Colorectal cancer is one of the leading causes of cancer-related deaths worldwide. Increased expression of CXCR4 has been associated with liver metastasis, disease progression, and shortened survival. Using in vitro cell binding studies and the in ovo model, we aimed to investigate the potential of [⁶⁸Ga]Ga-Pentixafor, a radiotracer specifically targeting human CXCR4, for CRC imaging. Specific membrane binding and internalisation of [⁶⁸Ga]Ga-Pentixafor was shown for HT29 cells, but not for HCT116 cells. Accordingly, [⁶⁸Ga]Ga-Pentixafor accumulated specifically in CAM-xenografts derived from HT29 cells, but not in HCT116 xenografts, as determined by µPET/MRI. The CAM-grown xenografts were histologically characterised, demonstrating vascularisation of the graft, preserved expression of human CXCR4, and viability of the tumour cells within the grafts. In vivo viability was further confirmed by µPET/MRI measurements using 2-[¹⁸F]FDG as a surrogate for glucose metabolism. [⁶⁸Ga]Ga-Pentixafor µPET/MRI scans showed distinct radiotracer accumulation in the chick embryonal heart, liver, and kidneys, whereas 2-[¹⁸F]FDG uptake was predominantly found in the kidneys and joints of the chick embryos. Our findings suggest that [⁶⁸Ga]Ga-Pentixafor is an interesting novel radiotracer for CRC imaging that is worth further investigation. Moreover, this study further supports the suitability of the CAM-xenograft model for the initial preclinical evaluation of targeted radiopharmaceuticals.

The first-in-class ruthenium-based chemotherapeutic agent BOLD-100 (formerly IT-139, NKP-1339, KP1339) is currently the subject of clinical evaluation for the treatment of gastric, pancreatic, colorectal and bile duct cancer. A radiolabeled version of the compound could present a helpful diagnostic tool. Thus, this study investigated the pharmacokinetics of BOLD-100 in more detail to facilitate the stratification of patients for the therapy. The synthesis of [¹⁰³Ru]BOLD-100, radiolabeled with carrier added (c.a.) ruthenium-103, was established and the product was characterized by HPLC and UV/Vis spectroscopy. In order to compare the radiolabeled and non-radioactive versions of BOLD-100, both complexes were fully evaluated in vitro and in vivo. The cytotoxicity of the compounds was determined in two colon carcinoma cell lines (HCT116 and CT26) and biodistribution studies were performed in Balb/c mice bearing CT26 allografts over a time period of 72 h post injection (p.i.). We report herein preclinical cytotoxicity and pharmacokinetic data for BOLD-100, which were found to be identical to those of its radiolabeled analog [¹⁰³Ru]BOLD-100.

Personalized treatment of cancer patients demands specific and validated biomarkers for tumor diagnosis and therapy. The development and validation of such require translational preclinical models that recapitulate human diseases as accurately as possible. Moreover, there is a need for convergence of different (pre)clinical disciplines that openly share their knowledge and methodologies. This review sheds light on the differential perception of biomarkers and gives an overview of currently used models in tracer development and approaches for biomarker discovery.

The organometallic technetium-99m tricarbonyl core, [^99mTc][Tc(CO)₃(H₂O)₃]⁺, is a versatile precursor for the development of radiotracers for single photon emission computed tomography (SPECT). A drawback of the ^99mTc-tricarbonyl core is its lipophilicity, which can influence the pharmacokinetic properties of the SPECT imaging probe. Addition of polar pharmacological modifiers to ^99mTc-tricarbonyl conjugates holds the promise to counteract this effect and provide tumor-targeting radiopharmaceuticals with improved hydrophilicities, e.g., resulting in a favorable fast renal excretion in vivo. We applied the “Click-to-Chelate” strategy for the assembly of a novel ^99mTc-tricarbonyl labeled conjugate made of the tumor-targeting, modified bombesin binding sequence [Nle¹⁴]BBN(7–14) and the carbohydrate sorbitol as a polar modifier. The ^99mTc-radiopeptide was evaluated in vitro with PC-3 cells and in Fox-1^nu mice bearing PC-3 xenografts including a direct comparison with a reference conjugate lacking the sorbitol moiety. The glycated ^99mTc-tricarbonyl peptide conjugate exhibited an increased hydrophilicity as well as a retained affinity toward the Gastrin releasing peptide receptor and cell internalization properties. However, there was no significant difference in vivo in terms of pharmacokinetic properties. In particular, the rate and route of excretion was unaltered in comparison to the more lipophilic reference compound. This could be attributed to the intrinsic properties of the peptide and/or its metabolites. We report a novel glycated (sorbitol-containing) alkyne substrate for the “Click-to-Chelate” methodology, which is potentially of general applicability for the development of ^99mTc-tricarbonyl based radiotracers displaying an enhanced hydrophilicity.

