Photoradiolabeling of onartuzumab with 99mTc and 188Re-tricarbonyl for radiotheranostics of gastric cancer

Abstract

The clinically relevant nuclear isomer of technetium-99 (99mTc) and the radionuclides rhenium-186/188 (186Re and 188Re) represent an almost ideal match for the development of radiotracers for applications in diagnostic imaging and molecularly targeted radionuclide therapy. Although the chemistry of Tc and Re are similar, important differences arise in the synthesis and properties of their complexes. Here, we report the synthesis and characterization of 99mTc- and 188Re-onartuzumab by photoradiolabeling of the cancer-specific mAb onartuzumab (MetMAb) with the corresponding metal-tricarbonyl complexes derived from a novel photoactivatable ligand. The acyclic tris-amine ligand L1, featuring a photoactive aryl-azide (ArN3) group, was synthesized from N1-(2-aminoethyl)ethane-1,2-diamine in 5 steps with an overall yield of 32%. Radiosynthesis of the [M(CO)3L1]+ (M = 99mTc or 188Re) photoactivatable complexes was accomplished via reduction of the [MVIIO4]- species to give the intermediate 99mTcI- and 188ReI-tricarbonyl-triaquo followed by ligand substitution with L1. The light-induced photoradiosynthesis of [M(CO)3L1-azepin]-onartuzumab (M-onartuzumab; M = 99mTc or 188Re) was achieved by irradiating the [M(CO)3L1]+ complexes in the presence of onartuzumab (formulated as MetMAb), with 395 nm light for 15 minutes at room temperature. Photoradiolabelling reactions produced M-onartuzumab radioimmunoconjugates in decay-corrected radiochemical yields of 20-to-30%, high radiochemical purities (RCP >95%), and in molar activities of 1.026-to-4.146 MBq nmol-1. Cellular binding assays confirmed the specificity of radiotracer binding toward human hepatocyte ‘growth factor’ receptor (c-MET) expression on the surface of MNK-45 gastric adenocarcinoma cells. Subsequent planar γ-ray scintigraphy imaging and ex vivobiodistribution experiments in mouse models bearing subcutaneous MKN-45 xenografts revealed specific tumor targeting compared against competitive inhibition (blocking) controls performed at 24 hours (99mTc and 188Re) and 72 hours (188Re). Tumor uptake reached 20.20 ± 4.05 %ID g-1 for 99mTc-onartuzumab and 22.13 ± 3.11 %ID g-1 for 188Re-onartuzumab after 24 hours. Blocking experiments confirmed tumor specificity, with a reduction in tumor uptake of approximately 70% for both 99mTc-onartuzumab and 188Re-onartuzumab. Experimental data also revealed the biochemical equvalence of 99mTc-onartuzumab and 188Re-onartuzumab in terms of stability and pharmacokinetics in vivo. For 188Re-onartuzumab, activity was retained in the tumor for over 72 hours, with uptake levels at 20.21 ± 1.47 %ID g-1. Overall, the experiments demonstrate that photoradiosynthesis can be employed to develop a variatiy of  rhenium based radioimmunoconjugates for future applications in tumor targeted radioimmunotherapy. Furthermore, these results underline the high potential of rhenium and technetium radioconjugates as theranostic paltforms.