In Vivo Evaluation and Dosimetry Estimate for a High Affinity Affibody PET Tracer Targeting PD-L1

Purpose In vivo imaging of programmed death ligand 1 (PD-L1) during immunotherapy could potentially monitor changing PD-L1 expression and PD-L1 expression heterogeneity within and across tumors. Some protein constructs can be used for same-day positron emission tomography (PET) imaging. Previously, we evaluated the PD-L1-targeting Affibody molecule [ 18 F]AlF-NOTA-Z PD-L1_1 as a PET tracer in a mouse tumor model of human PD-L1 expression. In this study, we evaluated the affinity-matured Affibody molecule Z PD-L1_4 , to determine if improved affinity for PD-L1 resulted in increased in vivo targeting of PD-L1. Procedures Z PD-L1_4 was conjugated with NOTA and radiolabeled with either [ 18 F]AlF or 68 Ga. [ 18 F]AlF-NOTA-Z PD-L1_4 and [ 68 Ga]NOTA-Z PD-L1_4 were evaluated in immunocompromised mice with LOX (PD-L1+) and SUDHL6 (PD-L1-) tumors with PET and ex vivo biodistribution measurements. In addition, whole-body PET studies were performed in rhesus monkeys to predict human biodistribution in a model with tracer binding to endogenous PD-L1, and to calculate absorbed radiation doses. Results Ex vivo biodistribution measurements showed that both tracers had > 25 fold higher accumulation in LOX tumors than SUDHL6 ([ 18 F]AlF-NOTA-Z PD-L1_4 : LOX: 8.7 ± 0.7 %ID/g ( N = 4) SUDHL6: 0.2 ± 0.01 %ID/g ( N = 6), [ 68 Ga]NOTA-Z PD-L1_4 : LOX: 15.8 ± 1.0 %ID/g ( N = 6) SUDHL6: 0.6 ± 0.1 %ID/g ( N = 6)), considerably higher than Z PD-L1_1 . In rhesus monkeys, both PET tracers showed fast clearance through kidneys and low background signal in the liver ([ 18 F]AlF-NOTA-Z PD-L1_4 : 1.26 ± 0.13 SUV, [ 68 Ga]NOTA-Z PD-L1_4 : 1.11 ± 0.06 SUV). PD-L1-expressing lymph nodes were visible in PET images, indicating in vivo PD-L1 targeting. Dosimetry estimates suggest that both PET tracers can be used for repeated clinical studies, although high kidney accumulation may limit allowable radioactive doses. Conclusions [ 18 F]AlF-NOTA-Z PD-L1_4 and [ 68 Ga]NOTA-Z PD-L1_4 are promising candidates for same-day clinical PD-L1 PET imaging, warranting clinical evaluation. The ability to use either [ 18 F] or [ 68 Ga] may expand access to clinical sites.