⁶⁴Cu PET Imaging of the CXCR4 Chemokine Receptor Using a Cross-Bridged Cyclam Bis-Tetraazamacrocyclic Antagonist

Abstract

Expression of the chemokine receptor chemokine C-X-C motif receptor 4 (CXCR4) plays an important role in cancer metastasis, in autoimmune diseases, and during stem cell-based repair processes after stroke and myocardial infarction. Previously reported PET imaging agents targeting CXCR4 suffer from either high nonspecific uptake or bind only to the human form of the receptor. The objective of this study was to develop a high-stability 64 Cu-labeled smallmolecule PET agent for imaging both human and murine CXCR4 chemokine receptors. Methods: Synthesis, radiochemistry, stability and radioligand binding assays were performed for the novel tracer 64 Cu-CuCB-bicyclam. In vivo dynamic PET studies were performed on mice bearing U87 (CXCR4 low-expressing) and U87.CXCR4 (human-CXCR4 high-expressing) tumors. Biodistribution and receptor blocking studies were performed on CD1-IGS immunocompetent mice. CXCR4 expression on tumor and liver disaggregates was confirmed using a combination of immunohistochemistry, quantitative polymerase chain reaction, and Western blot. Results: 64 Cu-CuCB-bicyclam has a high affinity for both the human and the murine variants of the CXCR4 receptor (half-maximal inhibitory concentration, 8 nM [human]/2 nM [murine]) and can be obtained from the parent chelator that has low affinity. In vitro and in vivo studies demonstrate specific uptake in CXCR4-expressing cells that can be blocked by more than 90% using a higher-affinity antagonist, with limited uptake in non-CXCR4-expressing organs and high in vivo stability. The tracer was also able to selectively displace the CXCR4 antagonists AMD3100 and AMD3465 from the liver. Conclusion: The tetraazamacrocyclic small molecule 64 Cu-CuCB-bicyclam has been shown to be an imaging agent for the CXCR4 receptor that is likely to be applicable across a range of species. It has high affinity and stability and is suitable for preclinical research in immunocompetent murine models.

A new class of PentixaFor- and PentixaTher-based theranostic agents with enhanced CXCR4-targeting efficiency

“…One potential reason for this observation might lie in the enhanced mCXCR4 affinity of the novel a-r-ABA-constructs; as demonstrated previously, enhanced and sometimes even dramatic (> 40% iD/g) liver accumulation of CXCR4-targeted tracers in mice is always observed for ligands with affinity for mCXCR4 56 , 60 . The finding that this uptake is (at least partially) blockable by an excess of unlabeled competitor such as AMD3100 (see Table S1 ) suggests involvement of a specific uptake mechanism for CXCR4-targeted tracers in the mouse liver based on hepatic mCXCR4 expression 61 .…”

“…It is important to note at this point, however, that due to its enhanced affinity towards mCXCR4, the biodistribution of [ 177 Lu]DOTA-r-a-ABA- iodo CPCR4 is “biased” with respect to background accumulation in comparison to the analogs with significantly lower mCXCR4 binding affinity, [ 177 Lu]PentixaTher and [ 177 Lu]DOTA-r-a-ABA-CPCR4 (Table 1 and Figure 5 ). It has been shown for other radioligands with high mCXCR4 affinity, that tracer uptake in liver, spleen, lung and bone (femur harboring bone marrow) is blockable with an excess of cold competitor 56 , 60 . In the case of [ 177 Lu]DOTA-r-a-ABA- iodo CPCR4, a blocking study has not been performed.…”

Influence of Multiple Animal Scanning on Image Quality for the Sedecal SuperArgus2R Preclinical PET Scanner

“…The primary application of preclinical PET at the University of Hull PET Research center is to assess the pharmacokinetics and biodistribution of novel imaging probes [36][37][38][39], for this application the degradation in image quality predicted for the two animal case is acceptable. For the application of existing tracers to provide physiological readouts in disease models, our data suggest that higher injected doses provide better image quality overall if dual animal scanning is warranted for logistical reasons [40].…”

Positron Emission Tomography for Response Evaluation in Microenvironment-Targeted Anti-Cancer Therapy

“…Since CXCR4 is known to be expressed in various CSCs and is associated with tumorigenicity, angiogenesis, invasion, and chemoresistance [ 144 ], CXCR4-directed therapies have been exploited using either alternative small-molecule CXCR4 antagonists [ 155 ], CXCL12 inhibitors [ 156 ], or anti-CXCR4 antibodies [ 157 ]. Radiolabeled compounds targeting CXCR4 have also been examined with lutetium-177 labeled pentixather, copper-64 labeled plerixafor, and another small molecule [ 158 , 159 , 160 ]. Fluorine-18 labeled pentixafor analog has recently shown high CXCR4 affinity and favorable tumor-to-normal organ ratios in a murine model of human lymphoma [ 161 ].…”