Nuclear Medicine and Biology - Articles in Press

Ex Vivo and In Vivo Evaluation of the Norepinephrine Transporter Ligand [11C]MRB for Brown Adipose Tissue Imaging - Corrected Proof

2012-05-18

Abstract: Introduction: It has been suggested that brown adipose tissue (BAT) in humans may play a role in energy balance and obesity. We conducted ex vivo and in vivo evaluation using [11C]MRB, a highly selective NET (norepinephrine transporter) ligand for BAT imaging at room temperature, which is not achievable with [18F]FDG.Methods: PET images of male Sprague–Dawley rats with [18F]FDG and [11C]MRB were compared. Relative [18F]FDG or [11C]MRB retention at 20, 40 and 60min post-injection was quantified on awake rats after exposing to cold (4°C for 4h) or remaining at room temperature. Rats pretreated with unlabeled MRB or nisoxetine 30min before [11C]MRB injection were also assessed. The [11C]MRB metabolite profile in BAT was evaluated.Results: PET imaging demonstrated intense [11C]MRB uptake (SUV of 2.9 to 3.3) in the interscapular BAT of both room temperature and cold-exposed rats and this uptake was significantly diminished by pretreatment with unlabeled MRB; in contrast, [18F]FDG in BAT was only detected in rats treated with cold. Ex vivo results were concordant with the imaging findings; i.e. the uptake of [11C]MRB in BAT was 3 times higher than that of [18F]FDG at room temperature (P=0.009), and the significant cold-stimulated uptake in BAT with [18F]FDG (10-fold, P=0.001) was not observed with [11C]MRB (P=0.082). HPLC analysis revealed 94%–99% of total radioactivity in BAT represented unchanged [11C]MRB.Conclusions: Our study demonstrates that BAT could be specifically labeled with [11C]MRB at room temperature and under cold conditions, supporting a NET-PET strategy for imaging BAT in humans under basal conditions.

[18F]desmethoxyfallypride as a novel PET radiotracer for quantitative in vivo dopamine D2/D3 receptor imaging in rat models of neurodegenerative diseases - Corrected Proof

2012-05-17

Abstract: Introduction: [18F]desmethoxyfallypride ([18F]DMFP) is a promising tracer for longitudinal assessment of striatal dopamine D2/D3-receptor (D2R) availability by positron emission tomography (PET) in small animal models. We explored the feasibility of [18F]DMFP-PET to image D2R availability in rat models of Huntington's (HD) and Parkinson's disease (PD).Methods: Animals received either unilateral intrastriatal quinolinic acid lesions or medial forebrain bundle injections of 6-OHDA to produce the loss of striatal projection neurones or deplete the striatal dopamine, corresponding to established animal models for HD and PD, respectively. Three weeks after lesioning, PET scans were acquired on a microPET Focus 120 system following the tail vein injection of [18F]DMFP.Results: [18F]DMFP-PET clearly visualized lesion induced decreases and increases of D2R availability. In vivo estimates of D2R binding and changes thereof gained by pharmacokinetic analyses correlated significantly with D2R density and its change provided by in vitro [3H]raclopride-autoradiography.Conclusions: In conclusion, [18F]DMFP-PET is a suitable method for in vivo D2R-assessment in preclinical research, e.g for monitoring cell-based therapies.

Automated GMP Synthesis of [18F]ICMT-11 for In Vivo Imaging of Caspase-3 Activity - Corrected Proof

Robin Fortt, Graham Smith, Ramla O. Awais, Sajinder K. Luthra, Eric O. Aboagye — 2012-05-10

Abstract: Introduction: Isatin-5-sulfonamide ([18F]ICMT-11) is a sub-nanomolar inhibitor of caspase-3 previously evaluated as an apoptosis imaging agent. Herein, an alternative radiosynthesis of [18F]ICMT-11 with increased purity and specific activity is presented. Finally, a GMP-applicable automated radiosynthesis of [18F]ICMT-11 is described.Methods: The preparation of [18F]ICMT-11 was evaluated under a variety of reaction conditions, including reaction solvent, by employing alternative phase transfer catalysts and under different deprotection conditions. Following initial investigations, the process was transferred onto a fully automated GE FASTlab synthesis platform for further development and optimisation.Results: The synthesis of [18F]ICMT-11 was successfully validated under GMP conditions, resulting in a yield of 4.6±0.4GBq with a radiochemical purity of >98% at EOS and a specific activity of 685±237GBq/μmol within 90 min. Quality control was carried out in accordance with the European Pharmacopoeia and demonstrated that [18F]ICMT-11 can be consistently manufactured on the FASTlab to meet specifications.Conclusions: A simplified methodology for the synthesis of the apoptosis imaging agent, [18F]ICMT-11, has been achieved by the SN2 displacement of a tosylate leaving group with [18F]fluoride ion. This results in an increased purity and specific activity over the original copper catalysed “Click” synthetic stratagem reaction involving 2-[18F]fluoroethylazide with an alkyne precursor and is now suitable for routine clinical application.

Synthesis of [18F]-labeled (2-(2-fluoroethoxy)ethyl)tris(4-methoxyphenyl)phosphonium cation as a potential agent for positron emission tomography myocardial imaging - Corrected Proof

Dong-Yeon Kim, Hee-Jung Kim, Kook-Hyun Yu, Jung-Joon Min — 2012-05-10

Abstract: Introduction: Established radiotracers for positron emission tomography (PET) myocardial perfusion study are commonly labeled with short-lived radio-isotopes that limit their widespread clinical use. Thus, we synthesized (2-(2-[18F]fluoroethoxy)ethyl)tris(4-methoxyphenyl)phosphonium salt ([18F]FETMP) as a novel myocardial perfusion agent that penetrates the hydrophobic barriers of the plasma and mitochondrial membranes and accumulate in mitochondria of cardiomyocytes in response to the negative inner-transmembrane potentials.Methods: The [18F]FETMP was synthesized via two-step nucleophilic substitution reactions of no-carrier-added [18F]fluoride with the precursor 2,2′-oxybis(ethane-2,1-diyl)bis(4-methylbenzenesulfonate) in the presence of Kryptofix 2.2.2 and K2CO3. The [18F]FETMP accumulation was measured in cell culture with rat embryonic cardiomyoblast (H9c2) and mouse normal fibroblast (NIH/3T3) cell lines. The mitochondrial membrane potential-dependent cellular uptake of [18F]FETMP was further assessed using the H9c2 cells treated with carbonyl cyanide m-chlorophenylhydrazone (CCCP) which is a protonophore that selectively abolishes the mitochondrial membrane potential. Biodistribution and micro-PET studies were performed in normal BALB/c mice to test and optimize the kinetics for radiolabeled phosphonium cation.Results: The radiolabeled compound was synthesized with 10%–20% yield. The radiochemical purity was >98% by analytical HPLC, and the specific activity was >5.92TBq/μmol. The cellular uptake assay showed preferential uptake of [18F]FETMP in cardiomyocytes. The results of biodistribution and micro-PET imaging studies of [18F]FETMP in mice and rats showed preferential accumulation in the myocardium.Conclusions: The results suggest that [18F]FETMP would be a promising candidate for myocardial imaging and might be useful for clinical cardiac PET/CT applications.

