Amplification of DNA damage by a γH2AX-targeted radiopharmaceutical

In-111-DTPA-anti-gamma H2AX-Tat, which combines an anti-gamma H2AX antibody with a cell-penetrating peptide, Tat, and the Auger electron-emitting radioisotope, In-111, targets the DNA damage signalling protein, gamma H2AX, and has potential as a probe for imaging DNA damage in vivo. The goal of this study was to investigate whether In-111-DTPA-anti-gamma H2AX-Tat labelled to high specific activity (6 MBq/mu g) can amplify treatment-related DNA damage for therapeutic gain. Methods: MDA-MB-468 and MDA-MB-231/H2N (231-H2N) breast cancer cells were incubated with In-111-DTPA-anti-gamma H2AX-Tat (3MBq, 6MBq/mu g) or a control radioimmunoconjugate, In-111-DTPA-mIgG-Tat, and exposed to IR or bleomycin. DNA damage was studied by counting gamma H2AX foci and by neutral comet assay. Cytotoxicity was evaluated using clonogenic assays. In-111-DTPA-anti-gamma H2AX-Tat was administered intravenously to 231-H2N-xenograft-bearing Balb/c nu/nu mice in tumor growth inhibition studies. Results: The number of gamma H2AX foci was greater after exposure of cells to IR (10Gy) plus In-111-DTPA-anti-gamma H2AX-Tat compared to IR alone (20.6 +/- 2.5 versus 10.4 +/- 2.3 foci/cell; P<.001). In-111-DTPA-anti-gamma H2AX-Tat resulted in a reduced surviving fraction in cells co-treated with IR (4Gy) versus IR alone (5.2%+/- 0.9% versus 47.8%+/- 2.8%; P<.001). Similarly, bleomycin (25-200 mu g/mL) plus In-111-DTPA-anti-gamma H2AX-Tat resulted in a lower SF compared to bleomycin alone. The combination of a single exposure to IR (10Gy) plus (111)InDTPA-anti-gamma H2AX-Tat significantly decreased the growth rate of 231-H2N xenografts in vivo compared to either In-111-DTPA-anti-gamma H2AX-Tat or IR alone (-0.002 +/- 0.004 versus 0.036 +/- 10.011 and 0.031 +/- 0.014 mm(3)/day, respectively, P<.001). Conclusion: In-111-DTPA-anti-gamma H2AX-Tat amplifies anticancer treatment-related DNA damage in vitro and has a potent anti-tumor effect when combined with IR in vivo. (C) 2012 Elsevier Inc. All rights reserved.