Imaging of HER2/neu expression in BT-474 human breast cancer xenografts in athymic mice using [99mTc]-HYNIC-trastuzumab (Herceptin) Fab fragments

Objective To evaluate the ability of trastuzumab (Herceptin) Fab, labelled with Tc-99m through introduced hydrazinenicotinamide (HYNIC) functionalities, to image HER2/neu-overexpressing human breast cancer xenografts in athymic mice. Methods Fab fragments were produced by immobilized papain digestion of trastuzumab immunoglobulin G (IgG), followed by purification by ultrafiltration. The immunoreactivity of trastuzumab Fab was evaluated by receptor-binding assays against HER2/neu-positive SK-BR-3 human breast cancer cells. Trastuzuma Alpha b Fab fragments were labelled with (TC)-T-99m following modification with HYNIC N-hydroxysuccinimide ester. Biodistribution and tumour imaging studies were performed in athymic mice bearing subcutaneous HER2/neu-overexpressing BT-474 human breast cancer xenografts following intravenous injection of 1.1 or 25 M Bq of [Tc-99m]-trastuzumab Fab (30 mu g), respectively. The specificity of tumour uptake was assessed by comparison with that of [Tc-99m]- labelled irrelevant antiCD33 HuM195 Fab. Results Trastuzumab Fab was pure and exhibited preserved immunoreactivity towards SK-BR-3 cells (K-d = 1.6 x 10(-8)M). Modification with HYNIC diminished its receptor-binding affinity fourfold. ((99m)Tcl-trastuzumab Fab localized avidly and specifically in BT-474 xenografts, achieving a tumour uptake of 10.7% of the injected dose (ID) per gram and a tumour to blood (T/B) ratio of 3: 1 at 24 h. The tumour uptake and T/B ratio for [Tc-99m]-trastuzumab Fab were significantly higher than those for control [Tc-99m]HuM195 Fab (2.6% ID - g(-1) and 0.9: 1, respectively; P < 0.05). Tumours were imaged as early as 2 h postinjection of [(99m)Tcl-trastuzumab Fab, but were more clearly visualized at 6 and 24 h post-injection. Conclusions [Tc-99m]-HYNIC-trastuzumab Fab localized specifically in HER2/neu-overexpressing human breast cancer xenografts in athymic mice, allowing imaging of the tumours within the useful lifetime of the radionuclide. (c) 2005 Lippincott Williams & Wilkins.