The synthesis and in vivo evaluation of [18F]PF-9811: a novel PET ligand for imaging brain fatty acid amide hydrolase (FAAH) - Corrected Proof

2012-05-09

Abstract: Introduction: Fatty acid amide hydrolase (FAAH) is responsible for the enzymatic degradation of the fatty acid amide family of signaling lipids, including the endogenous cannabinoid (endocannabinoid) anandamide. The involvement of the endocannabinoid system in pain and other nervous system disorders has made FAAH an attractive target for drug development. Companion molecular imaging probes are needed, however, to assess FAAH inhibition in the nervous system in vivo. We report here the synthesis and in vivo evaluation of [18F]PF-9811, a novel PET ligand for non-invasive imaging of FAAH in the brain.Methods: The potency and selectivity of unlabeled PF-9811 were determined by activity-based protein profiling (ABPP) both in vitro and in vivo. [18F]PF-9811 was synthesized in a 3-step, one-pot reaction sequence, followed by HPLC purification. Biological evaluation was performed by biodistribution and dynamic PET imaging studies in male rats. The specificity of [18F]PF-9811 uptake was evaluated by pre-administration of PF-04457845, a potent and selective FAAH inhibitor, 1h prior to radiotracer injection.Results: Biodistribution studies show good uptake (SUV~0.8 at 90min) of [18F]PF-9811 in rat brain, with significant reduction of the radiotracer in all brain regions (37%–73% at 90min) in blocking experiments. Dynamic PET imaging experiments in rat confirmed the heterogeneous uptake of [18F]PF-9811 in brain regions with high FAAH enzymatic activity, as well as statistically significant reductions in signal following pre-administration of the blocking compound PF-04457845.Conclusions: [18F]PF-9811 is a promising PET imaging agent for FAAH. Biodistribution and PET imaging experiments show that the tracer has good uptake in brain, regional heterogeneity, and specific binding as determined by blocking experiments with the highly potent and selective FAAH inhibitor, PF-04457845.

18F-fluorodeoxyglucose positron emission tomography in management of pancreatic cystic tumors - Corrected Proof

2012-05-07

Abstract: Objectives: To evaluate the effectiveness of PET in differentiating malignant from benign pancreatic cystic tumors.Methods: Between 2009 and 2010, all patients with pancreatic cystic tumors who had PET, triple phase contrast computed tomography (CT) and endoscopic ultrasound (EUS) were reviewed. Clinicopathological characteristics and final histology were correlated with preoperative PET, CT and EUS to assess the value of each modality in detecting malignant from benign lesions for clinical decision-making.Results: Twenty of a total of 116 patients with pancreatic cystic tumors had 18F-FDG PET because of diagnostic difficulties after evaluation with conventional modalities. Sensitivity and specificity of PET in differentiating malignant from benign pancreatic cystic tumors were 100% and 93.75%, with an accuracy of 95%. PET had the best sensitivity, specificity and accuracy for detecting malignant cystic tumors compared with CT and EUS. In 5 cases, the PET results altered the treatment options completely to follow-up instead of surgery (n=2), limited resection instead of Whipple's resection (n=1), and surgery instead of follow-up (n=2).Conclusions: PET is an accurate, non-invasive method to distinguish malignant from benign pancreatic cystic tumors and can be used as an adjunct to facilitate clinical decision making.

The effects of 5-fluoruracil treatment on 3′-fluoro-3′-deoxythymidine (FLT) transport and metabolism in proliferating and non-proliferating cultures of human tumor cells - Corrected Proof

David A. Plotnik, Lena J. McLaughlin, Kenneth A. Krohn, Jeffrey L. Schwartz — 2012-05-07

Abstract: 3′-Fluoro-3′-deoxythymidine (FLT) positron emission tomography (PET) has been proposed for imaging thymidylate synthase (TS) inhibition. Agents that target TS and shut down de novo synthesis of thymidine monophosphate increase the uptake and retention of FLT in vitro and in vivo because of a compensating increase in the salvage pathway. Increases in both thymidine kinase-1 (TK1) and the equilibrative nucleoside transporter hENT1 have been reported to underlie this effect. We examined whether the effects of one TS inhibitor, 5-fluorouracil (5FU), on FLT uptake require proliferating cells and whether the effects are limited to increasing TK1 activity.Methods: The effects of 5FU on FLT transport and metabolism, TK1 activity, and cell cycle progression were evaluated in the human tumor cell line, A549, maintained as either a proliferating or non-proliferating culture.Results: There were dose-dependent increases in FLT uptake that peaked after a 10μM 5FU exposure and then declined to baseline levels or below at higher doses in both proliferating and non-proliferating cultures. The dose-dependence for FLT uptake was mirrored by changes in TK1 activity. S phase fraction did not correlate with FLT uptake in proliferating cultures. Chemical inhibition of hENT1 reduced overall levels of FLT uptake but did not affect the low dose increase in FLT uptake.Conclusions: 5FU only affects FLT uptake in proliferating A549 cells and increases in FLT uptake are directly related to increased TK1 activity. Our studies did not support a role for hENT1 in the increased uptake of FLT after exposure to 5FU. Our studies with A549 cells support the suggestion that FLT-PET could provide a measure of TS inhibition in vivo.

Acute treatment with fluvoxamine elevates rat brain serotonin synthesis in some terminal regions: An autoradiographic study - Corrected Proof

Dorotea Muck-Seler, Nela Pivac, Mirko Diksic — 2012-05-07

Abstract: Introduction: A considerable body of evidence indicates the involvement of the neurotransmitter serotonin (5-HT) in the pathogenesis and treatment of depression.Methods: The acute effect of fluvoxamine, on 5-HT synthesis rates was investigated in rat brain regions, using α-14C-methyl-L-tryptophan as a tracer. Fluvoxamine (25 mg/kg) and saline (control) were injected intraperitoneally, one hour before the injection of the tracer (30 μCi).Results: There was no significant effect of fluvoxamine on plasma free tryptophan. After Benjamini–Hochberg False Discovery Rate correction, a significant decrease in the 5-HT synthesis rate in the fluvoxamine treated rats, was found in the raphe magnus (−32%), but not in the median (−14%) and dorsal (−3%) raphe nuclei. In the regions with serotonergic axon terminals, significant increases in synthesis rates were observed in the dorsal (+41%) and ventral (+43%) hippocampus, visual (+38%), auditory (+65%) and parietal (+37%) cortex, and the substantia nigra pars compacta (+56%). There were no significant changes in the 5-HT synthesis rates in the median (+11%) and lateral (+24%) part of the caudate-putamen, nucleus accumbens (+5%), VTA (+16%) or frontal cortex (+ 6%).Conclusions: The data show that the acute administration of fluvoxamine affects 5-HT synthesis rates in a regionally specific pattern, with a general elevation of the synthesis in the terminal regions and a reduction in some cell body structures. The reasons for the regional specific effect of fluvoxamine on 5-HT synthesis are unclear, but may be mediated by the presynaptic serotonergic autoreceptors.

Synthesis and evaluation of 18F labeled alanine derivatives as potential tumor imaging agents - Corrected Proof

2012-04-30

Abstract: Introduction: This paper reports the synthesis and labeling of 18F alanine derivatives. We also investigate their biological characteristics as potential tumor imaging agents mediated by alanine–serine–cysteine preferring (ASC) transporter system.Methods: Three new 18F alanine derivatives were prepared from corresponding tosylate-precursors through a two-step labeling reaction. In vitro uptake studies to evaluate and to compare these three analogs were carried out in 9L glioma and PC-3 prostate cancer cell lines. Potential transport mechanisms, protein incorporation and stability of 3-(1-[18F]fluoromethyl)-L-alanine (L-[18F]FMA) were investigated in 9L glioma cells. Its biodistribution was determined in a rat-bearing 9L tumor model. PET imaging studies were performed on rat bearing 9L glioma tumors and transgenic mouse carrying spontaneous generated M/tomND tumor (mammary gland adenocarcinoma).Results: New 18F alanine derivatives were prepared with 7%–34% uncorrected radiochemical yields, excellent enantiomeric purity (>99%) and good radiochemical purity (>99%). In vitro uptake of the L-[18F]FMA in 9L glioma and PC-3 prostate cancer cells was higher than that observed for the other two alanine derivatives and [18F]FDG in the first 1h. Inhibition of cell uptake studies suggested that L-[18F]FMA uptake in 9L glioma was predominantly via transport system ASC. After entering into cells, L-[18F]FMA remained stable and was not incorporated into protein within 2h. In vivo biodistribution studies demonstrated that L-[18F]FMA had relatively high uptake in liver and kidney. Tumor uptake was fast, reaching a maximum within 30min. The tumor-to-muscle, tumor-to-blood and tumor-to-brain ratios at 60min post injection were 2.2, 1.9 and 3.0, respectively. In PET imaging studies, tumors were visualized with L-[18F]FMA in both 9L rat and transgenic mouse.Conclusion: L-[18F]FMA showed promising properties as a PET imaging agent for up-regulated ASC transporter associated with tumor proliferation.

