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Background In pancreatic cancer surgery, accurate identification and resection of intrapancreatic residual tumors are quite difficult. We have developed a novel open-typed PET system (called 'OpenPET'), which enables high-resolution PET-guided surgery in real time, and demonstrated that OpenPET-guided surgery with intraperitoneally administered 64Cu-labeled anti-epidermal growth factor receptor antibody cetuximab is useful to detect and resect primary pancreatic cancer. Here, we investigated applicability of OpenPET-guided surgery for unexpected residual intrapancreatic tumors and examined its survival benefit over conventional surgery. Methods A mouse model with large (>1 cm) resectable pancreatic cancer of xPA-1-DC cells expressing red fluorescent protein was used. OpenPET-guided surgery was conducted 24 h after intraperitoneal administration of 64Cu-labeled cetuximab (7.4 MBq/mouse). For comparison, similar surgical procedures were conducted, and conventional tumor resection was attempted using only the naked eye (control). Survival rate after OpenPET-guided surgery was compared to that after control operations. Results Intraoperative OpenPET guidance enabled detection and resection of small residual tumors. Ten residual tumor specimens (3-10 mm in diameter) were intraoperatively isolated with OpenPET guidance (n = 7 mice). All isolated specimens showed tumor RFP signals. No resection of tumor tissue was performed in control group because the tumor could not be clearly detected with the naked eye alone. Mice after OpenPET-guided surgery showed significantly longer survival rates than those in control group. Conclusions OpenPET-guided surgery with 64Cu-labeled-cetuximab enabled intraoperative identification and resection of intrapancreatic small residual tumors. This technology could be useful to prevent tumor residuals during surgery and improve pancreatic cancer survival.

Peritoneal dissemination of pancreatic cancer has a poor prognosis. We have reported that intraperitoneal radioimmunotherapy using a 64Cu-labeled antibody (64Cu-ipRIT) is a promising adjuvant therapy option to prevent this complication. To achieve personalized 64Cu-ipRIT, we developed a new in vitro tumor cell-binding assay (64Cu-TuBA) system with a panel containing nine candidate 64Cu-labeled antibodies targeting seven antigens (EGFR, HER2, HER3, TfR, EpCAM, LAT1, and CD98), which are reportedly overexpressed in patients with pancreatic cancer. We investigated the feasibility of 64Cu-TuBA to select the highest-binding antibody for individual cancer cell lines and predict the treatment response in vivo for 64Cu-ipRIT. 64Cu-TuBA was performed using six human pancreatic cancer cell lines. For three cell lines, an in vivo treatment study was performed with 64Cu-ipRIT using high-, middle-, or low-binding antibodies in each peritoneal dissemination mouse model. The high-binding antibodies significantly prolonged survival in each mouse model, while low-and middle-binding antibodies were ineffective. There was a correlation between in vitro cell binding and in vivo therapeutic efficacy. Our findings suggest that 64Cu-TuBA can be used for patient selection to enable personalized 64Cu-ipRIT. Tumor cells isolated from surgically resected tumor tissues would be suitable for analysis with the 64Cu-TuBA system in future clinical studies.

Actinium-225 (225Ac) is a promising radionuclide used in targeted alpha therapy (TAT). Although 225Ac labeling of bifunctional chelating ligands is effective, previous in vivo studies reported that free 225Ac can be released from the drugs and that such free 225Ac is predominantly accumulated in the liver and could cause unexpected toxicity. To accelerate the clinical development of 225Ac TAT with a variety of drugs, preparing methods to deal with any unexpected toxicity would be valuable. The aim of this study was to evaluate the feasibility of various chelators for reducing and excreting free 225Ac and compare their chemical structures. Nine candidate chelators (D-penicillamine, dimercaprol, Ca-DTPA, Ca-EDTA, CyDTA, GEDTA TTHA, Ca-TTHA, and DO3A) were evaluated in vitro and in vivo. The biodistribution and dosimetry of free 225Ac were examined in mice before an in vivo chelating study. The liver exhibited pronounced 225Ac uptake, with an estimated human absorbed dose of 4.76 SvRBE5/MBq. Aminopolycarboxylate chelators with five and six carboxylic groups, Ca-DTPA and Ca-TTHA, significantly reduced 225Ac retention in the liver (22% and 30%, respectively). Significant 225Ac reductions were observed in the heart and remainder of the body with both Ca-DTPA and Ca-TTHA, and in the lung, kidney, and spleen with Ca-TTHA. In vitro interaction analysis supported the in vivo reduction ability of Ca-DTPA and Ca-TTHA. In conclusion, aminopolycarboxylate chelators with five and six carboxylic groups, Ca-DTPA and Ca-TTHA, were effective for whole-body clearance of free 225Ac. This feasibility study provides useful information for reducing undesirable radiation exposure from free 225Ac.

Pancreatic cancer (PC) has a poor prognosis owing to difficulties in the diagnosis of resectable PC at early stages. Several clinical studies have indicated that the detection and surgery of small resectable PC (64Cu-labeled anti-epidermal growth factor receptor antibody cetuximab was intraperitoneally administered into mice. It clearly identified PC tumors ≥3 mm. In contrast, neither OpenPET with intravenous-administered 64Cu-cetuximab nor intraperitoneal/intravenous-administered 18F-FDG (a traditional PET probe) could detect PC in this model. Immuno-OpenPET-guided surgery accurately resected small PC in mice and achieved significantly prolonged survival. This technology could provide a novel diagnostic and therapeutic strategy for small resectable PC to improve patient survival.

Purpose 211At, a promising alpha emitter, can easily volatilize and contaminate the environment. To safely manage this unique alpha emitter, we investigated the permeability of four types of plastic films and gloves against 211At and identified suitable materials to avoid contamination by 211At. Methods Four types of plastic films, polyethylene, polyvinylidene chloride, polyvinyl chloride, and a laminated film, and two types of rubber gloves, latex and nitrile, were examined. Small pieces of filter paper were covered with these materials, and a drop containing 100 kBq of 211At was placed on them. The radioactivity of pieces of filter paper under the materials was evaluated by measuring counts using a gamma counter and obtaining autoradiograms 3.5 h later. These experiments were also performed using 225Ac, 125I, 111In, 201Tl, and 99mTc. Results 211At solution easily penetrated polyethylene, polyvinyl chloride, and latex rubber. Similar results were obtained for 125I, while other radionuclides did not penetrate films or gloves. These results suggest that halogenic radionuclides under anionic conditions are likely to penetrate plastic films and rubber gloves. Conclusion Our evaluation revealed that, when 211At solution is used, the protection by polyvinylidene chloride, a laminated film, or nitrile rubber would be more effective than that by polyethylene, polyvinyl chloride, or latex rubber.

Four types of plastic films, polyethylene, polyvinylidene chloride, polyvinyl chloride, and a laminated film, and two types of rubber gloves, latex and nitrile, were examined. Small pieces of filter paper were covered with these materials, and a drop containing 100 kBq ofOur evaluation revealed that, when

Peritoneal dissemination is a common cause of death from gastrointestinal cancers and is difficult to treat using current therapeutic options, particularly late-phase disease. Here, we investigated the feasibility of integrated therapy using 64Cu-intraperitoneal radioimmunotherapy (ipRIT), alone or in combination with positron emission tomography (PET)-guided surgery using a theranostic agent (64Cu-labeled anti-epidermal growth factor receptor antibody cetuximab) to treat early- and late-phase peritoneal dissemination in mouse models. In this study, we utilized the OpenPET system, which has open space for conducting surgery while monitoring objects at high resolution in real time, as a novel approach to make PET-guided surgery feasible. 64Cu-ipRIT with cetuximab inhibited tumor growth and prolonged survival with little toxicity in mice with early-phase peritoneal dissemination of small lesions. For late-phase peritoneal dissemination, a combination of 64Cu-ipRIT for down-staging and subsequent OpenPET-guided surgery for resecting large tumor masses effectively prolonged survival. OpenPET clearly detected tumors (≥3 mm in size) behind other organs in the peritoneal cavity and was useful for confirming the presence or absence of residual tumors during an operation. These findings suggest that integrated 64Cu therapy can serve as a novel treatment strategy for peritoneal dissemination.

