High in-vivo stability in preclinical and first-in-human experiments with [F]AlF-RESCA-MIRC213: a F-labeled nanobody as PET radiotracer for diagnosis of HER2-positive cancers.

Purpose: [F]AlF-RESCA was introduced as a core particularly useful for F-labeling of heat-sensitive biomolecules. However, no translational studies have been reported up to now. Herein, we reported the first-in-human evaluation of an F-labeled anti-HER2 nanobody MIRC213 as a PET radiotracer for imaging HER2-positive cancers.

Methods: MIRC213 was produced by E. coli and conjugated with ( ±)-HRESCA-Mal. [F]AlF-RESCA-MIRC213 was prepared at room temperature. Its radiochemical purity and stability of were determined by radio-HPLC with the size-exclusion chromatographic column. Cell uptake was performed in NCI-N87 (HER2 +) and MCF-7 (HER2-) cells and the cell-binding affinity was verified in SK-OV-3 (HER2 +) cells. Small-animal PET/CT was performed using SK-OV-3, NCI-N87, and MCF-7 tumor-bearing mice at 30 min, 1 h, and 2 h post-injection. For blocking experiment, excess MIRC213 was co-injected with radiotracer. Biodistribution were performed on SKOV-3 and MCF-7 tumor-bearing mice at 2 h post-injection. For clinical study, PET/CT images were acquired at 2 h and 4 h after injection of [F]AlF-RESCA-MIRC213 (1.85-3.7 MBq/kg) in six breast cancer patients (3 HER2-positive and 3 HER2-negative). All patients underwent [F]-FDG PET/CT within a week for tissue selection purpose. Distribution and dosimetry were calculated. Standardized uptake values (SUV) were measured in tumors and normal organs.

Results: MIRC213 was produced with > 95% purity and modified with RESCA to obtain RESCA-MIRC213. [F]AlF-RESCA-MIRC213 was prepared within 20 min at room temperature with the radiochemical yield of 50.48 ± 7.6% and radiochemical purity of > 98% (n > 10), and remained stable in both PBS (88%) and 5% HSA (92%) after 6 h. The 2 h cellular uptake of [F]AlF-RESCA-MIRC213 in NCI-N87 cells was 11.22 ± 0.60 AD%/10 cells. Its binding affinity K value was determined to be 1.23 ± 0.58 nM. Small-animal PET/CT with [F]AlF-RESCA-MIRC213 can clearly differentiate SK-OV-3 and NCI-N87 tumors from MCF-7 tumors and background with a high uptake of 4.73 ± 1.18 ID%/g and substantially reduced to 1.70 ± 0.13 ID%/g for the blocking group (p < 0.05) in SK-OV-3 tumors at 2 h post-injection. No significant bone radioactivity was seen in the tumor-bearing animals. In all six breast cancer patients, there was no adverse reaction during study. The uptake of [F]AlF-RESCA-MIRC213 was mainly in lacrimal gland, parotid gland, submandibular gland, thyroid gland, gallbladder, kidneys, liver, and intestines. There was no significant bone radioactivity accumulation in cancer patients. [F]AlF-RESCA-MIRC213 had significantly higher tumor uptake in lesions from HER2-positive patients than that lesions from HER2-negative patients (SUVmax of 3.62 ± 1.56 vs. 1.41 ± 0.41, p = 0.0012) at 2 h post-injection. The kidneys received the highest radiation dose of 2.42 × 10 mGy/MBq, and the effective dose was 1.56 × 10 mSv/MBq.

Conclusions: [F]AlF-RESCA-MIRC213 could be prepared with high radiolabeling yield under mild conditions. [F]AlF-RESCA-MIRC213 has relatively high stability both in vitro and in vivo. The results from clinical transformation suggest that [F]AlF-RESCA-MIRC213 PET/CT is a safe procedure with favorable pharmacokinetics and dosimetry profile, and it is a promising new PET radiotracer for noninvasive diagnosis of HER2-positive cancers.