Synthesis, in vitro and in vivo characterization of novel 99mTc-‘4+1’-labeled 5-nitroimidazole derivatives as potential agents for imaging hypoxia - Corrected Proof

2012-04-25

Abstract: The evaluation of oxygenation status of solid tumors is an important field of radiopharmaceutical research. With the aim to develop new potential 99mTc-radiopharmaceuticals for imaging hypoxia, we have synthesized two novel isocyanide derivatives of metronidazole, which has demonstrated high affinity for hypoxic tumors in vitro and in vivo.Methods: Metronidazole derivatives 4-isocyano-N-[2-(2-methyl-5-nitro-1H-imidazol-1-yl)ethyl]butanamide (M1) and 1-(4-isocyanobutanoyl)-4-[2-(2-methyl-5-nitro-1H-imidazol-1-yl)ethyl]piperazine (M2) were synthesized, and labeling was performed through preparation of their corresponding 99mTc-(4+1) complexes, 99mTc-NS3M1 and 99mTc-NS3M2. The structure of the technetium complexes was corroborated by preparation and characterization of the corresponding rhenium complexes. We have studied the main physicochemical properties (stability, lipophilicity and plasma protein binding). Biological behavior in HCT-15 cells both in oxia and in hypoxia was assessed. Biodistribution in normal mice and in animals bearing induced 3LL Lewis murine lung carcinoma was also studied.Results: Metronidazole derivatives were successfully synthesized. Labeling with high radiochemical purity was achieved for both ligands. 99mTc complexes were stable in labeling milieu and human plasma. However, presence of the piperazine linker in M2 resulted in higher lipophilicity and protein binding. Although cell uptake in hypoxic conditions was observed for both radiotracers, 99mTc-NS3M2 biodistribution was considered unsuitable for a potential radiopharmaceutical due to high liver uptake and poor blood clearance. However, 99mTc-NS3M1 demonstrated a very favorable in vivo profile both in normal mice and in mice bearing induced tumors.Conclusion: Selective uptake and retention in tumor together with favorable tumor/muscle ratio make 99mTc-NS3M1 a promising candidate for further evaluation as potential hypoxia imaging agent in tumors.

Understanding the in vivo uptake kinetics of a phosphatidylethanolamine-binding agent 99mTc-Duramycin - Corrected Proof

Said Audi, Zhixin Li, Joseph Capacete, Yu Liu, Wei Fang, Laura G. Shu, Ming Zhao — 2012-04-25

Abstract: Introduction: 99mTc-Duramycin is a peptide-based molecular probe that binds specifically to phosphatidylethanolamine (PE). The goal was to characterize the kinetics of molecular interactions between 99mTc-Duramycin and the target tissue.Methods: High level of accessible PE is induced in cardiac tissues by myocardial ischemia (30 min) and reperfusion (120 min) in Sprague–Dawley rats. Target binding and biodistribution of 99mTc-duramycin were captured using SPECT/CT. To quantify the binding kinetics, the presence of radioactivity in ischemic versus normal cardiac tissues was measured by gamma counting at 3, 10, 20, 60 and 180 min after injection. A partially inactivated form of 99mTc-Duramycin was analyzed in the same fashion. A compartment model was developed to quantify the uptake kinetics of 99mTc-Duramycin in normal and ischemic myocardial tissue.Results: 99mTc-duramycin binds avidly to the damaged tissue with a high target-to-background radio. Compartment modeling shows that accessibility of binding sites in myocardial tissue to 99mTc-Duramycin is not a limiting factor and the rate constant of target binding in the target tissue is at 2.2 ml/nmol/min/g. The number of available binding sites for 99mTc-Duramycin in ischemic myocardium was estimated at 0.14 nmol/g. Covalent modification of D15 resulted in a 9-fold reduction in binding affinity.Conclusion: 99mTc-Duramycin accumulates avidly in target tissues in a PE-dependent fashion. Model results reflect an efficient uptake mechanism, consistent with the low molecular weight of the radiopharmaceutical and the relatively high density of available binding sites. These data help better define the imaging utilities of 99mTc-Duramycin as a novel PE-binding agent.

Preliminary studies of 99mTc-memantine derivatives for NMDA receptor imaging - Corrected Proof

2012-04-19

Abstract: Introduction: Novel technetium-labeled ligands, 99mTc-NCAM and 99mTc-NHAM were developed from the N-methyl-d-aspartate (NMDA) receptor agonist memantine as a lead compound by coupling with N2S2. This study evaluated the binding affinity and specificity of the ligands for the NMDA receptor.Methods: Ligand biodistribution and uptake specificity in the brain were investigated in mice. Binding affinity and specificity were determined by radioligand receptor binding assay. Three antagonists were used for competitive binding analysis. In addition, uptake of the complexes into SH-SY5Y nerve cells was evaluated.Results: The radiochemical purity of 99mTc-labeled ligands was more than 95%. Analysis of brain regional uptake showed higher concentration in the frontal lobe and specific uptake in the hippocampus. 99mTc-NCAM reached a higher target to nontarget ratio than 99mTc-NHAM. The results indicated that 99mTc-NCAM bound to a single site on the NMDA receptor with a Kd of 701.21 nmol/l and a Bmax of 62.47 nmol/mg. Specific inhibitors of the NMDA receptor, ketamine and dizocilpine, but not the dopamine D2 and 5HT1A receptor partial agonist aripiprazole, inhibited specific binding of 99mTc-NCAM to the NMDA receptor. Cell physiology experiments showed that NCAM can increase the viability of SH-SY5Y cells after glutamate-induced injury.Conclusions: The new radioligand 99mTc-NCAM has good affinity for and specific binding to the NMDA receptor, and easily crosses the blood–brain barrier; suggesting that it might be a potentially useful tracer for NMDA receptor expression.

A method to predict response of cell populations to cocktails of chemotherapeutics and radiopharmaceuticals: Validation with daunomycin, doxorubicin, and the alpha particle emitter 210Po - Corrected Proof

John M. Akudugu, Roger W. Howell — 2012-04-16

Abstract: There is considerable interest in the use of α-emitting radionuclides in radioimmunotherapy. However, the high toxicity of α-emitting radionuclides often does not permit administration of high activities for fear of normal tissue toxicity. Accordingly, targeting procedures need to be optimized for improved tumor control and minimized normal tissue toxicity. To guide design of effective cocktails of α-emitting radiopharmaceuticals and chemotherapy drugs, approaches that can predict biological response of a cell population on a cell-by-cell basis are needed.Methods: Cells were concomitantly treated with the α-particle emitting radiochemical 210Po-citrate and daunomycin, or with 210Po-citrate and doxorubicin. The responses of the treated cell populations were measured with a colony forming assay. The nonuniform cellular incorporation of the radiochemical and drugs was determined simultaneously on a cell-by-cell basis using flow cytometry. Monte Carlo methods were used to simulate cell survival on the basis of individual cell incorporation of each cytotoxic agent and validated by direct comparison with the experimental clonogenic cell survival.Results: Both daunomycin and doxorubicin enhanced the toxicity of the α-particles with a magnitude greater than expected based on single-agent toxicities. Cell survival obtained by Monte Carlo simulation was in good agreement with clonogenic cell survival for the combination treatments.Conclusion: Flow cytometry assisted Monte Carlo simulations can be used to predict toxicity of cocktails of α-emitting radiopharmaceuticals and chemotherapy drugs in a manner that takes into account the effects of nonuniform distributions of agents within cell populations.

