| Objective: Trastuzumab modified with nuclear translocation sequence (NLS) peptides and diethylenetriaminepentaacetic acid (DTPA) for complexing 111In (111In-DTPA-NLS-trastuzumab) has shown promise for Auger electron radioimmunotherapy of HER2-positive breast cancer (BC), but a limitation is the low specific activity (SA) achievable for labeling with 111In. Our objective was to synthesize and characterize trastuzumab modified site-specifically with a metal chelating polymer (MCP) that presents multiple DTPA chelators for complexing 111In to increase the SA and the amount of 111In delivered to HER2-overexpressing BC cells.;Methods: The first set of MCPs was synthesized with a polyglutamide backbone with 24 or 29 DTPA units, with or without NLS peptide modification, and a terminal hydrazide group (Hy-) for reaction with aldehydes generated by sodium meta periodate (NaIO4)-oxidation of glycans on the Fc domain of trastuzumab. The maximum SA for 111 In-labeling of trastuzumab modified with a MCP [trastuzumab-Hy-MCP-(NLS)] was determined. Cytotoxicity was evaluated by clonogenic assays in HER2-positive BC cell lines. The second set of MCPs was synthesized with a polyglutamide backbone with 23 or 30 DTPA units, with or without 7 polyethylene glycol (PEG) chains [trastuzumab-Hy-(PEG)-MCP]. Tumor and normal tissue accumulation of trastuzumab-Hy-(PEG)-MCP-111In, its elimination from the blood, and its normal tissue toxicity were evaluated in mice.;Results: The maximum SA for labeling trastuzumab-Hy-MCP-111In was 90-fold greater than for trastuzumab modified with two DTPA. Clonogenic survival (CS) of HER2 overexpressed SK-BR-3 cells (1.3x106 receptor/cell), ZR-75-1 cells with intermediate HER2 density (4x105 receptor/cell) but no HER2 gene amplification, and HER2 overexpressed (5x105 receptor/cell) but trastuzumab resistant TrR1 cells were decreased to 1.8 +/- 1.3%, 20.5 +/- 4.3% and 17.1 +/- 1.6% respectively by high SA trastuzumab-Hy-MCP- 111In (20 nmol/L). Trastuzumab-Hy-PEG-MCP-111In achieved 1.6 fold higher HER2-mediated uptake than trastuzumab-Hy-MCP-111In (1.3 +/- 0.3 %ID/g vs. 0.8 +/- 0.4 %ID/g) in s.c HER2 overexpressed BT-474 xenografts in mice. In non tumor bearing mice, kidney uptake of trastuzumab-Hy-PEG-MCP- 111In was 5.5-fold lower than trastuzumab-Hy-MCP-111In (3.0 +/- 0.4 % ID/g vs. 16.6 +/- 4.3 %ID/g). There was no normal tissue toxicity of trastuzumab-Hy-PEG-MCP-111In or trastuzumab-Hy-MCP- 111In (14 MBq, 10mug).;Conclusion: These results are promising for further development of trastuzumab-Hy-PEG-MCP-111In for Auger electron radioimmunotherapy of BC. |