In boron neutron capture therapy (BNCT), 10 B-4-borono-L-phenylalanine (BPA) is commonly used as a 10 B carrier. PET using 4-borono-2-18 F-fluoro-phenylalanine (18 F-FBPA PET) has been performed to estimate boron concentration and predict the therapeutic effects of BNCT; however, the association between tumor uptake of 18 F-FBPA and boron concentration in tumors remains unclear. The present study investigated the transport mechanism of 18 F-FBPA and BPA, and evaluated the utility of 18 F-FBPA PET in predicting boron concentration in tumors. The transporter assay revealed that 2-aminobicyclo-(2.2.1)-heptane-2-carboxylic acid, an inhibitor of the L-type amino acid transporter, significantly inhibited 18 F-FBPA and 14 C-4-borono-L-phenylalanine (14 C-BPA) uptake in FaDu and LN-229 human cancer cells. 18 F-FBPA uptake strongly correlated with 14 C-BPA uptake in 7 human tumor cell lines (r = .93; P < .01). PET experiments demonstrated that tumor uptake of 18 F-FBPA was independent of the administration method, and uptake of 18 F-FBPA by bolus injection correlated well with BPA uptake by continuous intravenous infusion. The results of this study revealed that evaluating tumor uptake of 18 F-FBPA by PET was useful for estimating 10 B concentration in tumors.

Glioblastoma is the most aggressive malignant brain tumor in humans and is difficult to cure using current treatment options. Hypoxic regions are frequently found in glioblastoma, and increased levels of hypoxia are associated with poor clinical outcomes of glioblastoma patients. Hypoxia plays important roles in the progression and recurrence of glioblastoma because of drug delivery deficiencies and induction of hypoxia-inducible factor-1α in tumor cells, which lead to poor prognosis. We focused on a promising hypoxia-targeted internal radiotherapy agent, 64Cu-diacetyl-bis (N4-methylthiosemicarbazone) (64Cu-ATSM), to address the need for additional treatment for glioblastoma. This compound can target the overreduced state under hypoxic conditions within tumors. Clinical positron emission tomography studies using radiolabeled Cu-ATSM have shown that Cu-ATSM accumulates in glioblastoma and its uptake is associated with high hypoxia-inducible factor-1α expression. To evaluate the therapeutic potential of this agent for glioblastoma, we examined the efficacy of 64Cu-ATSM in mice bearing U87MG glioblastoma tumors. Administration of single dosage (18.5, 37, 74, 111, and 148 MBq) and multiple dosages (37 MBq × 4) of 64Cu-ATSM was investigated. Single administration of 64Cu-ATSM in high-dose groups dose-dependently inhibited tumor growth and prolonged survival, with slight and reverse signs of adverse events. Multiple dosages of 64Cu-ATSM remarkably inhibited tumor growth and prolonged survival. By splitting the dose of 64Cu-ATSM, no adverse effects were observed. Our findings indicate that multiple administrations of 64Cu-ATSM have effective antitumor effects in glioblastoma without side effects, indicating its potential for treating this fatal disease.

// Yukie Yoshii 1 , Mitsuyoshi Yoshimoto 2 , Hiroki Matsumoto 3 , Takako Furukawa 1, 4 , Ming-Rong Zhang 1 , Masayuki Inubushi 5 , Atsushi B. Tsuji 1 , Yasuhisa Fujibayashi 1 , Tatsuya Higashi 1 and Tsuneo Saga 1, 6 1 National Institute of Radiological Sciences, National Institutes for Quantum and Radiological Science and Technology, Chiba, Japan 2 Division of Functional Imaging, National Cancer Center Hospital East, Kashiwa, Japan 3 Research Centre, Nihon Medi-Physics Co., Ltd., Sodegaura, Japan 4 Department of Radiological and Medical Laboratory Sciences, Nagoya University Graduate School of Medicine, Nagoya, Japan 5 Department of Nuclear Medicine, Kawasaki Medical School, Kurashiki, Japan 6 Department of Diagnostic Radiology, Kyoto University Hospital, Kyoto, Japan Correspondence to: Yukie Yoshii, email: yoshii.yukie@qst.go.jp Keywords: 64 Cu-ATSM, angiogenesis, bevacizumab, hypoxia, vascular decrease Received: July 19, 2017 Accepted: August 17, 2017 Published: September 28, 2017 ABSTRACT Bevacizumab, an anti-vascular endothelial growth factor (VEGF) antibody, is an antiangiogenic agent clinically used for various cancers. However, repeated use of this agent leads to tumor-decreased vascularity and hypoxia with activation of an HIF-1 signaling pathway, which results in drug delivery deficiency and induction of malignant behaviors in tumors. Here, we developed a novel strategy to treat tumors with bevacizumab-induced vascular decrease and hypoxia using 64 Cu-diacetyl-bis ( N 4 -methylthiosemicarbazone) ( 64 Cu-ATSM), a potential theranostic agent, which possesses high tissue permeability and can target over-reduced conditions under hypoxia in tumors, with a human colon carcinoma HT-29 tumor-bearing mouse model. The long-term treatment with bevacizumab caused decreased blood vessel density and activation of an HIF-1 signaling pathway; increased uptake of 64 Cu-ATSM was also observed despite limited blood vessel density in HT-29 tumors. In vivo high-resolution SPECT/PET/CT imaging confirmed reduced vascularity and increased proportion of 64 Cu-ATSM uptake areas within the bevacizumab-treated tumors. 64 Cu-ATSM therapy was effective to inhibit tumor growth and prolong survival of the bevacizumab-treated tumor-bearing mice without major adverse effects. In conclusion, 64 Cu-ATSM therapy effectively enhanced anti-tumor effects in tumors with bevacizumab-induced vascular decrease and hypoxia. 64 Cu-ATSM therapy could represent a novel approach as an add-on to antiangiogenic therapy.

Introduction To clarify the difference between system A and L amino acid transport imaging in PET clinical imaging, we focused on the use of α-[ N -methyl- 11 C ]-methylaminoisobutyric acid ([ 11 C ]MeAIB), and compared it with [ S -methyl- 11 C ]-L-methionine ([ 11 C ]MET). The aim of this study was to assess the correlation of accumulation of these two radioactive amino acid analogs with expression of amino acid transporters and cell proliferative activity in carcinoma cells. Methods Amino acid uptake inhibitor studies were performed in four human carcinoma cells (epidermal carcinoma A431, colorectal carcinoma LS180, and lung carcinomas PC14/GL and H441/GL) using the radioisotope analogs [ 3 H]MET and [ 14 C]MeAIB. MeAIB was used to inhibit the A system and 2-amino-2-norbornane-carboxylic acid (BCH) was used to inhibit the L system. The carcinoma gene expression levels of a number of amino acid transporters were measured by microarray and quantitative polymerase chain reaction. Carcinoma proliferative activity was assessed using accumulation of [methyl- 3 H]-3'-deoxy-3'-fluorothymidine ([ 3 H]FLT). Results and conclusion [ 14 C]MeAIB uptake occurred principally via a Na + -dependent A type mechanism whereas [ 3 H]MET uptake occurred predominantly via a Na + -independent L type mechanism although other transporters were also utilized depending on cell type. There was no correlation between [ 3 H]MET uptake and total system L amino acid transporter (LAT) expression. In contrast, [ 14 C]MeAIB uptake strongly correlated with total system A amino acid transporter (SNAT) expression and proliferative activity in this preliminary study using four human carcinoma cell lines. Carcinoma proliferative activity also correlated with total SNAT expression. Advances in Knowledge and Implications for Patient Care: Because there is a significant correlation between the accumulation of [ 14 C]MeAIB and the gene expression level of total SNAT as well as the accumulation of [ 3 H]FLT, it is suggested that use of the analog [ 11 C]MeAIB in PET may provide an indication of tumor cell proliferative activity. [ 11 C]MeAIB is therefore expected to be very useful in PET imaging.

