trapping efficiency of various commercial available cation exchange cartridges, the efficiency of subsequent (68)Ga-elution from these cartridges by means of various protocols and the influence of these variations on the labeling efficiency of [(68)Ga]CPCR4.2, [(68)Ga]TRAP(RGD)₃ and [(68)Ga]DOTATATE/[(68)Ga]DOTANOC. Finally, we transferred the optimized method to the automated, cassette based synthesis of [(68)Ga]CPCR4.2 and the aforementioned peptides. From seven tested cation exchange cartridges, Chromafix PS-H(+) gave the best extraction results (>95%). Moreover, we observed that acidified solutions of 5 M NaCl or 2.5 M CaCl₂ can be used for efficient cartridge elution. Using a disposable cGMP-compliant cassette system, we obtained [(68)Ga]CPCR4.2 in 80% decay-corrected yield and >99% purity. These data were confirmed by the production of [(68)Ga]DOTATATE, [(68)Ga]DOTANOC and [(68)Ga]TRAP(RGD)3 on the otherwise identical cassette system.