Examination of the metabolism of both MCT-imaging PET radioligands [18F]FACH and [18F]FACH-d6 in vitro by means of LC-MS/MS - support for understanding preclinical in vivo data from different species

Objectives: Positron Emission Tomography (PET) is a non-invasive molecular imaging technique, which continues to find increasing application and further development for diagnosis of a broad range of disorders and dysfunctions. Since their involvement in pathophysiological processes that lead to cancer, for example, monocarboxylate transporters (MCTs) are of interest for the development of new radioligands for PET, such as [18F]FACH [1, 2] and its deuterated isotopologue [18F]FACH-d6. Both candidates have been already examined previously in preclinical PET studies in healthy mice and piglets, also with regard to their biodistribution and metabolic fate [3]. However, the general finding that in piglets both radioligands showed significant lower metabolic stability than in mice left some important unanswered questions, especially those relevant to potential clinical application. Therefore, this study aimed to further explore the metabolic properties of both radioligands by means of in vitro examinations, which besides discovering species dependency in some respects, might allow predictions for their metabolic stability in humans. Methods: [18F]FACH, [18F]FACH-d6 as well as their corresponding non-radioactive references were incubated with murine (mouse MLM and rat RLM) and human (HLM) liver microsomes and with pig and human S9 fractions, in presence of NADPH, but also other co-factors (e.g. UDPGA). After collection of samples at defined time points, subsequently prepared samples were investigated by radio-HPLC, UV-HPLC or LC-MS/MS. Obtained data allowed calculation of basic metabolic parameters and identification of metabolites formed in vitro. Results: [18F]FACH, incubated in presence of NADPH at 37°C for up to 60 min, showed highest stability in MLM (t1/2= 29913 min), whereas lowest stability was found in RLM (t1/2= 201 min). A moderate stability was found for both HLM (t1/2= 948 min) and pig S9 fraction (t1/2= 522 min). However, in a recent head-to-head comparis