MET is a proto-oncogene protein, also known as c-Met or hepatocyte growth factor (HGF) receptor (HGFR) (1). MET is a membrane receptor with tyrosine kinase activity that is essential for embryonic development and wound healing. MET is normally expressed by stem cells, progenitor cells, and cells of epithelial origin, whereas expression of HGF, which is the only known ligand of MET, is restricted to cells of mesenchymal origin. Upon HGF stimulation, MET induces several biological responses that lead to invasive growth (2). Abnormal MET activation in cancer correlates with poor prognosis because deregulated MET triggers tumor growth, induces angiogenesis, and activates metastasis (3). MET and HGF are overexpressed in many types of human malignancies, including kidney, liver, stomach, breast, and brain cancers (2, 4, 5). Inhibition of MET function has been shown to inhibit pathological angiogenesis as well as tumor growth and metastasis (6). On the other hand, targeting HGF instead of the HGF receptor may also be beneficial to patients by blocking binding of HGF to MET.
Monoclonal antibody DN30 (150 kDa) is directed against the extracellular domain of MET, with a Kd of 2.64 nM (6). Inhibition of HGFR function with DN30 has been shown to inhibit pathological angiogenesis as well as tumor growth and metastasis. Perk et al. (7) prepared 89Zr-N-succinyldesferrioxamine-DN30 (89Zr-DN30) for imaging MET expression in tumors, showing tumor accumulation corresponding to MET expression levels. However, DN30 is a partial agonist with partial biological responses. Onartuzumab is a MET-selective humanized 1-armed monoclonal antibody (100 kDa) with anti-proliferation, anti-angiogenic, and pro-apoptotic properties against tumor growth in mouse tumor models (8). Onartuzumab is a pure antagonist with monovalent binding to MET. For use with positron emission tomography (PET), onartuzumab was conjugated with the bifunctional chelate p-isothiocyanatobenzyl-desferrioxamine (Df-Bz-NCS) and labeled with 89Zr to form 89Zr-Df-onartuzumab for imaging MET expression in tumors (9).
Df-Onartuzumab was prepared by incubation of onartuzumab with Df-Bz-NCS in buffer (pH 9) for 30 min at 37°C (9). Df-Onartuzumab was purified with column chromatography. The number of Df groups per onartuzumab was not reported. For 89Zr-labeling, a solution of 74–148 MBq (2–4 mCi) 89Zr in 1 M oxalic acid and Df-onartuzumab was incubated in HEPES buffer (pH 7.0) for 60 min at room temperature. The labeling yields were >90%. The specific activities of 89Zr-Df-onartuzumab were 37–185 MBq/nmol (1–5 mCi/nmol) at the end of synthesis.
In Vitro Studies: Testing in Cells and Tissues
In cell-based saturation assays with human gastric cancer MKN-45 cells, 89Zr-Df-onartuzumab exhibited an affinity constant (Kd) value of 6.12 ± 1.18 nM (n = 3) and a Bmax value of 0.94 ± 0.08 × 106 sites/cell (n = 3) (9). The immunoreactivity of 89Zr-Df-onartuzumab was 81.9 ± 0.7% (n = 3).
Jagoda et al. (9) performed ex vivo biodistribution studies of 1.85 MBq (0.05 mCi) 89Zr-Df-onartuzumab (~0.5 nmol) in nude mice bearing MKN-45 tumors (high MET expression) or U87MG tumors (low MET expression) (n = 4–5 mice/group) at 1, 2, 3, 4, and 5 d after injection. Accumulation in MKN-45 tumors was 10% injected dose/gram (ID/g) at 1 d and gradually increased to 23% ID/g at 5 d. Accumulation in the non-targeted tissues (blood, heart, lung, gastrointestinal tract, and muscle) decreased slowly, with ~50% reduction from 1 d to 5 d. On the other hand, radioactivity in the liver and kidney increased with time, suggesting hepatobiliary and renal clearance. Accumulation in the tumors was higher than in all other tissues at 2–5 d. Accumulation in U87MG tumors was 1.8- and 3.0-fold lower than that in the MKN-45 tumors at 3 d and 5 d, respectively. The tumor/muscle ratios for MKN-45 tumors were 13.2, 26.5, and 26.6 at 2, 3, and 5 d, respectively. On the other hand, the tumor/muscle ratios for U87MG tumors were 7 and 9 at 2 d and 5 d, respectively. Co-injection of excess onartuzumab (10 nmol/mouse) was performed with mice bearing MKN-45 tumors (n = 4 mice) with 0.37 MBq (0.01 mCi) 89Zr-Df-onartuzumab (~0.03 nmol) at 3 d after injection. Tumor accumulation was reduced by 67%.