Hypoxia in tumors is known to contribute to the malignant characteristics such as therapy resistance and metastatic potential. Cu-diacetyl-bis (N4-methylthiosemicarbazone) (Cu-ATSM), labeled with several Cu radioisotopes (60Cu, 62Cu and 64Cu) is a potential PET imaging agent that targets tumor hypoxia with over-reduced conditions. Clinical studies of Cu-ATSM PET has been conducted worldwide in recent years and the usefulness of this agent is being recognized. Multicenter clinical study of 62Cu-ATSM PET has been conducted in Japan. Among Cu-ATSM tracers, 64Cu-ATSM can be used not only as a PET imaging agent but also as a therapeutic agent, because 64Cu emits β− particle and Auger electron, which can damage the cells. Here, we will discuss the present status and future prospects of Cu-ATSM.

Pancreatic cancer (PC) has a very poor prognosis. Surgery is the primary treatment for patients with resectable PC; however, local recurrence, hepatic metastasis, and peritoneal dissemination often occur even after extensive surgery. Adjuvant chemotherapy, typically with gemcitabine, has been used clinically but with only a modest survival benefit. To achieve a better outcome, we investigated the efficacy of (64)Cu-intraperitoneal radioimmunotherapy (ipRIT) with (64)Cu-labeled antiepidermal growth factor receptor antibody cetuximab as an adjuvant treatment after PC surgery using an orthotopic xenografted mouse model. Methods: The efficacy of adjuvant (64)Cu-ipRIT was investigated in a human PC mouse model harboring orthotopic xenografts of xPA-1-DC cells. To reproduce the clinical situation, PC xenografts were surgically resected when pancreatic tumors were readily visible but not metastatic tumors. Increasing doses of (64)Cu-cetuximab were intraperitoneally injected, and the mice were monitored for toxicity to determine the safe therapeutic dose. For adjuvant (64)Cu-ipRIT, the day after tumor resection, the mice were intraperitoneally administered 22.2 MBq of (64)Cu-PCTA-cetuximab and the survival was compared with that in surgery-only controls. For comparison, adjuvant chemotherapy with gemcitabine was also examined using the same model. Results: The mouse model not only developed primary tumors in the pancreas but also subsequently reproduced local recurrence, hepatic metastasis, and peritoneal dissemination after surgery, which is similar to the manifestations that occur with human PC. Adjuvant (64)Cu-ipRIT with (64)Cu-labeled cetuximab after surgery effectively suppressed local recurrence, hepatic metastasis, and peritoneal dissemination in this model. Significant improvement of the survival with minimal toxicity was achieved by adjuvant (64)Cu-ipRIT compared with that in control mice that underwent surgery only. Adjuvant chemotherapy with gemcitabine nominally prolonged the survival, but the effect was not statistically significant. Conclusion: (64)Cu-ipRIT with cetuximab can be an effective adjuvant therapy after PC surgery.

Monoamine oxidase B (MAO-B), predominantly expressed in glial cells, plays an important role in neurotransmitter regulation, and MAO-B activity relates to several neuronal diseases. Here, we aimed to develop a radiofluorinated MAO-B imaging probe based on the structure of a selective MAO-B inhibitor, MD-230254. We synthesized and evaluated a series of compounds in vitro and in vivo. A series of fluorinated analogs of MD-230254 were synthesized and evaluated for inhibitory potency and selectivity toward MAO-B. 5-[4-(2-[18 F]Fluorobenzyloxy)phenyl]-3-(2-cyanoethyl)-1,3,4-oxadiazol-2(3H)-one (2-[18 F]FBPO) was synthesized from a corresponding tributylstannyl precursor and [18 F]CH3 COOF. Biodistribution after intravenous injection of 2-[18 F]FBPO was evaluated in male ddY mice with or without pretreatment by inhibitors. Among the compounds synthesized and evaluated, 2-FBPO showed high inhibitory potency and selectivity toward MAO-B comparable with MD-230254. 2-[18 F]FBPO was successfully synthesized by an electrophilic reaction with a high radiochemical purity of more than 99%. 2-[18 F]FBPO was efficiently taken up by the brain and showed rapid blood clearance, which provided a brain/blood radioactivity ratio of 3.7 at 90 minutes postinjection. The brain radioactivity was significantly decreased by pretreatment with an MAO-B selective inhibitor. The great potential of 2-[18 F]FBPO as an MAO-B imaging probe, applicable to a variety of diseases, is indicated.

In boron neutron capture therapy (BNCT), 10B‐4‐borono‐L‐phenylalanine (BPA) is commonly used as a 10B carrier. PET using 4‐borono‐2‐18F‐fluoro‐phenylalanine (18F‐FBPA PET) has been performed to estimate boron concentration and predict the therapeutic effects of BNCT; however, the association between tumor uptake of 18F‐FBPA and boron concentration in tumors remains unclear. The present study investigated the transport mechanism of 18F‐FBPA and BPA, and evaluated the utility of 18F‐FBPA PET in predicting boron concentration in tumors. The transporter assay revealed that 2‐aminobicyclo‐(2.2.1)‐heptane‐2‐carboxylic acid, an inhibitor of the L‐type amino acid transporter, significantly inhibited 18F‐FBPA and 14C‐4‐borono‐L‐phenylalanine (14C‐BPA) uptake in FaDu and LN‐229 human cancer cells. 18F‐FBPA uptake strongly correlated with 14C‐BPA uptake in 7 human tumor cell lines (r = .93; P < .01). PET experiments demonstrated that tumor uptake of 18F‐FBPA was independent of the administration method, and uptake of 18F‐FBPA by bolus injection correlated well with BPA uptake by continuous intravenous infusion. The results of this study revealed that evaluating tumor uptake of 18F‐FBPA by PET was useful for estimating 10B concentration in tumors.

Glioblastoma is the most aggressive malignant brain tumor in humans and is difficult to cure using current treatment options. Hypoxic regions are frequently found in glioblastoma, and increased levels of hypoxia are associated with poor clinical outcomes of glioblastoma patients. Hypoxia plays important roles in the progression and recurrence of glioblastoma because of drug delivery deficiencies and induction of hypoxia-inducible factor-1α in tumor cells, which lead to poor prognosis. We focused on a promising hypoxia-targeted internal radiotherapy agent, 64Cu-diacetyl-bis (N4-methylthiosemicarbazone) (64Cu-ATSM), to address the need for additional treatment for glioblastoma. This compound can target the overreduced state under hypoxic conditions within tumors. Clinical positron emission tomography studies using radiolabeled Cu-ATSM have shown that Cu-ATSM accumulates in glioblastoma and its uptake is associated with high hypoxia-inducible factor-1α expression. To evaluate the therapeutic potential of this agent for glioblastoma, we examined the efficacy of 64Cu-ATSM in mice bearing U87MG glioblastoma tumors. Administration of single dosage (18.5, 37, 74, 111, and 148 MBq) and multiple dosages (37 MBq × 4) of 64Cu-ATSM was investigated. Single administration of 64Cu-ATSM in high-dose groups dose-dependently inhibited tumor growth and prolonged survival, with slight and reverse signs of adverse events. Multiple dosages of 64Cu-ATSM remarkably inhibited tumor growth and prolonged survival. By splitting the dose of 64Cu-ATSM, no adverse effects were observed. Our findings indicate that multiple administrations of 64Cu-ATSM have effective antitumor effects in glioblastoma without side effects, indicating its potential for treating this fatal disease.

