Synthesis and Preliminary Evaluations of [F]fluorinated Pyridine-2- Carboxamide Derivatives for Targeting PD-L1 in Cancer

Background: Treatment with immune checkpoint inhibitors has improved both progressionfree survival and overall survival in a subset of patients with tumors. However, the selection of patients who benefit from immune checkpoint inhibitor treatment remains challenging. Positron Emission Tomography (PET) is a non-invasive molecular imaging tool that offers a promising alternative to the current IHC for detecting the PD-L1 expression in malignant cells in vivo, enabling patient selection and predicting the response to individual patient immunotherapy treatment.

Objectives: Herein, we report the development of novel [F]labeled pyridine-2-carboxamide derivatives [F]2 and [F]3 as small-molecule probes for imaging immune checkpoint (PD-1/PD-L1) in cancer using PET.

Methods And Results: [F]2 and [F]3 were prepared by a one-step radiofluorination in 44 ± 5% and 30 ± 4% radiochemical yield and > 98% radiochemical purity for a potential clinical translation. The total synthesis time, including HPLC purification, was less than 45 min. [F]2 and [F]3 showed excellent stability in injection solution and a significant accumulation and retention in PD-1/PD-L1 expressing MDA-MB-231 breast cancer and in HeLa cervix carcinoma cells (2- 5 cpm/1000 cells). In addition, autoradiographic analysis and inhibition experiments on tumor slices confirm the potential of both compounds as specific imaging probes for the PD-1/PD-L1 axis in tumors.

Conclusion: The in vitro evaluation in PD-L1 expressing cells together with results from autoradiographic analysis in PD-L1 positive tumor sections, suggest that [F]2 and [F]3 could be potential imaging probes for assessing PD-L1 expression in tumors and warrant further biological evaluations in vivo.