In vitro characterization of [18F]-florbetaben, an Aβ imaging radiotracer - Corrected Proof

2012-04-13

Abstract: Purpose: Amyloid-β (Aβ) plaques are a major pathological hallmark of Alzheimer's disease (AD). The noninvasive detection of Aβ plaques may increase the accuracy of clinical diagnosis as well as monitor therapeutic interventions. While [11C]-PiB is the most widely used Aβ positron emission tomography (PET) radiotracer, due to the short half-life of 11C (20 min), its application is limited to centers with an on-site cyclotron and 11C radiochemistry expertise. Therefore, novel [18F] (half-life 110 min)-labeled Aβ PET tracers have been developed. We have demonstrated that [18F]-florbetaben-PET can differentiate individuals diagnosed with AD from healthy elderly, Parkinson's disease and frontotemporal lobe dementia (FTLD-tau) patients. While [18F]-florbetaben-PET retention matched the reported postmortem distribution of Aβ plaques, the nature of [18F]-florbetaben binding to other pathological lesions comprising misfolded proteins needs further assessment. The objective of this study was to determine whether Florbetaben selectively binds to Aβ plaques in postmortem tissue specimens containing mixed pathological hallmarks (i.e., tau and α-synuclein aggregates).Method: Human AD, FTLD-tau and dementia with Lewy bodies (DLB) brain sections were analyzed by [18F]-florbetaben autoradiography and [3H]-florbetaben high-resolution emulsion autoradiography and [19F]-florbetaben fluorescence microscopy.Results: Both autoradiographical analyses demonstrated that Florbetaben exclusively bound Aβ plaques in AD brain sections at low nanomolar concentrations. Furthermore, at concentrations thousand-folds higher than those during a PET scan, [19F]-florbetaben did not bind to α-synuclein or tau aggregates in DLB and FTLD-tau brain sections, respectively. Detection of [19F]-florbetaben staining by fluorescence microscopy in several AD brain regions demonstrated that Florbetaben identified Aβ plaques in all brain regions examined.Conclusion: This study provides further evidence that [18F]-florbetaben-PET is a highly selective radiotracer to assess Aβ plaque deposition in the brain.

Microwave-assisted one-pot radiosynthesis of 2′-deoxy-2′-[18F]fluoro-5-methyl-1-β-d-arabinofuranosyluracil ([18F]-FMAU) - Corrected Proof

Kai Chen, Zibo Li, Peter S. Conti — 2012-04-13

Abstract: Objectives: [18F]-FMAU is a PET tracer being evaluated for imaging cell proliferation. Current multi-step procedures of [18F]-FMAU synthesis are time-consuming, resulting in low radiochemical yield and inconvenient applications for the clinic. We have previously reported the use of Friedel-Crafts catalysts for an improved synthesis of [18F]-FMAU. In this study, we investigated the efficiency of microwave-assisted radiosynthesis of [18F]-FMAU in comparison with conventional thermal conditions.Methods: A simplified one-pot synthesis of [18F]-FMAU was developed under microwave conditions. Various reaction times, temperatures, and microwave powers were systematically explored to optimize the coupling reaction of 2-deoxy-2-[18F]fluoro-1,3,5-tri-O-benzoyl-d-arabinofuranose ([18F]-sugar) and bis-2,4-(trimethylsilyloxy)-5-methyluracil (silylated uracil) in the presence of a Friedel-Crafts catalyst, trimethylsilyl trifluoromethanesulfonate (TMSOTf).Results: Microwave significantly enhanced the coupling efficiency of [18F]-sugar and silylated uracil by reducing the reaction time to 10 min (6-fold reduction as compared to conventional heating) at 95 °C. Base hydrolysis followed by high-performance liquid chromatography purification produced the desired [18F]-FMAU. The overall radiochemical yield was 20±4% (decay corrected, n=3). Radiochemical purity was >99% and specific activity was >400 mCi/μmol. The α/β anomer ratio was 1:2. The radiosynthesis time was about 90 min from the end of bombardment.Conclusions: A reliable microwave-assisted approach has been developed for routine synthesis of [18F]-FMAU. The new approach affords a simplified process with shorter synthesis time and higher radiochemical yield as compared to conventional heating. A fully automated microwave-assisted synthesis of [18F]-FMAU can be readily achieved under new reaction conditions.

A comparison of in vitro methods for determining the membrane receptor expression in cell lines - Corrected Proof

Zbynek Novy, Pavel Barta, Jana Mandikova, Milan Laznicek, Frantisek Trejtnar — 2012-04-12

Abstract: Introduction: Determining the number of expressed receptors per cell (NRPC) in cell lines is an important prerequisite for many experimental procedures in biomedical research. This paper focuses on the comparison of a newly developed method of determining NRPC — the Kinetic extrapolation method (KEX) — with the standard saturation method. These two methods, both based on radiolabeled ligand–receptor binding, were compared with the data on receptor expression found using quantified western blotting.Methods: Four cell lines with different expressions of epidermal growth factor receptor (EGFR) were chosen for the experiment: A431, HaCaT, HCT116 and HepG2. Two radiolabeled monoclonal antibodies specific for EGFR were used as ligands: [131I]-cetuximab and [131I]-panitumumab. The classic manual technique based on the saturation of cell receptors was performed on cells seeded in 24-well plates. The KEX method uses the LigandTracer, a special instrument which detects ligand retention in real time from seeded cells onto a rotating Petri dish. The western blot analysis was performed according to the routinely used procedure.Results: A very close accordance between the manual saturation technique and the KEX method was found in all four cell lines used. The NRPC in the cell lines follows the same order using both ligands: A431>HaCaT>HCT116≈HepG2. Similarly, consistent data on EGFR expression in the studied cell lines were obtained using western blot analysis and the radiolabeled ligand binding assays.Conclusions: The KEX method could be as similarly useful for determining receptor expression as is the classic saturation method and western blotting.

Re-evaluation of in vivo selectivity of [11C]SA4503 to σ1 receptors in the brain: Contributions of emopamil binding protein - Corrected Proof

Jun Toyohara, Muneyuki Sakata, Kiichi Ishiwata — 2012-04-12

Abstract: Introduction: Carbon-11-labeled 1-[2-(3,4-dimethoxyphenyl)ethyl]-4-(3-phenylpropyl)piperazine ([11C]SA4503) was shown to be a promising PET ligand for mapping σ1 receptors, and was applied to human subjects. However, an in vitro study indicated that SA4503 also binds to the emopamil binding protein (EBP), vertebral Δ8-Δ7 sterol isomerase. To our knowledge, no information is available about the possibility of [11C]SA4503 binding to EBP in the brain in vivo.Methods: To confirm the selectivity of [11C]SA4503, we carried out an in vivo blocking experiment using high-affinity EBP and σ1 selective blocker.Results: The brain uptake of [11C]SA4503 was dose-dependently decreased by SA4503 and the high-affinity σ1 blockers haloperidol, ifenprodil, and trifluperidol. On the other hand, the effects of the high-affinity EBP blockers tamoxifen and trifluoperazine were negligible.Conclusions: Our results confirmed the σ1-selective binding of [11C]SA4503 in the brain.

Comparison of 3′-deoxy-3′-[18F]fluorothymidine PET and O-(2-[18F]fluoroethyl)-L-tyrosine PET in patients with newly diagnosed glioma - Corrected Proof

Su Young Jeong, Sang Moo Lim — 2012-04-09

Abstract: Purpose: The purpose of this prospective study was to clarify the value of FLT PET and FET PET for the noninvasive grading and prognosis of newly diagnosed gliomas.Materials and methods: Twenty patients with newly diagnosed gliomas were investigated with FLT and FET PET before surgery. FLT and FET uptakes were assessed by the maximum standardized uptake (SUVmax) of tumor, and the ratio to uptake in the normal brain parenchyma (TNR). All tumors were graded by WHO system.Results: FLT PET detected all 17 high-grade gliomas (HGG) and did not detect all 3 low-grade gliomas (LGG). FET PET detected all 20 HGG and LGG regardless of grading. The average FLT SUVmax in HGG and LGG was 1.51±0.72 and 0.30±0.07, and the average FLT TNR in HGG and LGG was 5.52±3.09 and 1.12±0.14, respectively. The differences of FLT SUVmax and TNR between HGG and LGG were statistically significant (p=0.0069, p=0.0070). The average FET SUVmax in HGG and LGG was 2.68±0.86 and 1.36±0.15, and the average FET TNR in HGG and LGG was 2.31±0.73 and 1.27±0.12, respectively. The differences of FET SUVmax and TNR between HGG and LGG were statistically significant (p=0.0129, p=0.0095).Conclusions: FET PET has higher sensitivity in detection of gliomas rather than FLT PET, but it seems that FLT PET is better than FET PET for noninvasive grading and predicting prognosis of newly diagnosed gliomas, considering high contrast of FLT and overlap of FET uptakes between HGG and LGG.