Background and purpose The aim of this study was to compare the accumulation of 4-borono-2- 18 F-fluoro-phenylalanine ( 18 F-BPA) with that of 18 F-fluorodeoxyglucose ( 18 F-FDG) in head and neck cancers, and to assess the usefulness of 18 F-FDG PET for screening candidates for boron neutron capture therapy (BNCT). Material and methods Twenty patients with pathologically proven malignant tumors of the head and neck were recruited from March 2012 to January 2014. All patients underwent both whole-body 18 F-BPA PET/CT and 18 F-FDG PET/CT within 2weeks of each other. The uptakes of 18 F-BPA and 18 F-FDG at 1h after injection were evaluated using the maximum standardized uptake value (SUVmax). Results The accumulation of 18 F-FDG was significantly correlated with that of 18 F-BPA. The SUVmax of 18 F-FDG ⩾5.0 is considered to be suggestive of high 18 F-BPA accumulation. Conclusions 18 F-FDG PET might be an effective screening method performed prior to 18 F-BPA for selecting patients with head and neck cancer for treatment with BNCT.

Lung cancer is one of the leading causes of cancer-related deaths worldwide, including Japan. Although computed tomography (CT) can detect small lung lesions such as those appearing as ground glass opacity, it cannot differentiate between malignant and non-malignant lesions. Previously, we have shown that single photon emission computed tomography (SPECT) imaging using (111)In-1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid-cyclo-(Arg-Gly-Asp-D-Phe-Lys) (DOTA-c(RGDfK)), an imaging probe of αvβ3 integrin, is useful for the early detection of pancreatic cancer in a hamster pancreatic carcinogenesis model. In this study, we aimed to assess the usefulness of SPECT/CT with (111)In-DOTA-c(RGDfK) for the evaluation of the malignancy of lung cancer.Lung tumors were induced by a single intraperitoneal injection (250 mg/kg) of urethane in male A/J mice. Twenty-six weeks after the urethane treatment, SPECT was performed an hour after injection of (111)In-DOTA-c(RGDfK). Following this, the radioactivity ratios of tumor to normal lung tissue were measured by autoradiography (ARG) in the excised lung samples. We also examined the expression of αvβ3 integrin in mouse and human lung samples.Urethane treatment induced 5 hyperplasias, 41 adenomas and 12 adenocarcinomas in the lungs of 8 A/J mice. SPECT with (111)In-DOTA-c(RGDfK) could clearly visualize lung nodules, though we failed to detect small lung nodules like adenoma and hyperplasias (adenocarcinoma: 66.7%, adenoma: 33.6%, hyperplasia: 0.0%). ARG analysis revealed significant uptake of (111)In-DOTA-c(RGDfK) in all the lesions. Moreover, tumor to normal lung tissue ratios increased along with the progression of carcinogenesis. Histopathological examination using human lung tissue samples revealed clear up-regulation of αvβ3 integrin in well-differentiated adenocarcinoma (Noguchi type B and C) rather than atypical adenomatous hyperplasia.Although there are some limitations in evaluating the malignancy of small lung tumors using (111)In-DOTA-c(RGDfK), SPECT with (111)In-DOTA-c(RGDfK) might be a useful non-invasive imaging approach for evaluating the characteristics of lung tumors in mice, thus showing potential for use in humans.

To clarify the difference between system A and L amino acid transport imaging in PET clinical imaging, we focused on the use of α-[N-methyl-Amino acid uptake inhibitor studies were performed in four human carcinoma cells (epidermal carcinoma A431, colorectal carcinoma LS180, and lung carcinomas PC14/GL and H441/GL) using the radioisotope analogs [[

Early detection of pancreatic cancer is key to overcoming its poor prognosis. α(v)β(3)-integrin is often overexpressed in pancreatic tumor cells, whereas it is scarcely expressed in normal pancreatic cells. In this study, we investigated the usefulness of SPECT imaging with (111)In-1,4,7,10-tetraazacylododecane-N,N',N″,N'''-tetraacetic acid-cyclo-(Arg-Gly-Asp-d-Phe-Lys) [(111)In-DOTA-c(RGDfK)], an imaging probe of α(v)β(3)-integrin, for the early detection of pancreatic cancer in a hamster pancreatic carcinogenesis model.Hamsters were subcutaneously injected with the pancreatic duct carcinogen N-nitrosobis(2-oxopropyl)amine to induce pancreatic cancer. N-nitrosobis(2-oxopropyl)amine-treated hamsters underwent in vivo SPECT with (111)In-DOTA-c(RGDfK). After imaging, the tumor-to-normal pancreatic tissue radioactivity ratios in excised pancreatic samples were measured with autoradiography (ARG) and compared with the immunopathologic findings for α(v)β(3)-integrin. In a mouse model in which inflammation was induced with turpentine, the uptake of (111)In-DOTA-c(RGDfK) in inflammatory regions was evaluated with ARG and compared with that of (18)F-FDG.(111)In-DOTA-c(RGDfK) was clearly visualized in pancreatic cancer lesions as small as 3 mm in diameter. ARG analysis revealed high tumor-to-normal pancreatic tissue radioactivity ratios (4.6 ± 1.0 [mean ± SD] in adenocarcinoma and 3.3 ± 1.4 in atypical hyperplasia). The uptake of (111)In-DOTA-c(RGDfK) strongly correlated with α(v)β(3)-integrin expression. In the inflammatory model, inflammation-to-muscle ratios for (18)F-FDG and (111)In-DOTA-c(RGDfK) were 8.37 ± 4.37 and 1.98 ± 0.60, respectively. These results imply that (111)In-DOTA-c(RGDfK) has a lower rate of false-positive tumor detection than (18)F-FDG.Our findings suggest that SPECT with (111)In-DOTA-c(RGDfK) has great potential for the early and accurate detection of pancreatic cancer.

Epidermal growth factor receptor tyrosine kinase (EGFR-TK) represents an attractive target for tumor diagnosis agents. Previously, the radioiodinated 4-(3-iodoanilino)-6,7-diethoxyquinazoline ([(125)I]m-IPQ) has been reported to possess good characteristics as a tumor imaging agent; however, it was also found to have low in vivo stability. To improve the in vivo stability, m-IPQ derivatives, 4-(3-iodophenoxy)-6,7-diethoxyquinazoline (PHY) and 4-(3-iodobenzylamino)-6,7-diethoxyquinazoline (BAY) were designed and synthesized, and the biological studies of [(125)I]PHY and [(125)I]BAY were performed to evaluate these new ligands as in vivo tumor diagnosis agents.PHY and BAY were synthesized according to previous reports. The EGFR-TK inhibitory potency of these new compounds was measured and compared to other EGFR-TK inhibitors. Radiolabeled [(125)I]PHY and [(125)I]BAY were synthesized by an iododestannylation reaction. Biodistribution studies of these radioligands were conducted in normal mice and tumor-bearing mice. Furthermore, selectivity and binding characteristics of [(125)I]PHY were analyzed by in vitro blocking studies and a binding assay.The new derivatives were found to have high inhibitory potency against EGFR-TK (PHY: IC50 = 12.7 ± 7.2 nM, BAY: IC50 = 51.0 ± 8.9 nM). [(125)I]PHY and [(125)I]BAY were conveniently synthesized from tributylstannyl precursors. In in vivo biodistribution studies, [(125)I]PHY and [(125)I]BAY were observed to have lower uptake in the stomach, an indication of deiodination, than [(125)I]m-IPQ. Moreover, [(125)I]PHY showed high uptake and prolonged retention in tumors and low accumulation in blood and muscle tissue resulting in a good tumor-to-blood ratio (0.94-1.50) and tumor-to-muscle ratio (1.02-1.95). The EGFR-TK selectivity of [(125)I]PHY was confirmed by pretreatment experiments with specific EGFR-TK inhibitors.New radioiodinated quinazoline derivatives were synthesized, which were found to have improved in vivo stability. In particular, [(125)I]PHY showed higher tumor accumulation than the other ligands which was indicative of selective binding to EGFR-TK. These desirable characteristics for [(125)I]PHY suggest that the (123)I-labeled counterpart, [(123)I]PHY, could be a possible candidate for cancer diagnosis radiopharmaceutical.

