Panitumumab labeled with Auger electron-emitting 197gHg/197mHg: cytotoxicity on human breast cancer cells and tumour and normal tissue uptake in mice with EGFR-overexpressing breast cancer xenografts

197g Hg and 197m Hg are attractive radionuclides for radioimmunotherapy of cancer due to their abundant Auger electron emissions (23.2 and 19.4 electrons/decay, respectively). Our aim was to study the cytotoxicity and DNA-damaging properties of panitumumab labeled with 197g Hg/ 197m Hg on a panel of human breast cancer cells expressing different levels of epidermal growth factor receptor (EGFR) (10 4 to 10 6 EGFR/cell) and evaluate the tumour and normal tissue uptake of these radioimmunoconjugates in mice with EGFR-overexpressing MDA-MB-468 human breast cancer xenografts (10 6 EGFR/cell). Panitumumab was conjugated to 2-[4,7,10-tris(2-amino-2-oxoethyl)-1,4,7,10-tetrazacyclododec-1-yl]acetamide (TCMC) or a sulphur-rich (NS 4 ) bifunctional chelator to complex 197g Hg/ 197m Hg or directly labeled with 197g Hg/ 197m Hg through endogenous mercury binding sites. All radioimmunoconjugates bound specifically (87–91%) to EGFR on MDA-MB-468 cells. The radioimmunoconjugates were bound, internalized and imported into the nucleus of breast cancer cells dependent on their EGFR expression. Absorbed doses in the nucleus from the Auger electron emissions of 197g Hg/ 197m Hg were correlated with EGFR expression level. The radioimmunoconjugates were most cytotoxic to MDA-MB-468 cells reducing their clonogenic survival to < 5% while unlabeled immunoconjugates did not decrease survival. Blocking EGFR with excess panitumumab restored clonogenic survival to ~ 60%. DNA double-strand breaks were highest in MDA-MB-468 cells and were decreased by blocking EGFR. Subcutaneous MDA-MB-468 tumours in athymic (nude) mice were imaged by SPECT/CT at 1 d and 3 d post-injection of the radioimmunoconjugates. Tumour uptake on images and in biodistribution studies was decreased by co-administering an excess of panitumumab. High kidney uptake was observed. Panitumumab was complexed to 197g Hg/ 197m Hg after conjugation to a bifunctional chelator or labeled directly with 197g Hg/ 197m Hg. These radioimmunoconjugates bound specifically to EGFR on human breast cancer cells and caused DNA double-strand breaks that decreased their clonogenic survival. The radioimmunoconjugates localized in EGFR-overexpressing MDA-MB-468 tumours in mice but there was high kidney uptake, suggesting in vivo release of some 197g Hg/ 197m Hg. Competition between complexation of 197g Hg/ 197m Hg to a bifunctional chelator and binding to endogenous mercury binding sites in panitumumab presents challenges for radiolabeling and may explain the apparent release of some 197g Hg/ 197m Hg in vivo. Alternative labeling strategies such as a 2-step method involving conjugation of the preformed 197g Hg/ 197m Hg bifunctional chelator to panitumumab could be explored to address these challenges.