[123I]Epidepride neuroimaging of dopamine D2/D3 receptor in chronic MK-801-induced rat schizophrenia model - Corrected Proof

2012-03-30

Abstract: Purpose: [123I]Epidepride is a radio-tracer with very high affinity for dopamine D2/D3 receptors in brain. The importance of alteration in dopamine D2/D3 receptor binding condition has been wildly verified in schizophrenia. In the present study we set up a rat schizophrenia model by chronic injection of a non-competitive NMDA receptor antagonist, MK-801, to examine if [123I]epidepride could be used to evaluate the alterations of dopamine D2/D3 receptor binding condition in specific brain regions.Method: Rats were given repeated injection of MK-801 (dissolved in saline, 0.3mg/kg) or saline for 1month. Afterwards, total distance traveled (cm) and social interaction changes were recorded. Radiochemical purity of [123I]epidepride was analyzed by Radio-Thin-Layer Chromatography (chloroform: methanol, 9:1, v/v) and [123I]epidepride neuroimages were obtained by ex vivo autoradiography and small animal SPECT/CT. Data obtained were then analyzed to determine the changes of specific binding ratio.Result: Chronic MK-801 treatment for a month caused significantly increased local motor activity and induced an inhibition of social interaction. As shown in [123I]epidepride ex vivo autoradiographs, MK-801 induced a decrease of specific binding ratio in the striatum (24.01%), hypothalamus (35.43%), midbrain (41.73%) and substantia nigra (37.93%). In addition, [123I]epidepride small animal SPECT/CT neuroimaging was performed in the striatum and midbrain. There were statistically significant decreases in specific binding ratio in both the striatum (P<.01) and midbrain (P<.05) between the saline and MK-801 group.Conclusion: These results suggest that [123I]epidepride is a useful radio-tracer to reveal the alterations of dopamine D2/D3 receptor binding in a rat schizophrenia model and is also helpful to evaluate therapeutic effects of schizophrenia in the future.

Preliminary studies on 177Lu-labeled sodium pyrophosphate (177Lu-PYP) as a potential bone-seeking radiopharmaceutical for bone pain palliation - Corrected Proof

Imtiaz Ahmed Abbasi — 2012-03-30

Abstract: Objective: 99mTc-Sn-PYP (Technetium-99m labeled tin pyrophosphate) has been widely used as a radiopharmaceutical for bone scanning as well as in nuclear cardiology. It is also found in the body in trace amounts. 177Lu is presently considered as an excellent radionuclide for developing bone pain palliation agents. PYP is an analogue of MDP and MDP has been labeled with 177Lu. No study on preparing a complex of 177Lu with PYP has been reported yet. Based on these facts, it was hypothesized that a bone-seeking 177Lu-PYP (Lutetium-177 labeled Pyrophosphate) radiopharmaceutical could be developed as an agent for palliative radiotherapy of bone pain due to skeletal metastases.Methods: 177Lu was produced by irradiating lutetium foil (11 mg) natural target at a flux ∼1.0×1014n/cm2/s for 12 h in the swimming pool type reactor. 177Lu in the form of 177LuCl3 was labeled with PYP. The radiochemical purity and labeling efficiencies were determined by paper chromatography. Labeling of 177Lu with PYP was optimized and a labeled sample was subjected to HPLC analysis. To determine the charge on the 177Lu-PYP complex, radio-electrophoresis was conducted for 1 h under a voltage of 300 V and 45 mA current using 0.025 M phosphate buffer (pH 6.9). Bioevaluation studies with rabbit under γ-camera were also performed to verify the skeletal uptake.Results: The quality control using paper radio-chromatography has shown >99% radiochemical purity of 177Lu-PYP complex. Radio-chromatography also showed maximum labeling at ligand/metal ratio=60:1. HPLC analysis showed 1.42±0.01 min retention time of 177Lu-PYP complex. No decrease in labeling was observed at higher temperatures. Gamma-camera images of 177Lu-PYP in normal rabbit at 24 h post injection also showed high skeletal uptake.Conclusion: The study demonstrated that sodium pyrophosphate could be labeled with 177Lu with high radiochemical yields (>99%). Negatively charged 177Lu-PYP complex retained stability for a day and at high temperatures too. Gamma-camera images of 177Lu-PYP in normal rabbit at 24 h post injection showed high skeletal uptake, suggesting that it may be useful as a bone-pain palliation agent for the treatment of bone metastases.

A preclinical investigation of the saturation and dosimetry of 153Sm-DOTMP as a bone-seeking radiopharmaceutical - Corrected Proof

2012-03-29

Abstract: Introduction: The therapeutic potential of the bone-seeking radiopharmaceutical 153Sm-labeled 1,4,7,10-tetraazacyclododecanetetramethylenephosphonic acid (153Sm-DOTMP) was assessed by measuring its dosage-dependent skeletal uptake at two chelant-to-metal ratios and its source organ residence times at a chelant-to-metal ratio of 1.5:1. A similar agent, 153Sm-labeled ethylenediaminetetramethylenephosphonic acid (153Sm-EDTMP), has been reported to exhibit dosage-limiting skeletal saturation.Methods: Sm-DOTMP was prepared with tracer activity of 153Sm and sufficient stable, unenriched Sm to simulate different activities. Cohorts of seven 280-g Sprague–Dawley rats were administered the equivalent of 296, 592, 888, 1184 and 1480 MBq (8, 16, 24, 32 and 40 mCi) at a fixed chelant-to-metal ratio of 1.5:1 and euthanized 3 h after administration. Cohorts of three 128-g Sprague–Dawley rats were administered equivalent dosages of 10.4, 592 and 888 (0.28, 16 and 32 mCi) at a fixed chelant-to-metal ratio of 270:1 and euthanized 2 h after administration. A simulated activity of 1480 MBq (40 mCi) at a chelant-to-metal ratio of 1.5:1 was administered to cohorts of seven rats that were euthanized at 2, 4, 24 or 48 h postadministration. The heart, lungs, liver, spleen, kidneys, small intestine, large intestine, urinary bladder, muscle and a femur were excised, weighed and counted. The data were analyzed to determine skeletal uptake and source organ residence times.Results: No statistically significant skeletal saturation was observed up to human-equivalent dosages of 370 GBq (10 Ci) at a chelant-to-metal ratio of 1.5:1, but the skeletal uptake dropped by 40% over the range of dosages at a chelant-to-metal ratio of 270:1. At a chelant-to-metal ratio of 1.5:1, the preferred ratio, the skeletal uptake fraction in rats was 0.408 (95% confidence interval 0.396–0.419) with an effective half-life of 47.3 h (95% confidence interval 42.3–53.7; the physical half-life of 153Sm is 46.3 h). Extrapolating to an adult human model, 52.9 GBq (1.43 Ci) of 153Sm-DOTMP would deliver 40 Gy to the red marrow.Conclusion: 153Sm-DOTMP has dosimetry equivalent to that of 153Sm-EDTMP at low dosages, yet with no skeletal saturation at higher administered activities.