[¹¹C]SNAP-7941 and its radiofluorinated, fluoro-ethyl derivative [¹⁸F]FE@SNAP have been developed as the first positron emission tomography tracers for melanin-concentrating hormone receptor 1 (MCHR1) imaging. Accumulation of these MCHR1 PET-tracers in rat brown adipose tissue (BAT) in vivo provided first indication of MCHR1 expression in rodent BAT. To rule out off-target binding, affinity of both MCHR1 ligands toward adrenergic beta-3 receptors (ADRB3) was examined. Further, specific binding of [¹¹C]SNAP-7941 to brown adipocytes and effects of MCHR1 ligands on brown adipocyte activation were investigated. SNAP-7941 and FE@SNAP evinced to be highly selective toward MCHR1. [¹¹C]SNAP-7941 binding to brown adipocytes was shown to be mainly MCHR1-specific. This data strongly indicates MCHR1 expression in rodent BAT and moreover, a peripheral, anti-obesity effect of MCHR1 antagonists directly exerted in BAT is proposed. Moreover, MCHR1 expression in murine brown adipocytes was confirmed by protein and mRNA analysis. We conclude that MCHR1 PET imaging contributes to basic research in endocrinology by elucidating the involvement of the MCH system in peripheral tissues, such as BAT.

The fertilised chicken egg (in ovo) is a promising, ethically acceptable intermediate model for preselection of radiotracers prior to testing in rodents. The behaviour of monoclonal antibodies in ovo for application in positron emission tomography is yet to be explored. Here, we investigated the potential of this model for development of radiolabelled antibodies using simultaneous PET and magnetic resonance imaging. The biodistribution of the model antibody [⁸⁹Zr]Zr-nSuc-Df-Denosumab was assessed at different time points and specific activities. The study was complemented with the more established [⁸⁹Zr]Zr-nSuc-Df-Atezolizumab. Both ⁸⁹Zr-labelled antibodies were synthesised with radiochemical yields >70 %, radiochemical purities >98 % and immune reactive fractions greater than 99 %. [⁸⁹Zr]Zr-nSuc-Df-Denosumab showed initial uptake in the chick embryonic heart, spleen and liver, while brain accumulation was negligible at all investigated time points. Using radiotracer in lower specific activities appeared to slow down biodistribution and clearance. By 23 h p.i., most activity was localised in the gallbladder, suggesting hepatobiliary excretion. A comparable biodistribution profile was observed for [⁸⁹Zr]Zr-nSuc-Df-Atezolizumab at 19 h p.i. The rapid clearance of both IgG antibodies is likely caused by the absence of neonatal Fc receptors (FcRn) in birds, which in mammals mediate IgG recycling and thereby prolong antibody half-life. In conclusion, while the overall biodistribution patterns were broadly comparable to those reported in mammals, this first exploratory investigation of ⁸⁹Zr-labelled antibodies in ovo revealed markedly altered pharmacokinetics in chick embryos, most likely due to the absence of FcRn. These findings indicate predominantly non-specific distribution and clearance of these radiolabelled antibodies in fertilised chicken eggs. This study underscores the importance of considering species-specific differences when interpreting in ovo data and the necessity of further research into the suitability of the in ovo model for evaluating different classes of radiotracers.