Automated radiofluorination of HER2 single domain antibody: the road towards the clinical translation of [18F]FB-HER2 sdAb

Background: With the next generation of Human Epidermal Growth Factor Receptor 2 (HER2) -targeting therapies, such as antibody-drug conjugates, showing benefit in "HER2 low" and even "HER2 ultralow" patients, the need for novel methods to quantify HER2 expression accurately becomes even more important for clinical decision making. A HER2 PET/CT imaging assessment could evaluate HER2 positive disease locations while improving patient care, reducing the need for invasive biopsies. A single-domain antibody (sdAb)-based PET tracer could combine the high specificity of sdAbs with short-lived radionuclides such as fluorine-18 (F-18) and gallium-68 (Ga-68). SdAb-based PET tracers have clinically been used via a Ga-68-chelator approach. However, the distribution of Ga-68-labelled pharmaceuticals to peripheral PET centres is more challenging to organize due to the short half-life of Ga-68, most certainly when the available activity is limited by a generator. Cyclotron produced Ga-68 has removed this limitation. Distribution of F-18-labelled pharmaceuticals remains less challenging due to its slightly longer half-life, and radiofluorination of sdAbs via N-succinimidyl-4-[F-18]fluorobenzoate ([F-18]SFB) has shown to be a promising strategy for developing sdAb-based PET tracers. Although [F-18]SFB automation has been reported, automating protein conjugation proves challenging. Herein we report the fully automated, cartridge-based production of [F-18]FB-HER2 sdAb on a single synthesis module. Results: [F-18]FB-HER2 sdAb (> 6 GBq) was obtained after a fully automated production (95 min), with a RCP > 95%, apparent molar activity > 20 GBq/mu mol and decay-corrected radiochemical yield (RCY d.c.) of 14 +/- 2% (n = 4). Further upscaling amounted to production batches of 16 GBq with an apparent molar activity > 40 GBq/mu mol and RCY d.c. of 8 +/- 1% (n = 4). Ex vivo biodistribution and PET imaging showed specific HER2-positive tumour targeting and low kidney retention. Conclusion: The [F-18]FB-HER2 sdAb tracer was produced with clinically relevant activities using a fully automated production method. The automated production method was designed to ease the translation to the clinic and has the potential to be used not only in mono-centre but also multi-centre clinical trials with one central production site. [F-18]FB-HER2 sdAb showed a favourable biodistribution pattern and could be a valuable alternative to Ga-68-labelled sdAb-based PET tracers in the clinic.