In vivo evaluation of [18F]FEAnGA-Me: a PET tracer for imaging β-glucuronidase (β-GUS) activity in a tumor/inflammation rodent model - Corrected Proof

2012-03-27

Abstract: Introduction: The PET tracer, 1-O-(4-(2-fluoroethyl-carbamoyloxymethyl)-2-nitrophenyl)-O-β-d-glucopyronuronate ([18F]FEAnGA), was recently developed for PET imaging of extracellular β-glucuronidase (β-GUS). However, [18F]FEAnGA exhibited rapid renal clearance, which resulted in a relatively low tracer uptake in the tumor. To improve the pharmacokinetics of [18F]FEAnGA, we developed its more lipophilic methyl ester analog, [18F]FEAnGA-Me.Methods: [18F]FEAnGA-Me was obtained by alkylation of the O-protected glucuronide methyl ester precursor with [18F]-fluoroethylamine ([18F]FEA), followed by removal of the acetate protecting groups with NaOMe/MeOH. The PET tracer was evaluated by in vitro and in vivo studies.Results: [18F]FEAnGA-Me was obtained in 5%–10% overall radiochemical yield. It is 10-fold less hydrophilic than [18F]FEAnGA and it is stable in PBS and in the presence of β-GUS for 1 h. However, in the presence of esterase or plasma [18F]FEAnGA-Me is converted to [18F]FEAnGA, and subsequently converted to [18F]FEA by β-GUS. MicroPET studies in Wistar rats bearing a C6 glioma and a sterile inflammation showed similar uptake in tumors after injection of either [18F]FEAnGA-Me or [18F]FEAnGA. Both tracers had a rapid two-phase clearance of total plasma radioactivity with a half-life of 1 and 8 min. The [18F]FEAnGA fraction generated from [18F]FEAnGA-Me by in vivo hydrolysis had a circulation half-life of 1 and 11 min in plasma. Similar distribution volume in the viable part of the tumor was found after injection of either [18F]FEAnGA-Me or [18F]FEAnGA.Conclusion: The imaging properties of [18F]FEAnGA-Me were not significantly better than those of [18F]FEAnGA. Therefore, other strategies should be applied in order to improve the kinetics of these tracers.

Nephrotoxicity profiles and threshold dose values for [177Lu]-DOTATATE in nude mice - Corrected Proof

2012-03-27

Abstract: Introduction: In peptide receptor radionuclide therapy for neuroendocrine tumors the main dose-limiting tissue is found in the kidneys because of tubular reabsorption and retention of radioactivity. The aim of this study was to quantify late effects in renal cortex of nude mice exposed to high amounts of [177Lu]-DOTA-Tyr3-octreotate ([177Lu]-DOTATATE), and to determine whether a threshold dose value exists for these findings.Methods: Nude mice were exposed to 90, 120 or 150 MBq of [177Lu]-DOTATATE. Renal toxicity was evaluated up to 6 months after injection. Blood samples were collected to examine renal functional markers, and after sacrifice at 6 months changes in renal morphology were explored. Tissue damage was estimated by quantifying the relative area of the different subunits in the renal cortex using point counting. Additional morphological signs of radiation damage were also noted. The absorbed doses to the kidneys were estimated by previously determined kidney pharmacokinetics and Monte Carlo simulations for different assumptions regarding the activity distribution.Results: Increased serum creatinine and urea values indicated long-term renal toxicity. The tissue area occupied by proximal tubules decreased with increasing doses of [177Lu]-DOTATATE, whereas the other subunits in cortex slightly increased. The mean absorbed dose in the renal cortex for [177Lu]-DOTATATE was estimated to be 35–58 Gy for the different groups of animals. A dose–response relationship was observed for proximal tubular damage, and a threshold dose value of 24 Gy (BED 37 Gy) was determined.Conclusions: Selective morphological changes in kidney cortex of nude mice were quantified and appeared in a dose dependent manner after injection of high amounts of [177Lu]-DOTATATE.

Novel synthesis and preclinical evaluation of folic acid derivatives labeled with 18F-[FDG] for PET imaging of folate receptor-positive tumors - Corrected Proof

I. Al Jammaz, B. Al-Otaibi, S. Amer, N. Al-Hokbany, S. Okarvi — 2012-03-27

Abstract: There is a need to develop more potent radiofluorinated folic acid conjugates for a better visualization of folate receptors that overexpress on many human cancers. Due to the clinical importance of [18F]-fluoro-2-deoxy-d-glucose ([18F]-FDG) and its availability in almost every positron-emission tomography center, new radiofluorinated [18F]-FDG-folate and methotrexate conjugates ([18F]-5 and [18F]-8) were synthesized using [18F]-FDG as a prosthetic group. In a convenient and simple one-step radiosynthesis, [18F]-5 and [18F]-8 conjugates were prepared in high radiochemical yields (>80%) with total synthesis time of almost 20 min, and radiochemical purities were found to be greater than 98% without high-performance liquid chromatography purification, which make these approaches amenable for automation. In vitro tests on KB cell line showed that a significant amount of the radioconjugates were associated with the cell fractions. In vivo characterization in normal Balb/c mice revealed rapid blood clearance of these radioconjugates with excretion predominantly by the urinary and hepatobiliary systems for [18F]-5 and [18F]-8 conjugates, respectively. Biodistribution studies in nude mice-bearing human KB cell line xenografts demonstrated significant tumor uptake and favorable kinetics profile for [18F]-5 over the other conjugate. The uptake in the tumors was blocked by the excess coinjection of cold folic acid, suggesting the receptor-mediated process. These results demonstrate that [18F]-5 may be useful as a molecular probe for detecting and staging of folate receptor-positive cancers, such as ovarian cancer and their metastasis, as well as monitoring tumor response to the treatment.

68Ga-NODAGA-RGD is a suitable substitute for 18F-Galacto-RGD and can be produced with high specific activity in a cGMP/GRP compliant automated process - Corrected Proof

2012-03-23

Abstract: Introduction: 18F-Galacto-cyclo(RGDfK) is a well investigated tracer for imaging of ανβ3 expression in vivo, but suffers from the drawback of a time consuming multistep synthesis that can hardly be established under GMP conditions. In this study, we present a direct comparison of the pharmacokinetic properties of this tracer with 68Ga-NODAGA-cyclo(RGDyK), in order to assess its potential as an alternative for 18F-Galacto-cyclo(RGDfK).Methods: 68Ga labeling of NODAGA-cyclo(RGDyK) was done in full automation using HEPES-buffered eluate of an SnO2 based 68Ga-generator. Using M21 (human melanoma) xenografted BALB/c nude mice, biodistribution studies and micro-PET scans were performed for both 18F-Galacto-cyclo(RGDfK) and 68Ga-NODAGA-cyclo(RGDyK), and for the latter, in vivo stability was assessed. IC50 was determined in a displacement assay on M21 cells against 125I-echistatin.Results: 68Ga-NODAGA-cyclo(RGDyK) was produced with high specific activity (routinely ca. 500 GBq/μmol) within 15 min. IC50 values are similar for both substances. Tracer uptake was similar in ανβ3 positive tumors (1.45%±0.11% ID/g and 1.35%±0.53% ID/g for 68Ga-NODAGA-RGD and 18F-Galacto-RGD, respectively) as well as for all other organs and tissues, with the exception of gall bladder and intestines, where 18F-Galacto-cyclo(RGDfK) uptake was significantly higher, which can be explained by the higher hydrophilicity of 68Ga-NODAGA-cyclo(RGDyK) (logP=−4.0 vs. −3.2 for 18F-Galacto-RGD). Only intact tracer was detected 30 min p.i. in organs and tumor; however, minor amounts of metabolites were found in the urine (6% of total urine activity).Conclusion: 68Ga-labeling of NODAGA-RGD can be performed rapidly and efficiently within 15 min in a GMP compliant process. Similar preclinical results were obtained in comparison with 18F-Galacto-RGD. Therefore, 68Ga-NODAGA-cyclo(RGDyK) is a suitable replacement for 18F-Galacto-cyclo(RGDfK).

