[18F]2 was eluted by back-flushing with ethanol in a portion of 0.5mL ( 2 mL total). 13C NMR studies. Computational studies of model compounds also support the above proposed mechanism. Similarly, the ring-opening and re-closure method was Oligomycin used successfully in the synthesis of the 11C labeled isatin sulfonamide analogue [11C]4 (WC-98). A microPET imaging study using [11C]4 in the Fas liver apoptosis model demonstrated retained activity in the target organ (liver) of the treated mice. Increased caspase-3 activation in the liver was verified by the fluorometric caspase-3 enzyme assay. Therefore, this study provides a useful method for radio-synthesis of isatin derivative radiotracers for PET and SPECT studies, and [11C]4 is a potential PET radiotracer for noninvasive imaging of apoptosis. imaging techniques used in molecular Oligomycin imaging, is being used more frequently in clinical and research fields because of its high sensitivity, minimal physiological effect from PET tracers, good spatial resolution and ease of accurate quantification. One important application of molecular imaging is to study programmed cell death (apoptosis) at the molecular level. Apoptosis is critical for the normal development and function of multicellular organisms as a common and universal mechanism of cell death.2 The abnormal regulation of Oligomycin cellular death via apoptosis is believed to play a key role in a variety of human diseases.3 In addition, the beneficial effect of chemotherapy, radiotherapy, and other antitumor Mouse monoclonal to OPN. Osteopontin is the principal phosphorylated glycoprotein of bone and is expressed in a limited number of other tissues including dentine. Osteopontin is produced by osteoblasts under stimulation by calcitriol and binds tightly to hydroxyapatite. It is also involved in the anchoring of osteoclasts to the mineral of bone matrix via the vitronectin receptor, which has specificity for osteopontin. Osteopontin is overexpressed in a variety of cancers, including lung, breast, colorectal, stomach, ovarian, melanoma and mesothelioma. therapies can be attributed to their activation of the apoptotic process.4 Therefore, the development of a noninvasive imaging procedure that can study the process of apoptosis in a variety of disease states and monitor the ability of a drug or other treatment either to induce or to halt apoptosis would be of tremendous value to the research and clinical community. One of the most commonly used agents so far for imaging apoptosis is based on Annexin V, which is a 36 kDa protein that binds selectively with high affinity to phosphatidylserine, a protein that is externalized in the early stages of apoptosis after the activation of caspase-3. Annexin V has been labeled with different radioisotopes for PET and single photon emission computed tomography (SPECT) studies.5 99mTc-labeled Annexin V using SPECT showed promising results and is undergoing clinical trials.6 However, since the externalization of phosphatidylserine also occurs in necrosis, radiolabeled Annexin V is not specific for imaging apoptosis microPET imaging study with the Fas liver injury model in mice. Results and Discussion Synthesis Oligomycin and Radiolabeling The syntheses of standard compounds 2, 4 and precursors 7a, 7b and 12 for 18F labeling of 2 and 8, and 13 for 11C labeling Oligomycin of 4, are shown in Scheme 1. The isatin nitrogen of 5-(2-phenoxymethyl-pyrrolidine-sulfonyl)-1H-2,3-dione 3 was alkylated by treatment of 3 with sodium hydride in DMF at 0 C for 20 min and followed by addition of various alkyl halides to give compounds 2, 4, 5, and 6 respectively. Compound 5 was then heated to reflux with silver methanesulfonate or silver microPET study of Fas-treated liver injury apoptosis model Once the radiolabeling conditions were established, [11C]4 (WC-98) was evaluated in a well-characterized mouse model of liver apoptosis induced by administration of anti-Fas (Jo2) antibody, which results in massive caspase-3 activation in the liver. Both microPET imaging and biodistribution studies were performed.25 The microPET imaging results are shown in Figure 1. There is clearly retained activity in the liver in the Fas-treated mice compared to the control, with retention of the activity in the liver demonstrated on time-activity curves (Figure 1). The analysis of the liver is illustrated in Figure 2. At 5 min, there is no difference in liver uptake between treated and control mice; however, at 30 min, there is clearly increased tracer activity in the liver samples taken from the treated mice compared with controls. Liver samples from the biodistribution study were taken and analyzed for caspase-3 activation by fluorometric enzyme analysis using the caspase-3 fluorogenic substrate Ac-DEVD-AMC (Biomol) (Figure 3). Enzyme analysis verified increased caspase-3 enzymatic activity in the treated animals and correlated with the findings from the microPET imaging and biodistribution studies. Since [11C]4 (WC-98) is a competitive inhibitor of caspase-3 and binds to the activated form of caspase-3 in tissues undergoing apoptosis, we see expected retention of the tracer in the liver of the treated mice. The results obtained with [18F]4 suggest again that radiolabeled evaluation of [11C]4 (WC-98) uptake in liver comparing mice treated with the Fas antibody to untreated controls..
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