Whole-body PET imaging studies were performed in mice bearing MKN-45 xenografts at 1–5 d after injection of 7.4 MBq (0.2 mCi) 89Zr-Df-onartuzumab (1.2 nmol) (9). The tumors could be visualized at 1 d with improving quality over the 5-d period. The tumor/muscle ratio was 26 at 5 d.
Other Non-Primate Mammals
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Intramural Research Program
- Boccaccio C., Comoglio P.M. Invasive growth: a MET-driven genetic programme for cancer and stem cells. Nat Rev Cancer. 2006;6(8):637–45. [PubMed: 16862193]
- Di Renzo M.F., Olivero M., Giacomini A., Porte H., Chastre E., Mirossay L., Nordlinger B., Bretti S., Bottardi S., Giordano S. et al. Overexpression and amplification of the met/HGF receptor gene during the progression of colorectal cancer. Clin Cancer Res. 1995;1(2):147–54. [PubMed: 9815967]
- Lengyel E., Prechtel D., Resau J.H., Gauger K., Welk A., Lindemann K., Salanti G., Richter T., Knudsen B., Vande Woude G.F., Harbeck N. C-Met overexpression in node-positive breast cancer identifies patients with poor clinical outcome independent of Her2/neu. Int J Cancer. 2005;113(4):678–82. [PubMed: 15455388]
- Smith H.S., Stern R., Liu E., Benz C. Early and late events in the development of human breast cancer. Basic Life Sci. 1991;57:329–37. [PubMed: 1814293]
- Di Renzo M.F., Olivero M., Serini G., Orlandi F., Pilotti S., Belfiore A., Costantino A., Vigneri R., Angeli A., Pierotti M.A. et al. Overexpression of the c-MET/HGF receptor in human thyroid carcinomas derived from the follicular epithelium. J Endocrinol Invest. 1995;18(2):134–9. [PubMed: 7629381]
- Petrelli A., Circosta P., Granziero L., Mazzone M., Pisacane A., Fenoglio S., Comoglio P.M., Giordano S. Ab-induced ectodomain shedding mediates hepatocyte growth factor receptor down-regulation and hampers biological activity. Proc Natl Acad Sci U S A. 2006;103(13):5090–5. [PMC free article: PMC1458799] [PubMed: 16547140]
- Perk L.R., Stigter-van Walsum M., Visser G.W., Kloet R.W., Vosjan M.J., Leemans C.R., Giaccone G., Albano R., Comoglio P.M., van Dongen G.A. Quantitative PET imaging of Met-expressing human cancer xenografts with (89)Zr-labelled monoclonal antibody DN30. Eur J Nucl Med Mol Imaging. 2008;35(10):1857–67. [PubMed: 18491091]
- Martens T., Schmidt N.O., Eckerich C., Fillbrandt R., Merchant M., Schwall R., Westphal M., Lamszus K. A novel one-armed anti-c-Met antibody inhibits glioblastoma growth in vivo. Clin Cancer Res. 2006;12(20 Pt 1):6144–52. [PubMed: 17062691]
- Jagoda E.M., Lang L., Bhadrasetty V., Histed S., Williams M., Kramer-Marek G., Mena E., Rosenblum L., Marik J., Tinianow J.N., Merchant M., Szajek L., Paik C., Cecchi F., Raffensperger K., Jose-Dizon J.M., Bottaro D.P., Choyke P. Immuno-PET of the Hepatocyte Growth Factor Receptor Met Using the 1-Armed Antibody Onartuzumab. J Nucl Med. 2012;53(10):1592–600. [PMC free article: PMC3982858] [PubMed: 22917884]
Created: December 1, 2012; Last Update: March 14, 2013.
National Center for Biotechnology Information (US), Bethesda (MD)
Leung K. 89Zr-Desferrioxamine p-isothiocyanatobenzyl-anti-hepatocyte growth factor receptor 1-armed antibody onartuzumab. 2012 Dec 1 [Updated 2013 Mar 14]. In: Molecular Imaging and Contrast Agent Database (MICAD) [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2004-2013.