Tryptophan metabolism in breast cancers: molecular imaging and immunohistochemistry studies - Corrected Proof

2012-03-23

Abstract: Introduction: Tryptophan oxidation via the kynurenine pathway is an important mechanism of tumoral immunoresistance. Increased tryptophan metabolism via the serotonin pathway has been linked to malignant progression in breast cancer. In this study, we combined quantitative positron emission tomography (PET) with tumor immunohistochemistry to analyze tryptophan transport and metabolism in breast cancer.Methods: Dynamic α-[11C]methyl-l-tryptophan (AMT) PET was performed in nine women with stage II–IV breast cancer. PET tracer kinetic modeling was performed in all tumors. Expression of L-type amino acid transporter 1 (LAT1), indoleamine 2,3-dioxygenase (IDO; the initial and rate-limiting enzyme of the kynurenine pathway) and tryptophan hydroxylase 1 (TPH1; the initial enzyme of the serotonin pathway) was assessed by immunostaining of resected tumor specimens.Results: Tumor AMT uptake peaked at 5–20 min postinjection in seven tumors; the other two cases showed protracted tracer accumulation. Tumor standardized uptake values (SUVs) varied widely (2.6–9.8) and showed a strong positive correlation with volume of distribution values derived from kinetic analysis (P<.01). Invasive ductal carcinomas (n=6) showed particularly high AMT SUVs (range, 4.7–9.8). Moderate to strong immunostaining for LAT1, IDO and TPH1 was detected in most tumor cells.Conclusions: Breast cancers show differential tryptophan kinetics on dynamic PET. SUVs measured 5–20 min postinjection reflect reasonably the tracer's volume of distribution. Further studies are warranted to determine if in vivo AMT accumulation in these tumors is related to tryptophan metabolism via the kynurenine and serotonin pathways.

In vivo and in vitro evaluation of Cy5.5 conjugated epidermal growth factor receptor binding peptide - Corrected Proof

2012-03-15

Abstract: The epidermal growth factor receptor (EGFR) is a tyrosine kinase receptor and plays an important role in carcinogenesis. In this study, the epidermal growth factor receptor binding peptide (EGBP) was identified using a phage display method and evaluated in U87MG cells in order to investigate the possibility to target the EGFR using an optical imaging system. Cyanine dye 5.5 (Cy5.5) was conjugated with EGBP-GGG-SC, EGBP-AOC-SC, and EGBP-AM2BA-SC. Cellular binding study of EGBP-Linker-Cy5.5 conjugates or 125I-EGBP-Linker compounds was performed in U87MG cells. Optical imaging studies were performed in U87MG bearing mice. Three of seven clones from the 12-mer peptide library showed a specific binding affinity to rhEGFR, and they encoded the same 12 amino acid peptide sequence, FPMFNHWEQWPP. Confocal images show that the fluorescent signal of EGBP-Linker-Cy5.5 conjugates was decreased in the order: EGBP-AOC-Cy5.5≫EGBP-AM2BA-Cy5.5>EGBP-GGG-Cy5.5. EGBP-AOC-Cy5.5 appeared in cell cytoplasm and surface, and it was inhibited by free EGBP apparently. The cellular binding of EGBP-AOC-Cy5.5 exhibited a higher average radiance value than EGBP-GGG-Cy5.5 and EGBP-AM2BA-Cy5.5. Among various 125I-EGBP-Linker compounds, EGBP-GGG showed a higher binding than other compounds. However, uptake of 125I-EGBP-AOC was clearly inhibited by free EGBP in inhibition study. In an in vivo study, the fluorescent signal of EGBP-AOC-Cy5.5 treated mouse was mainly detected in the tumor and kidney. Among the three derivatives, EGBP-AOC-Cy5.5 was the optimized optical imaging agent for U87MG EGFR positive tumors in the animal model. This study demonstrated the EGBP-Linker-Cy5.5 conjugates may be useful as a potential EGFR target optical probe.

Whole-body distribution and radiation dosimetry of [11C]telmisartan as a biomarker for hepatic organic anion transporting polypeptide (OATP) 1B3 - Corrected Proof

2012-03-15

Abstract: Introduction: Telmisartan, a nonpeptide angiotensin II AT1 receptor antagonist used as an antihypertensive drug, is specifically taken up by the liver through the OATP1B3. PET imaging with [11C]telmisartan is expected to provide information about the whole body pharmacokinetics of telmisartan as well as its transport property by OATP1B3. The purpose of the study was to determine the biodistribution and radiation dosimetry of [11C]telmisartan in humans.Methods: Biodistribution of [11C]telmisartan was measured in three rats and six healthy male human volunteers. In the rat study, a dynamic emission scan was performed for 90 min. In the human study, dynamic whole-body PET images were acquired after intravenous injection of [11C]telmisartan. ROIs were defined for source organs on the PET images to measure time-course of [11C]telmisartan uptake as percentage injected dose and the number of disintegration for each organ. Radiation dosimetry was calculated with OLINDA/EXM.Results: In the rat study, most radioactivity was rapidly taken up by the liver and part of it was excreted into the biliary tract and intestine. Extrapolating from the rat data, the effective dose for the adult human being was estimated to be 3.65±0.01 microSv/MBq (n=3). In the human study, most of the tracer was taken up by the liver as well, although not as rapidly as in the rat. The activity in the gall bladder and intestine increased gradually. The effective dose for the adult human being was 4.24±0.09 microSv/MBq (n=6).Conclusions: [11C]Telmisartan is a safe PET tracer with a dosimetry profile comparable to other common 11C PET tracers.

Evaluation of the angiogenesis inhibitor KR-31831 in SKOV-3 tumor-bearing mice using 64Cu-DOTA-VEGF121 and microPET - Corrected Proof

2012-03-14

Abstract: KR-31831 ((2R,3R,4S)-6-amino-4-[N-(4-chloropheyl)-N-(1H-imidazol-2ylmethyl)amino]-3-hydroxyl-2-methyl-2-dimethoxymethyl-3,4-dihydro-2H-1-benzopyran), an angiogenesis inhibitor, was evaluated in tumor-bearing mice using molecular imaging technology. Pre-treatment microPET images were acquired on SKOV-3 cell-implanted nude mice after injection with 64Cu-DOTA-VEGF121. KR-31831 (50 mg/kg) was then injected intraperitoneally into the treatment group (n=3), while injection vehicle was injected into the control (n=4) and blocking (n=3) groups. After injections occurred daily for 28 days, all groups of mice underwent post-treatment microPET imaging after injection with 64Cu-DOTA-VEGF121. The post-treatment images showed high tumor uptake in the control group and reduced tumor uptake in both the blocking and treatment groups. ROI analysis of the tumor images revealed 6.25%±1.18% ID/g at 1 h, 6.55%±0.69% ID/g at 2 h, and 4.68%±0.63% ID/g at 16 h in the control group; 3.87%±0.45% ID/g at 1 h, 4.50%±0.44% ID/g at 2 h, and 3.63%±0.25% ID/g at 16 h in the blocking group; and 4.03%±0.74% ID/g at 1 h, 4.37%±0.67% ID/g at 2 h, and 3.83%±0.90% ID/g at 16 h in the treatment group. Biodistribution obtained after the post-treatment microPET imaging also demonstrated high tumor uptake (3.74%±0.27% ID/g) in the control group and reduced uptakes in both the blocking group (2.69%±0.73% ID/g, P<.05) and the treatment group (3.11%±0.25% ID/g, P<.05), which correlated well with microPET imaging data. Immunofluorescence analysis showed higher levels of VEGFR2 and CD31 expressions in tumor tissues of the control and blocking groups than in tumor tissues of the treatment group. These results suggest that the antiangiogenic activity of KR-31831 is mediated through VEGFR2 and microPET serves as a useful molecular imaging tool for evaluation of a newly developed angiogenesis inhibitor, KR-31831.

