Dual-targeting CD73/PD-L1 bifunctional inhibitor: a promising cancer immunotherapy strategy

This work aims to design and characterize a novel bifunctional small molecule that simultaneously targets PD-L1 and CD73 to enhance the therapeutic efficacy of tumor immunotherapy.

Multiple methodologies were integrated for compound screening and biological characterization, including computer-aided molecular docking, homogeneous time-resolved fluorescence (HTRF) binding assay, surface plasmon resonance (SPR), and PD-1/PD-L1 NFAT reporter cell assay.

The lead compound CP-1 exhibited potent dual-target inhibitory activities. It blocked the PD-1/PD-L1 interaction with an IC₅₀ of 10.27 nM and suppressed CD73 activity with an IC₅₀ of 300.2 nM. Molecular docking simulations revealed that CP-1 stably binds to the functional domains of PD-L1 and CD73 via specific non-covalent interactions. Cellular functional assays further demonstrated that CP-1 effectively restored T cell function in the PD-1/PD-L1 reporter system, with an EC₅₀ of 0.9 μM.

CP-1 exhibits balanced, dual-nanomolar inhibitory activity against PD-L1 and CD73 and displays potent immunomodulatory effects at the cellular level. It serves as a promising lead candidate for developing novel bifunctional agents to advance tumor immunotherapy.