68 Ga-Labeled Trastuzumab Fragments for ImmunoPET Imaging of Human Epidermal Growth Factor Receptor 2 Expression in Solid Cancers

Background: Trastuzumab, the first humanized antibody approved for therapeutic use has shown promising results for the treatment of patients with human epidermal growth factor receptor 2 (HER2) positive cancers. The aim of this study was to formulate immunoPET agents based on trastuzumab fragments and demonstrate their potential for early diagnosis of HER2-positive tumors. Materials and Methods: F(ab')₂ and F(ab') fragments of trastuzumab were prepared by enzymatic digestion and conjugated with chelator NOTA for labeling with ⁶⁸Ga. For comparison, intact trastuzumab was also radiolabeled. In vitro stability, immunoreactivity, and binding affinity of radio formulations toward HER2 receptors were evaluated by performing in vitro studies in cancer cell lines. Biodistribution and PET imaging studies were performed in animal model bearing tumors. Results: ⁶⁸Ga-NOTA-F(ab')-trastuzumab, ⁶⁸Ga-NOTA-F(ab')₂-trastuzumab, and ⁶⁸Ga-NOTA-trastuzumab could be prepared with >98% radiochemical purity (% RCP) and were found to be stable when studied up to 4 h. In vitro binding studies revealed high affinity and specificity of formulations toward HER2 receptors. Specific tumor uptake of ⁶⁸Ga-NOTA-F(ab')-trastuzumab and ⁶⁸Ga-NOTA-F(ab')₂-trastuzumab in HER2-positive tumors was observed in biodistribution and PET imaging studies. Conclusions: This study describes optimization of protocol for the formulation of ⁶⁸Ga-NOTA-F(ab')-trastuzumab and ⁶⁸Ga-NOTA-F(ab')₂-trastuzumab for targeting HER2-overexpressing tumors. Further studies with these radioformulations are warranted to confirm their potential as immunoPET agents for management of HER2-positive breast and other solid tumors.