Trans-1-amino-3-(18)F-fluorocyclobutanecarboxylic acid (anti-(18)F-FACBC) is an amino acid PET tracer that has shown promise for visualizing prostate cancer. Therefore, we aimed to clarify the anti-(18)F-FACBC transport mechanism in prostate cancer cells. We also studied the fate of anti-(18)F-FACBC after it is transported into cells.For convenience, because of their longer half-lives, (14)C compounds were used instead of (18)F-labeled tracers. Trans-1-amino-3-fluoro-1-(14)C-cyclobutanecarboxylic acid ((14)C-FACBC) uptake was examined in human prostate cancer DU145 cells with the following substrates of amino acid transporters: α-(methylamino) isobutyric acid (a system A-specific substrate) and 2-amino-2-norbornanecarboxylic acid (a system L-specific substrate). The messenger RNA expression of amino acid transporters in human prostate cancer specimens was analyzed by complementary DNA microarray and quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). Gene expression in DU145 cells was analyzed by qRT-PCR. We also examined the knockdown effect of the amino acid transporters system ASC transporter 2 (ASCT2) and sodium-coupled neutral amino acid transporter 2 (SNAT2) on (14)C-FACBC uptake. In addition, the possibility of (14)C-FACBC incorporation into proteins was examined.(14)C-FACBC uptake by DU145 cells was markedly decreased to approximately 20% in the absence of Na(+), compared with that in its presence, indicating that Na(+)-dependent transporters are mainly responsible for the uptake of this tracer. Moreover, 2-amino-2-norbornanecarboxylic acid inhibited the transport of (14)C-FACBC to the basal level in Na(+)-free buffer. In contrast, α-(methylamino) isobutyric acid did not inhibit (14)C-FACBC accumulation in DU145 cells. Human prostate tumor specimens and DU145 cells had similar messenger RNA expression patterns of amino acid transporter genes. Although SNAT2 and ASCT2 are 2 major amino acid transporters expressed in prostate tumor tissues and DU145 cells, ASCT2 knockdown using small interfering RNA was more effective in lowering (14)C-FACBC transport than SNAT2. Almost all intracellular (14)C-FACBC was recovered from the nonprotein fraction.ASCT2, which is a Na(+)-dependent amino acid transporter, and to a lesser extent Na(+)-independent transporters play a role in the uptake of (14)C-FACBC by DU145 cells. Among the Na(+)-independent transporters, system L transporters are also involved in the transport of (14)C-FACBC. Moreover, (14)C-FACBC is not incorporated into proteins in cells. These findings suggest a possible mechanism of anti-(18)F-FACBC PET for prostate cancer.

A radioiodinated analog of PD153035 (m-IPQ) was evaluated as a potential epidermal growth factor receptor tyrosine kinase (EGFR-TK) activity imaging ligand for SPECT. The 50% inhibition concentration (IC50) value of m-IPQ for EGFR-TK phosphorylation inhibition was evaluated and compared to various EGFR-TK inhibitors. [125I]m-IPQ was synthesized by iododestannylation reaction. Biodistribution study of [125I]m-IPQ was conducted in normal mice and tumor-bearing mice. The selectivity and binding characteristics (B max and K d) were analyzed. The quinazoline derivative m-IPQ was found to have high inhibitory potency (IC50: 50.5 ± 3.5 nM) and selectivity toward EGFR-TK. In vivo biodistribution studies of [125I]m-IPQ demonstrated its rapid clearance and low retention in normal tissue. On the other hand, high tumor uptake was observed. However, the increase in [125I]m-IPQ uptake in the stomach as a deiodination parameter was found. Thus, [125I]m-IPQ showed low in vivo stability. The selectivity toward EGFR-TK of m-IPQ was confirmed by the pretreatment experiment with EGFR-TK specific inhibitors, PD153035, Genistein. [125I]m-IPQ bound to single population of binding sites with high affinity and kinetic parameter. In addition, [125I]m-IPQ was bound to EGFR-TK according to the amount of EGFR-TK expression in the tumor. [125I]m-IPQ showed a relatively high tumor accumulation with selective EGFR-TK binding. Moreover, the tumor uptake of [125I]m-IPQ might be reflected in the amount of EGFR-TK expression in the tumor. These good characteristics of [125I]m-IPQ suggested that a 123I-labeled counterpart, [123I]m-IPQ, would have great potential for EGFR-TK imaging with SPECT. However, the in vivo stability of this compound needs to improve.

金沢大学新学術創成研究機構 / 金沢大学医薬保健研究域薬学系, It has been reported that sigma receptors are highly expressed in a variety of human tumors. In this study, we selected (+)-2-[4-(4-iodophenyl)piperidino] cyclohexanol [(+)-pIV] as a sigma receptor ligand and evaluated the potential of radioiodinated (+)-pIV for tumor imaging and therapy. (+)-[125/131I]pIV was prepared by an iododestannylation reaction under no-carrier-added conditions with radiochemical purity over 99% after HPLC purification. Biodistribution experiments were performed by the intravenous injection of (+)-[125I]pIV into mice bearing human prostate tumors (DU-145). Blocking studies were performed by intravenous injection of (+)-[125I]pIV mixed with an excess amount of unlabeled sigma ligand into DU-145 tumor-bearing mice. For therapeutic study, (+)-[131I]pIV was injected at a dose of 7.4 MBq followed by measurement of the tumor size. In biodistribution experiments, (+)-[125I]pIV showed high uptake and long residence in the tumor. High tumor to blood and muscle ratios were achieved because the radioactivity levels of blood and muscle were low. However, the accumulations of radioactivity in non-target tissues, such as liver and kidney, were high. The radioactivity in the non-target tissues slowly decreased over time. Co-injection of (+)-[125I]pIV with an excess amount of unlabeled sigma ligand resulted in a significant decrease in the tumor/blood ratio, indicating sigma receptor-mediated tumor uptake. In therapeutic study, tumor growth in mice treated with (+)-[131I]pIV was significantly inhibited compared to that of an untreated group. These results indicate that radioiodinated (+)-pIV has a high potential for sigma receptor imaging in tumor and radionuclide receptor therapy. (Cancer Sci 2009). © 2009 Japanese Cancer Association.

