Yoichi Shimizu1,2,3, Songji Zhao4, Hironobu Yasui5,4, Ken-Ichi Nishijima5, Hiroki Matsumoto6, Tohru Shiga4, Nagara Tamaki4, Mikako Ogawa7, Yuji Kuge5,4. 1. Laboratory of Bioanalysis and Molecular Imaging, Graduate School of Pharmaceutical Sciences, Hokkaido University, Kita 12, Nishi 6, Kita-ku, Sapporo, 060-0812, Japan. yshimizu@pharm.hokudai.ac.jp. 2. Central Institute of Isotope Science, Hokkaido University, Kita 15 Nishi 7, Kita-ku, Sapporo, 060-0815, Japan. yshimizu@pharm.hokudai.ac.jp. 3. Kyoto University Hospital, 54 Shogoinkawahara-cho, Sakyo-ku, Kyoto, 606-8507, Japan. yshimizu@pharm.hokudai.ac.jp. 4. Graduate School of Medicine, Hokkaido University, Kita 15 Nishi 7, Kita-ku, Sapporo, 060-8638, Japan. 5. Central Institute of Isotope Science, Hokkaido University, Kita 15 Nishi 7, Kita-ku, Sapporo, 060-0815, Japan. 6. Research Centre, Nihon Medi-Physics Co., Ltd., 3-1 Kitatode, Sodegaura, Chiba, 299-0266, Japan. 7. Laboratory of Bioanalysis and Molecular Imaging, Graduate School of Pharmaceutical Sciences, Hokkaido University, Kita 12, Nishi 6, Kita-ku, Sapporo, 060-0812, Japan.
Abstract
PURPOSE: Hypoxia in tumor has close relationship with angiogenesis and tumor progression. Previously, we developed 2,2-dihydroxymethyl-3-[18F]fluoropropyl-2-nitroimidazole ([18F]DiFA) as a novel positron emission tomography (PET) probe for diagnosis of hypoxia. In this study, we elucidated whether the accumulation of [18F]DiFA in cells is dependent on the hypoxic state and revealed how [18F]DiFA accumulates in hypoxic cells in combination with imaging mass spectrometry (IMS). PROCEDURES: FaDu human head and neck cancer cells were treated with [18F]DiFA and then incubated under normoxia (21% O2) or hypoxia (1% O2) for 2 h. The cells were extracted using methanol, and the radioactivities of the precipitates (macromolecule fraction) and supernatants (low-molecular-weight fraction) were measured. FaDu-bearing mice were injected intravenously with [18F]DiFA and with pimonidazole 1 h later. The tumors were excised 2 h after the injection of [18F]DiFA. Autoradiography, IMS, and immunohistochemical (IHC) staining for pimonidazole were performed with serial tumor sections. RESULTS: In the in vitro study, the radioactivity of FaDu cells was significantly higher under hypoxia than that under normoxia (0.53 ± 0.02 vs. 0.27 ± 0.02 %dose/mg protein, p < 0.05). The radioactivity of the low-molecular-weight fraction was 66.3 ± 0.6% in the hypoxic cell. In the in vivo study, [18F]DiFA accumulated in the tumor tissues existed mainly as low-molecular-weight compounds (90.4 ± 0.9%). In addition, the glutathione conjugate of reductive DiFA metabolite (amino-DiFA-GS) existed in tumor tissues revealed by the IMS study, and the distribution pattern of amino-DiFA-GS was very similar to that of the radioactivity and the positive staining area of pimonidazole. CONCLUSIONS: Our results suggest that [18F]DiFA undergoes the glutathione conjugation reaction following reductive metabolism in hypoxic cells, which leads hypoxia-specific PET imaging with [18F]DiFA.
PURPOSE:Hypoxia in tumor has close relationship with angiogenesis and tumor progression. Previously, we developed 2,2-dihydroxymethyl-3-[18F]fluoropropyl-2-nitroimidazole ([18F]DiFA) as a novel positron emission tomography (PET) probe for diagnosis of hypoxia. In this study, we elucidated whether the accumulation of [18F]DiFA in cells is dependent on the hypoxic state and revealed how [18F]DiFA accumulates in hypoxic cells in combination with imaging mass spectrometry (IMS). PROCEDURES: FaDu human head and neck cancer cells were treated with [18F]DiFA and then incubated under normoxia (21% O2) or hypoxia (1% O2) for 2 h. The cells were extracted using methanol, and the radioactivities of the precipitates (macromolecule fraction) and supernatants (low-molecular-weight fraction) were measured. FaDu-bearing mice were injected intravenously with [18F]DiFA and with pimonidazole 1 h later. The tumors were excised 2 h after the injection of [18F]DiFA. Autoradiography, IMS, and immunohistochemical (IHC) staining for pimonidazole were performed with serial tumor sections. RESULTS: In the in vitro study, the radioactivity of FaDu cells was significantly higher under hypoxia than that under normoxia (0.53 ± 0.02 vs. 0.27 ± 0.02 %dose/mg protein, p < 0.05). The radioactivity of the low-molecular-weight fraction was 66.3 ± 0.6% in the hypoxic cell. In the in vivo study, [18F]DiFA accumulated in the tumor tissues existed mainly as low-molecular-weight compounds (90.4 ± 0.9%). In addition, the glutathione conjugate of reductive DiFA metabolite (amino-DiFA-GS) existed in tumor tissues revealed by the IMS study, and the distribution pattern of amino-DiFA-GS was very similar to that of the radioactivity and the positive staining area of pimonidazole. CONCLUSIONS: Our results suggest that [18F]DiFA undergoes the glutathione conjugation reaction following reductive metabolism in hypoxic cells, which leads hypoxia-specific PET imaging with [18F]DiFA.
Entities:
Keywords:
DiFA; Glutathione; Hypoxia; Imaging mass spectrometry; Molecular imaging
Authors: Jason Gobey; Mark Cole; John Janiszewski; Thomas Covey; Tung Chau; Peter Kovarik; Jay Corr Journal: Anal Chem Date: 2005-09-01 Impact factor: 6.986