Development 68Ga trastuzumab Fab and bioevaluation by PET imaging in HER2/neu expressing breast cancer patients.

Yogesh Rathore, Jaya Shukla, Ishita Laroiya, Amar Deep, Tamanna Lakhanpal, Rajender Kumar, Harmandeep Singh, Amanjit Bal, Gurpreet Singh, Krishan Gopal Thakur, B R Mittal

Nuclear medicine communications 2022 Apr 01

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Receptors on breast cancer cells play a crucial role in the management of patients. Trastuzumab is a widely used drug for the treatment of HER2/neu expressing tumors. ImmunoPET with trastuzumab is not feasible due to slow pharmacokinetics. Fragment of antigen-binding (Fab) radiolabeled with positron emitters can be used for immunoPET. Fab has been generated by papain digestion and conjugated with the bifunctional chelating agent NOTA. The SDS-PAGE and MALDI-TOF were used to see the integrity of Fab and conjugated Fab. In-vitro stability and target specificity for HER2/neu receptors were performed in plasma and receptor binding with bio-layer interferometry (BLI) techniques. Radiolabeling was standardized with 68GaCl3 and PET imaging was performed in seven patients showing 18F fluorodeoxyglucose (18F-FDG) uptake and correlated with HER2/neu expression by immunohistochemistry. Fab production was optimized at molar ratio 23:1 of trastuzumab and papain at 37 °C with a constant stirrer at 850 rpm for 22-24 h, at pH 8. Conjugation with NOTA was standardized at molar ratio 1:25 of trastuzumab Fab and NOTA. Molecular mass of trastuzumab Fab-NOTA was found approximately 46.3 kDa (~1/3 of intact antibody). Trastuzumab Fab-NOTA showed radiolabelling efficiency of 48-70% with incubation time 15 min at 37-40 °C and pH 4.5-5.0. BLI demonstrated the affinity of trastuzumab, trastuzumab Fab and trastuzumab Fab-NOTA towards HER2/neu receptor with KD of <1pM, ~0.5nM and ~20nM, respectively. All immunohistochemistry proven patients showed uptake in primary breast lesion and lymph nodes. Trastuzumab Fab-NOTA is suitable for radiolabelling with 68Ga and ImmunoPET imaging of HER2/neu receptor. Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.

Citation

Yogesh Rathore, Jaya Shukla, Ishita Laroiya, Amar Deep, Tamanna Lakhanpal, Rajender Kumar, Harmandeep Singh, Amanjit Bal, Gurpreet Singh, Krishan Gopal Thakur, B R Mittal. Development 68Ga trastuzumab Fab and bioevaluation by PET imaging in HER2/neu expressing breast cancer patients. Nuclear medicine communications. 2022 Apr 01;43(4):458-467