The alpha(v)beta(3) integrin plays a pivotal role in angiogenesis and tumor metastasis. Angiogenic blood vessels overexpress alpha(v)beta(3) integrin, as in tumor neovascularization, and alpha(v)beta(3) integrin expression in other microvascular beds and organs is limited. Therefore, alpha(v)beta(3) integrin is a suitable receptor for tumor-targeting imaging and therapy. Recently, tetrameric and dimeric RGD peptides have been developed to enhance specificity to alpha(v)beta(3) integrin. In comparison to the corresponding monomeric peptide, however, these peptides show high levels of accumulation in kidney and liver. The purpose of this study is to evaluate tumor-targeting properties and the therapeutic potential of 111In- and 90Y-labeled monomeric RGD peptides in BALB/c nude mice with SKOV-3 human ovarian carcinoma tumors. DOTA-c(RGDfK) was labeled with 111In or 90Y and purified by HPLC. A biodistribution study and scintigraphic images revealed the specific uptake to alpha(v)beta(3) integrin and the rapid clearance from normal tissues. These peptides were renally excreted. At 10 min after injection of tracers, 111In-DOTA-c(RGDfK) and 90Y-DOTA-c(RGDfK) showed high uptake in tumors (7.3 +/- 0.6% ID/g and 4.6 +/- 0.8% ID/g, respectively) and gradually decreased over time (2.3 +/- 0.4% ID/g and 1.5 +/- 0.5% ID/g at 24 hr, respectively). High tumor-to-blood and -muscle ratios were obtained from these peptides. In radionuclide therapeutic study, multiple-dose administration of 90Y-DOTA-c(RGDfK) (3 x 11.1 MBq) suppressed tumor growth in comparison to the control group and a single-dose administration (11.1 MBq). Monomeric RGD peptides, 111In-DOTA-c(RGDfK) and (90)Y-DOTA-c(RGDfK), could be promising tracers for alpha(v)beta(3) integrin-targeting imaging and radiotherapy.

(64)Cu-diacetyl-bis (N(4)-methylthiosemicarbazone) ((64)Cu-ATSM) is a potential theranostic agent targeting the over-reduced state under hypoxia within tumors. Recent clinical Cu-ATSM positron emission tomography studies have revealed a correlation between uptake and poor prognosis; however, the reason is unclear. Here, using a human colon carcinoma HT-29 model, we demonstrated that the intratumoral (64)Cu-ATSM high-uptake regions exhibited malignant characteristics, such as upregulated DNA repair and elevated %CD133(+) cancer stem-like cells. Based on this evidence, we developed a strategy to enhance the efficacy of (64)Cu-ATSM internal radiotherapy (IRT) by inhibiting DNA repair with a nucleic acid (NA) antimetabolite. The results of the analyses showed upregulation of pathways related to DNA repair along with NA incorporation (bromodeoxyuridine uptake) and elevation of %CD133(+) cells in (64)Cu-ATSM high-uptake regions. In an in vivo(64)Cu-ATSM treatment study, co-administration of an NA antimetabolite and (64)Cu-ATSM synergistically inhibited tumor growth, with little toxicity, and effectively reduced %CD133(+) cells. (64)Cu-ATSM therapy targeted malignant tumor regions with activated DNA repair and high concentrations of CD133(+) cells in the HT-29 model. NA antimetabolite co-administration can be an effective approach to enhance the therapeutic effect of (64)Cu-ATSM IRT.

Introduction Angiogenesis is essential for tumor growth or metastasis. A method involving noninvasive detection of angiogenic activity in vivo would provide diagnostic information regarding antiangiogenic therapy targeting vascular endothelial cells as well as important insight into the role of vascular endothelial growth factor (VEGF) and its receptor (flt-1 and KDR) system in tumor biology. We evaluated radioiodinated VEGF 121 , which displays high binding affinity for KDR, and VEGF 165 , which possesses high binding affinity for flt-1 and low affinity for KDR, as angiogenesis imaging agents using the LS180 tumor xenograft model. Methods VEGF 121 and VEGF 165 were labeled with 125 I by the chloramine-T method. Biodistribution was observed in an LS180 human colon cancer xenograft model. Additionally, autoradiographic imaging and immunohistochemical staining of tumors were performed with 125 I-VEGF 121 . Results 125 I-VEGF 121 and 125 I-VEGF 165 exhibited strong, continuous uptake by tumors and the uterus, an organ characterized by angiogenesis. 125 I-VEGF 121 uptake in tumors was twofold higher than that of 125 I-VEGF 165 (9.12±98 and 4.79±1.08 %ID/g at 2 h, respectively). 125 I-VEGF 121 displayed higher tumor to nontumor (T/N) ratios in most normal organs in comparison with 125 I-VEGF 165 . 125 I-VEGF 121 accumulation in tumors decreased with increasing tumor volume. Autoradiographic and immunohistochemical analyses confirmed that the difference in 125 I-VEGF 121 tumor accumulation correlated with degree of tumor vascularity. Conclusion Radioiodinated VEGF 121 is a promising tracer for noninvasive delineation of angiogenesis in vivo.

64 Cu-labeled diacetyl-bis(N 4 -methylthiosemicarbazone) ( 64 Cu-ATSM) is a promising agent for internal radiation therapy and imaging of hypoxic tissues. In the present study, the intra-tumoral distribution of 64 Cu-ATSM was investigated by comparing it to that of [ 18 F]FDG and histological findings. VX2 tumors were implanted into Japanese white rabbits subcutaneously. 64 Cu-ATSM and [ 18 F]FDG were co-injected intravenously and the tumor was dissected and cut into 1 mm thick slices 1 h after the injection. The uptake of 64 Cu-ATSM and [ 18 F]FDG was measured using a dual-tracer autoradiographic technique. Histological cell biology was estimated from the optical microscopy of tumor sections. The major accumulation of 64 Cu-ATSM was observed around the outer rim of the tumor masses which consisted mainly of active cells and expected to be hypoxic. [ 18 F]FDG was distributed more widely with highest levels in the inner regions where pre-necrotic cells were mainly observed. 64 Cu-ATSM appears to be useful for the detection of hypoxic but active tumor cell regions in vivo.

Introduction Several lines of evidence suggest that C-7α-substituted estradiol derivatives are well tolerated by estrogen receptor (ER). In line with this hypothesis, we are interested in the design and synthesis of C-7α-substituted estrogens as molecular probes to visualize ER function. Methods We have synthesized 7α-(3-[ 18 F]fluoropropyl) estradiol (C3-7α-[ 18 F]FES) as a potential radiopharmaceutical for ER imaging by positron emission tomography (PET). In vitro receptor binding and in vivo biodistribution and blocking studies in mature female mice, and in vivo metabolite analysis were carried out. Furthermore, in vivo ER-selective uptake was confirmed using ER-positive T-47D and ER-negative MDA-MB-231 tumor-bearing mice. We also compared the in vivo biodistribution of C3-7α-[ 18 F]FES with 16α-[ 18 F]FES. Results C3-7α-[ 18 F]FES was produced in moderate yields (30.7% ± 15.1%, decay corrected) with specific activity of 32.0 ± 18.1 GBq/μmol (EOS). The in vitro binding affinity of C3-7α-FES to the ERα isoform was sufficient and equivalent to that of estradiol. C3-7α-[ 18 F]FES showed selective uptake in ER-rich tissues, such as the uterus (4.7%ID/g ± 1.2%ID/g at 15 minutes) and ovary (4.0%ID/g ± 1.0%ID/g at 5 minutes). The tissue time activity curves of these organs showed reversible kinetics, indicating suitability for quantitative analysis. The highest contrast was obtained at 120 minutes after injection of C3-7α-[ 18 F]FES in the uterus (uterus/blood = 18, uterus/muscle = 17.3) and ovary (ovary/blood = 6.3, ovary/muscle = 6.0). However, the level of selective uptake of C3-7α-[ 18 F]FES was significantly lower than that of 16α-[ 18 F]FES. Most radioactivity in the uterus was detected in unchanged form, although peripherally C3-7α-[ 18 F]FES was rapidly degraded to hydrophilic metabolites. In accordance with this peripheral metabolism, gradual increases in bone radioactivity were observed, indicating defluorination. Coinjection with estradiol dose-dependently inhibited C3-7α-[ 18 F]FES uptake in the uterus and ovary. The in vivo IC 50 values of estradiol in the uterus and ovary were 34.4 and 38.5 nmol/kg, respectively. Furthermore, in vivo tumor uptake of C3-7α-[ 18 F]FES was significantly higher (unpaired t test with Welch’s correction; p = 0.015) in ER-positive T-47D tumors (2.3%ID/g ± 0.4%ID/g) than ER-negative MDA-MB-231 tumors (0.9%ID/g ± 0.1%ID/g). Conclusions Although extensive metabolism was observed in rodents, C3-7α-[ 18 F]FES showed promising results for quantitative analysis of ER density in vivo . However, the selective uptake of C3-7α-[ 18 F]FES was lower than that of 16α-[ 18 F]FES. Further optimizations and structure–activity relationship studies of the C-7α-substituted estradiol are needed.

