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Site-Specific Radiolabeling of a Human PD-L1 Nanobody via Maleimide–Cysteine Chemistry

In Vivo Cellular and Molecular Imaging Laboratory (ICMI), Medical Imaging Department (MIMA), Vrije Universiteit Brussel, 1090 Brussels, Belgium
Laboratory for Molecular and Cellular Therapy (LMCT), Department of Biomedical Sciences, Vrije Universiteit Brussel, 1090 Brussels, Belgium
Department of Nuclear Medicine, UZ Brussel, Laarbeeklaan 101, 1090 Brussels, Belgium
Author to whom correspondence should be addressed.
Academic Editor: Abdelwahab Omri
Pharmaceuticals 2021, 14(6), 550;
Received: 29 April 2021 / Revised: 2 June 2021 / Accepted: 4 June 2021 / Published: 8 June 2021
Immune checkpoint inhibitors targeting the programmed cell death-1 (PD-1) and its ligand PD-L1 have proven to be efficient cancer therapies in a subset of patients. From all the patients with various cancer types, only 20% have a positive response. Being able to distinguish patients that do express PD-1/PD-L1 from patients that do not allows patients to benefit from a more personalized and efficient treatment of tumor lesion(s). Expression of PD-1 and PD-L1 is typically assessed via immunohistochemical detection in a tumor biopsy. However, this method does not take in account the expression heterogeneity within the lesion, nor the possible metastasis. To visualize whole-body PD-L1 expression by PET imaging, we developed a nanobody-based radio-immunotracer targeting PD-L1 site-specifically labeled with gallium-68. The cysteine-tagged nanobody was site-specifically conjugated with a maleimide (mal)-NOTA chelator and radiolabeling was tested at different nanobody concentrations and temperatures. Affinity and specificity of the tracer, referred to as [68Ga]Ga-NOTA-mal-hPD-L1 Nb, were assayed by surface plasmon resonance and on PD-L1POS or PD-L1NEG 624-MEL cells. Xenografted athymic nude mice bearing 624-MEL PD-L1POS or PD-L1NEG tumors were injected with the tracer and ex vivo biodistribution was performed 1 h 20 min post-injection. Ideal 68Ga-labeling conditions were found at 50 °C for 15 min. [68Ga]Ga-NOTA-mal-hPD-L1 Nb was obtained in 80 ± 5% DC-RCY with a RCP > 99%, and was stable in injection buffer and human serum up to 3 h (>99% RCP). The in vitro characterization showed that the NOTA-functionalized Nb retained its affinity and specificity. Ex vivo biodistribution revealed a tracer uptake of 1.86 ± 0.67% IA/g in the positive tumors compared with 0.42 ± 0.04% IA/g in the negative tumors. Low background uptake was measured in the other organs and tissues, except for the kidneys and bladder, due to the expected excretion route of Nbs. The data obtained show that the site-specific 68Ga-labeled NOTA-mal-hPD-L1 Nb is a promising PET radio-immunotracer due to its ease of production, stability and specificity for PD-L1. View Full-Text
Keywords: cancer; nanobodies; PD-L1; site-specific; PET; gallium-68; maleimide-NOTA cancer; nanobodies; PD-L1; site-specific; PET; gallium-68; maleimide-NOTA
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MDPI and ACS Style

Chigoho, D.M.; Lecocq, Q.; Awad, R.M.; Breckpot, K.; Devoogdt, N.; Keyaerts, M.; Caveliers, V.; Xavier, C.; Bridoux, J. Site-Specific Radiolabeling of a Human PD-L1 Nanobody via Maleimide–Cysteine Chemistry. Pharmaceuticals 2021, 14, 550.

AMA Style

Chigoho DM, Lecocq Q, Awad RM, Breckpot K, Devoogdt N, Keyaerts M, Caveliers V, Xavier C, Bridoux J. Site-Specific Radiolabeling of a Human PD-L1 Nanobody via Maleimide–Cysteine Chemistry. Pharmaceuticals. 2021; 14(6):550.

Chicago/Turabian Style

Chigoho, Dora M., Quentin Lecocq, Robin M. Awad, Karine Breckpot, Nick Devoogdt, Marleen Keyaerts, Vicky Caveliers, Catarina Xavier, and Jessica Bridoux. 2021. "Site-Specific Radiolabeling of a Human PD-L1 Nanobody via Maleimide–Cysteine Chemistry" Pharmaceuticals 14, no. 6: 550.

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