89Zr-Labeled Anti-PD-L1 Antibody PET Monitors Gemcitabine Therapy-Induced Modulation of Tumor PD-L1 Expression

• Published in: The Journal of nuclear medicine (1978) Vol. 62; no. 5; pp. 656 - 664
• Main Authors: Jung, Kyung-Ho, Park, Jin Won, Lee, Jin Hee, Moon, Seung Hwan, Cho, Young Seok, Lee, Kyung-Han
• Format: Journal Article
• Language: English
• Published: New York Society of Nuclear Medicine 10.05.2021
• Subjects: AKT protein; Antibodies; Apoptosis; Basic Science Investigation; Binding; Blood; Chemotherapy; Cold treatment; Colon; Colon cancer; Colorectal cancer; Computed tomography; Conjugation; Deferoxamine; Disulfide bonds; Dosage; Electrophoresis; Flow cytometry; Gel electrophoresis; Gemcitabine; Homology; Image analysis; Image processing; Immune clearance; Intravenous administration; Ionization; Kinases; L1 protein; Medical imaging; Modulation; PD-L1 protein; Pharmacokinetics; Polyacrylamide; Positron emission; Proteins; PTEN protein; Radiolabelling; Reduction; Sodium dodecyl sulfate; Sodium lauryl sulfate; Tensin; Tomography; Tumors; Western blotting; Zirconium isotopes
• ISSN: 0161-5505; 1535-5667; 1535-5667
• DOI: 10.2967/jnumed.120.250720

We developed an 89Zr-labeled anti–programmed death ligand 1 (anti-PD-L1) immune PET that can monitor chemotherapy-mediated modulation of tumor PD-L1 expression in living subjects. Methods: Anti-PD-L1 underwent sulfohydryl moiety–specific conjugation with maleimide-deferoxamine followed by 89Zr radiolabeling. CT26 colon cancer cells and PD-L1–overexpressing CT26/PD-L1 cells underwent binding assays, flow cytometry, and Western blotting. In vivo pharmacokinetics, biodistribution, and PET imaging were evaluated in mice. Results: 89Zr-anti-PD-L1 synthesis was straightforward and efficient. Sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that reduction produced half-antibody fragments, and matrix-assisted laser desorption ionization time-of-flight analysis estimated 2.18 conjugations per antibody, indicating specific conjugation at the hinge-region disulfide bonds. CT26/PD-L1 cells showed 102.2 ± 6.7-fold greater 89Zr-anti-PD-L1 binding than that of weakly expressing CT26 cells. Excellent target specificity was confirmed by a drastic reduction in binding by excess cold antibody. Intravenous 89Zr-anti-PD-L1 followed biexponential blood clearance. PET/CT image analysis demonstrated decreases in major organ activity over 7 d, whereas high CT26/PD-L1 tumor activity was maintained. Again, this was suppressed by excess cold antibody. Treatment of CT26 cells with gemcitabine for 24 h augmented PD-L1 protein to 592.4% ± 114.2% of the control level and increased 89Zr-anti-PD-L1 binding, accompanied by increased AKT (protein kinase B) activation and reduced phosphatase and tensin homolog (PTEN). In CT26 tumor–bearing mice, gemcitabine treatment substantially increased tumor uptake from 1.56% ± 0.48% to 6.24% ± 0.37% injected dose per gram (tumor-to-blood ratio, 34.7). Immunoblots revealed significant increases in tumor PD-L1 and activated AKT and a decrease in PTEN. Conclusion: 89Zr-anti-PD-L1 showed specific targeting with favorable imaging properties. Gemcitabine treatment upregulated cancer cell and tumor PD-L1 expression and increased 89Zr-anti-PD-L1 uptake. 89Zr-anti-PD-L1 PET may thus be useful for monitoring chemotherapy-mediated tumor PD-L1 modulation in living subjects.