During the past decade, the efficacy of new molecular targeted drugs such as tyrosine kinase inhibitors (TKIs) and monoclonal antibodies has been proven worldwide, and molecular targeted therapies have become the mainstream in cancer therapy. However, clinical use of these new drugs presents unexpected adverse effects or poor therapeutic effects. Therefore, we require diagnostic tools to estimate the target molecule status in cancer tissues and predict therapeutic efficacy and adverse effects. Although immunohistochemical, polymerase chain reaction (PCR) and fluorescence in situ hybridization (FISH) analyses of biopsy samples are conventional and popular for this diagnostic purpose, molecular imaging modalities such as positron emission tomography (PET) and single photon emission computed tomography (SPECT) are also useful for noninvasive estimation of gene and protein expression and drug pharmacokinetics. In this review, we introduce new radiolabeled TKIs, antibodies, and their clinical application in molecular targeted therapy and discuss the issues of these imaging probes.

A series of iodinated analogues of MD-230254 was synthesized and evaluated for inhibitory potency and selectivity toward monoamine oxidase B (MAO-B). Among them, 5-[4-(2-iodobenzyloxy)phenyl]-3-(cyanoethyl)-1,3,4-oxadiazole-2(3H)one (2-IBPO) was found to have high inhibitory potency and selectivity toward MAO-B (IC50=2.0 nM, MAO-A/MAO-B >50000). Analysis of the inhibition kinetics indicated that 2-IBPO acts in a two-step mechanism as a competitive, slow, and tight-binding inhibitor of MAO-B with a Ki value of 2.4 nM and an overall Ki* value at an equilibrium of 3.8 nM. The new radioligand for MAO-B, [125I]2-IBPO was conveniently synthesized from a tributylstannyl precursor by an iododestannylation reaction using sodium [125I]iodide and hydrogen peroxide with high radiochemical yield. The in vivo tissue distribution studies of [125I]2-IBPO demonstrated its high initial uptake and prolonged retention in the brain. A selective interaction of [125I]2-IBPO with MAO-B was confirmed by the pretreatment experiment with well known MAO specific inhibitors, l-deprenyl, Ro-16-6491, clorgyline, and Ro-41-1049. These very desirable characteristics of [125I]2-IBPO suggested that a 123I-labeled counterpart, [123I]2-IBPO, would have great potential in in vivo studies of MAO-B in the human brain with single photon emission computed tomography (SPECT).

To reveal the metabolic fate of acetate in neoplasms that may characterize the accumulation patterns of [1-(11)C]acetate in tumors depicted by positron emission tomography. Four tumor cell lines (LS174T, RPMI2650, A2780, and A375) and fibroblasts in growing and resting states were used. In uptake experiments, cells were incubated with[1-(14)C]acetate for 40 min. [(14)C]CO(2) was measured in the tight-air chamber, and the metabolites in cells were identified by thin layer chromatography and paper chromatography. The glucose metabolic rate of each cell line was measured with [2,6-(3)H]2-deoxy-glucose (DG), and the growth activity of each cell line was estimated by measuring the incorporation of [(3)H]methyl thymidine into DNA. Compared with resting fibroblasts, all four tumor cell lines showed higher accumulation of (14)C activity from [1-(14)C]acetate. These tumor-to-normal ratios of [1-(14)C]acetate were larger than those of DG. Tumor cells incorporated (14)C activity into the lipid-soluble fraction, mostly of phosphatidylcholine and neutral lipids, more prominently than did fibroblasts. The lipid-soluble fraction of (14)C accumulation in cells showed a positive correlation with growth activity, whereas the water-soluble and CO(2) fractions did not. These findings suggest that the high tumor-to-normal ratio of [1-(14)C]acetate is mainly due to the enhanced lipid synthesis, which reflects the high growth activity of neoplasms. This in vitro study suggests that [1-(11)C]acetate is appropriate for estimating the growth activity of tumor cells.

To investigate age-related changes in cerebral energy production, we compared senescence-accelerated prone mice (SAMP8) as an animal model of accelerated aging and senescence-accelerated resistant mice (SAMR1) as a control. Considering that the cerebral glucose metabolic rate (CMRglc) at the time of O2 deprivation and 2,4-dinitrophenol (DNP) loading would reflect anaerobic glycolytic capacity and mitochondrial function, respectively, we investigated dynamic changes in CMRglc before and after loading with these perturbations. Fresh brain slices were incubated with [18F]2-fluoro-2-deoxy-D-glucose ([18F]FDG) in oxygenated Krebs-Ringer solution at 36°C, and serial two-dimensional time-resolved images of [18F]FDG uptake in these slices were obtained on the imaging plates. The fractional rate constant (=k3*) of [18F]FDG proportional to the CMRglc was evaluated by applying the Gjedde-Patlak graphical method to the image data. The k3* value before the hypoxic perturbation in all of the brain sites analyzed was higher in SAMP8 than SAMR1 in both the 2- and 10-month-old groups. With O2 deprivation, k3* values were higher without site specificity in the 2-month-old SAMP8 than in 2-month-old SAMR1, whereas in 10-month-old mice, there was no significant difference between the two groups. In contrast, with DNP loading, while no significant difference was noted between 2-month-old SAMP8 and 2-month-old SAMR1, in 10-month-old mice, the SAMP8 group showed lower values in certain regions than SAMR1 mice. These results suggest that in the brain tissue of SAMP8, a marked transient enhancement of anaerobic glycolytic capacity in the 2-month-olds and a decrease in mitochondrial function in the subsequent period occur, as a result of which glucose metabolism appears to be enhanced in both the 2- and 10-month-old groups compared to SAMR1 mice.

Using a positron autoradiography technique, dynamic changes in the cerebral glucose metabolic rate (CMRglc) induced by hypoxia/reoxygenation were investigated in living brain slices. After incubating fresh rat brain slices (300 microm thick) with [(18)F]2-fluoro-2-deoxy-D-glucose ([(18)F]FDG) in oxygenated Krebs-Ringer solution at 36 degrees C, serial two-dimensional time-resolved images of [(18)F]FDG uptake in the slices were obtained on imaging plates. As compared to the unloaded control values, with hypoxia-loading [(18)F]FDG uptake increased markedly, suggesting enhanced glycolysis. The net influx constant (K) of [(18)F]FDG at pre-hypoxia-loading and after reoxygenation with loading hypoxia for various periods of time was quantitatively evaluated by applying the Patlak graphical method to the image data. Regardless of the brain region, with hypoxia of/=10-min duration, the K value returned to the preloading level, whereas with hypoxia of/=20 min duration only partial or no recovery was seen, indicating irreversible neuronal damage. The 30-min administration of either N-methyl-D-aspartate (NMDA)/non-NMDA antagonist or a free radical scavenger at the same time as reoxygenation after 20 min hypoxia showed a neuroprotective effect inhibiting the decrease in the post-hypoxia-loading K value. In contrast, no such neuroprotective effect was evident with administration of either of these agents only during hypoxia loading, possibly indicating that immediately after reoxygenation neuronal damage was induced mediated by excitatory amino acids and free radicals in tandem. These results demonstrate that serial quantitative evaluation of CMRglc using this technique may be of use in investigating the brain tissue injury associated with hypoxia/reoxygenation as well as clarifying the underlying mechanisms and protective effect of various drugs against such injury.

Dynamic changes in the regional cerebral glucose metabolic rate induced by hypoxia/reoxygenation or ischemia/reperfusion were investigated with a positron autoradiography technique. Fresh rat brain slices were incubated with [18F]2-fluoro-2-deoxy-D-glucose ([18F]FDG) in oxygenated Krebs-Ringer solution at 36°C, and serial two-dimensional time-resolved images of [18F]FDG uptake in the slices were obtained. In the case of loading hypoxia (oxygen deprivation)/pseudoischemia (oxygen and glucose deprivation) for various periods of time, the net influx constant ( K) of [18F]FDG at preloading and after reoxygenation/pseudoreperfusion (post-loading) was quantitatively evaluated by applying the Patlak graphical method to the image data. Regardless of the brain region, with hypoxia lasting ≥20 minutes, the postloading K value was decreased compared with the unloaded control, whereas with pseudoischemia of ≤40 minutes, approximately the same level as the unloaded control was maintained. Next, the neuroprotective effect against hypoxia/pseudoischemia loading induced by the addition of a free radical scavenger or an N-methyl-D-aspartate (NMDA) antagonist was assessed by determining whether a decrease in the postloading K value was prevented. Whereas with 20-minute hypoxia, both agents exhibited a neuroprotective effect, in the case of 50-minute pseudoischemia, only the NMDA antagonist did so, with the free radical scavenger being ineffective. These results demonstrate that hypoxia causes irreversible neuronal damage within a shorter period than ischemia, with both free radicals and glutamate suggested to be involved in tandem in the neurotoxicity induced by hypoxia, whereas glutamate alone is involved in ischemic neurotoxicity.

Epidermal growth factor receptor (EGFR) is attractive target for tumor diagnosis and therapy, as it is specifically and abundantly expressed in tumor cells. EGFR-tyrosine kinase (TK) inhibitors such as gefitinib and erlotinib are widely used in the treatment of non-small cell lung cancer (NSCLC). In this study, we investigated whether radioiodinated 4-(3-iodo-phenoxy)-6,7-diethoxy-quinazoline (PHY), which is a candidate EGFR-TK imaging agent for single photon emission computed tomography (SPECT) is able to predict gefitinib sensitivity. We used four NSCLC cell lines-A549 (wild-type EGFR), H1650 (mutant EGFR; del E746_A750), H1975 (mutant EGFR; L858R, T790M) and H3255 (mutant EGFR; L858R)-and one epidermoid carcinoma cell line, A431 (wild-type EGFR). Cell proliferation assay and Western blotting revealed that A431 and H3255 with high EGFR expression showed high sensitivity to gefitinib. On the other hand, A549, H1650 and H1975 showed much lower sensitivity to gefitinib. The blocking study revealed that gefitinib decreased tumor uptake in (125)I-PHY in A431-bearing mice. Moreover, in vivo tumor uptake of (125)I-PHY was correlated with the IC50 of gefitinib for cell proliferation. In the present study, tumor uptake of (125)I-PHY was correlated with the gefitinib sensitivity and this uptake was based on expression levels of EGFR, but not on mutation status. Although the mutation status is the most important factor for predicting gefitinib sensitivity, the abundant expression of EGFR is essential for therapy with EGFR-TK inhibitors. Therefore, radioiodinated PHY is a potential imaging agent to predict gefitinib sensitivity based on EGFR expression levels though further modifications of the imaging agent is needed to accurately estimate the mutation status.

Purpose (64)Cu-diacetyl-bis (N (4)-methylthiosemicarbazone) ((64)Cu-ATSM) is a promising theranostic agent that targets hypoxic regions in tumors related to malignant characteristics. Its diagnostic usefulness has been recognized in clinical studies. Internal radiotherapy (IRT) with (64)Cu-ATSM is reportedly effective in preclinical studies; however, for clinical applications, improvements to reduce radiation exposure in non-target organs, particularly the liver, are required. We developed a strategy to reduce radiation doses to critical organs while preserving tumor radiation doses by controlled administration of copper chelator penicillamine during (64)Cu-ATSM IRT. Methods Biodistribution was evaluated in HT-29 tumor-bearing mice injected with (64)Cu-ATSM (185 kBq) with or without oral penicillamine administration. The appropriate injection interval between (64)Cu-ATSM and penicillamine was determined. Then, the optimal penicillamine administration schedule was selected from single (100, 300, and 500 mg/kg) and fractionated doses (100 mg/kg×3 at 1- or 2-h intervals from 1 h after (64)Cu-ATSM injection). PET imaging was performed to confirm the effect of penicillamine with a therapeutic (64)Cu-ATSM dose (37 MBq). Dosimetry analysis was performed to estimate human absorbed doses. Results Penicillamine reduced (64)Cu accumulation in the liver and small intestine. Tumor uptake was not affected by penicillamine administration at 1 h after (64)Cu-ATSM injection, when radioactivity was almost cleared from the blood and tumor uptake had plateaued. Of the single doses, 300 mg/kg was most effective. Fractionated administration at 2-h intervals further decreased liver accumulation at later time points. PET indicated that penicillamine acts similarly with the therapeutic (64)Cu-ATSM dose. Dosimetry demonstrated that appropriately scheduled penicillamine administration reduced radiation doses to critical organs (liver, ovaries, and red marrow) below tolerance levels. Laxatives reduced radiation doses to the large intestine. Conclusions We developed a novel strategy to reduce radiation exposure in critical organs during (64)Cu-ATSM IRT, thus promoting its clinical applications. This method could be beneficial for other (64)Cu-labeled compounds.

Introduction 4-Borono-2- 18 F-fluoro-phenylalanine ( 18 F-FBPA) has been used to anticipate the therapeutic effects of boron neutron capture therapy (BNCT) with 4-borono-L-phenylalanine (BPA). Similarly, L-[methyl- 11 C]-methionine ( 11 C-MET), the most popular amino acid PET tracer, is a possible candidate for this purpose. We investigated the transport mechanism of 18 F-FBPA and compared it with that of 14 C-MET in human glioblastoma cell lines. Methods Uptake of 18 F-FBPA and 14 C-MET was examined in A172, T98G, and U-87MG cells using 2-aminobicyclo-(2.2.1)-heptane-2-carboxylic acid (a system L-specific substrate), 2-(methylamino)-isobutyric acid (a system A-specific substrate), and BPA. Gene expression was analyzed by quantitative real time polymerase chain reaction. Results System L was mainly involved in the uptake of 18 F-FBPA (74.5%–81.1% of total uptake) and 14 C-MET (48.3%–59.4%). System A and ASC also contributed to the uptake of 14 C-MET. Inhibition experiments revealed that BPA significantly decreased the uptake of 18 F-FBPA, whereas 31%–42% of total 14 C-MET uptake was transported by BPA non-sensitive transporters. In addition, 18 F-FBPA uptake correlated with LAT1 and total LAT expressions. Conclusion This study demonstrated that 18 F-FBPA was predominantly transported by system L in human glioblastoma cells compared to 14 C-MET. Although further studies are needed to elucidate the correlation between 18 F-FBPA uptake and BPA content in tumor tissues, 18 F-FBPA is suitable for the selection of patients who benefit from BNCT with BPA.