Repeatability of FDG quantification in tumor imaging: averaged SUVs are superior to SUVmax - Corrected Proof

2012-03-02

Abstract: Purpose: Reliable 18F-fluorodeoxyglucose (FDG) uptake quantification is crucial for cancer treatment monitoring. While interobserver variability has been found to be lower for a maximum standard uptake value (SUV)max than for an averaged SUV (SUVmean), the repeatability has not been investigated yet. In this study, we determined the repeatability of SUV values in two sequential measurements 5 min apart.Methods: Positron emission tomography data of malignant chest tumors were acquired dynamically during 45 min in 20 patients. SUV values were derived from the hottest (SUVmax), the mean of the 5 (SUV5) and 10 (SUV10) hottest voxels and the mean of a volume of interest (SUVmean). The repeatability of the SUV measurements was determined as the standard deviation of the difference between the values at 40 and 45 min and represented as Bland–Altman graphs.Results: The standard deviation of the difference between the two sequential scans for SUVmax, SUV5, SUV10 and SUVmean was 1.01, 0.53, 0.37 and 0.28.Conclusion: The repeatability of SUV is markedly increased by deriving the value from multiple voxels. Compared to SUVmax, the variability in SUV measurements is reduced by a factor of 2.7 (2.7=1.01/0.37) if 10 voxels are pooled.

High molecular mass radioimmunoconjugates are promising for intraperitoneal α-emitter immunotherapy due to prolonged retention in the peritoneum - Corrected Proof

2012-03-02

Abstract: Introduction: Therapeutic efficacy of intraperitoneal radioimmunotherapy is dependent on the time of retention of the radioimmunoconjugates within the peritoneal cavity. Therefore, the aim of this study was to investigate intraperitoneal retention of Fab, IgG and IgM radioimmunoconjugates.Methods: Female Balb/c mice were injected with 213Bi- or 111In-labeled IgM, IgG and recombinant Fab conjugates intraperitoneally or intravenously. At different time points after injection, whole body distribution of radionuclides was imaged using a gamma camera. Distribution of radionuclides in selected organs was determined via γ-counting after sacrifice. Biological half-lives of the conjugates were calculated from whole body activities.Results: After i.p. injection 213Bi-Fab rapidly accumulated in the kidneys indicative of glomerular filtration and reabsorption. Accumulation of 213Bi-IgG in the kidneys was significantly lower. 213Bi-IgM showed a striking accumulation in the liver 180 min after i.p. injection. 111In-IgG persisted in the circulation up to 72 h both after i.p. and i.v. injection. 111In-IgM showed a continuous accumulation in the liver. Moreover, 111In-IgM was significantly higher 24 h after i.v. injection than i.p. injection both in liver and spleen. These differences could be confirmed via scintigraphy. After injection of 111In-IgG differences in scintigraphic images between i.v. and i.p. were clearly visible only at 3 h. Biological half lives were 24 h, 45 h and 165 h for 111In-IgM, 111In-Fab and 111In-IgG, respectively.Conclusions: Retention of radioimmunoconjugates in the peritoneal cavity positively correlates with the molecular mass of the antibody. Therefore, IgM radioimmunoconjugates should be preferably used in radioimmunotherapy of free floating tumor cells and small tumor cell clusters in the ascites of the peritoneal cavity.

11C-5-hydroxytryptophan positron emission tomography after radiofrequency ablation of neuroendocrine tumor liver metastases - Corrected Proof

2012-03-02

Abstract: Aim: The aim was to assess the feasibility of 11C-5-hydroxy-tryptophan positron emission tomography (11C-5-HTP-PET) in the follow-up after radiofrequency ablation (RFA) of liver metastases from neuroendocrine tumors (NETs).Background: Contrast-enhanced computed tomography (CECT) and contrast-enhanced ultrasound (CEUS) are commonly used to evaluate the liver after RFA of NETs. In general, 11C-5-HTP-PET is more sensitive in the visualization of NETs, but no studies have investigated its role after RFA.Methods: Six consecutive patients with liver metastases from NETs were subjected to RFA treatment. All patients underwent baseline imaging before RFA and on two occasions (1–2 and 6–11 months) after RFA. The imaging consisted of 11C-5-HTP-PET, CEUS and CECT on all three occasions.Results: Thirty RFA areas were evaluated, and residual tumors (RTs) were depicted in eight areas (22%). 11C-5-HTP-PET depicted RTs after RFA with maximum sensitivity (100%) and specificity (100%), using radiological follow-up as the gold standard. 11C-5-HTP-PET detected five out of eight RTs earlier than CECT or CEUS. In general, the sensitivity of 11C-5-HTP-PET exceeded that of CECT and CEUS for early visualization of NET liver metastases.Conclusion: 11C-5-HTP-PET can be used in the follow-up after RFA for the purpose of detecting RT, and it provides additional information to CEUS and CECT by detecting new lesions.

Preparation and evaluation of bombesin peptide derivatives as potential tumor imaging agents: effects of structure and composition of amino acid sequence on in vitro and in vivo characteristics - Corrected Proof

Subhani M. Okarvi, Ibrahim A. Jammaz — 2012-03-02

Abstract: Objectives: Among the many clinically relevant peptide receptor systems, bombesin (BN) receptors have attracted enormous attraction due to their overexpression in various frequently occurring human tumors including breast and prostate, thus making such receptors promising targets with radiolabeled BN analogs. The present study describes the preparation and evaluation of a series of new BN derivatives as potential tumor imaging agents.Methods: Several new BN derivatives with the common structure MAG3-X-BN(1–14 or 6–14), where X=Asp or Asp-Asp, were synthesized by solid-phase peptide synthesis. S-benzoylmercaptoacetic acid was incorporated at the end of synthesis via manual conjugation to yield MAG3-BN conjugates. Radiolabeling with 99mTc was accomplished by ligand exchange method. The receptor-binding affinity assays were performed in MDA-MB-231, MCF-7, T47-D and PC-3 cancer cell lines. In vivo biodistribution and clearance kinetics were assessed in Balb/c mice, and tumor targeting efficacy was determined in nude mice bearing breast tumor xenografts.Results: The peptides were prepared conveniently and radiolabeled efficiently with 99mTc (up to 95% labeling efficiency). In vitro cell binding assays demonstrated high affinity (values in the nanomolar range) of 99mTc peptides towards breast and prostate cancer cell lines. In addition, the radioconjugates displayed significant internalization (values ranged between 19% and 35%) in tumor cells. In vivo biodistribution and biokinetics are characterized by efficient clearance from the blood and variable degrees of excretion through the renal pathway. In vivo tumor targeting studies displayed variable uptake capacity of different BN derivatives, underlining the influence of specific amino acid sequence on tumor targeting profiles. Tumor uptake was always higher than the radioactivity in the blood and muscle, with good tumor retention and good tumor-to-blood and tumor-to-muscle ratios, indicating the potential of these agents for targeting tumors in vivo.Conclusions: The combination of favorable in vitro and in vivo properties may render these BN peptides as potential candidates for targeting BN/GRP receptor-positive tumors. They deserve further evaluation to determine their real strength. The present data indeed provide useful information regarding peptide structure–pharmacologic activity relationship, which might be useful in designing and developing new BN-like peptides for efficient targeting of tumors in vivo.

RGD conjugates of the H2dedpa scaffold: synthesis, labeling and imaging with 68Ga - Corrected Proof

2012-03-02

Abstract: Introduction: The rekindled interest in the 68Ga generator as an attractive positron emission tomography generator system has led us and others to investigate novel chelate systems for 68Ga. We have previously reported our findings with the acyclic, rapidly coordinating chelate H2dedpa and its model derivatives.Methods: In this report, we describe the synthesis of the corresponding bifunctional chelate scaffolds (H2dp-bb-NCS and H2dp-N-NCS) as well as the radiolabeling properties, transferrin stability, binding to the target using in vitro cell models and in vivo behavior the corresponding conjugates with the αvβ3 targeting cyclic pentapeptide cRGDyK (monomeric H2RGD-1 and dimeric H2RGD-2).Results: The ability of the conjugated ligands to coordinate Ga isotopes within 10 min at room temperature at concentrations of 1 nmol was confirmed. Complex [67Ga(RGD-1)]+ was more stable (92% after 2 h) than [67Ga(RGD-2)]+ (73% after 2 h) in a transferrin challenge experiment. IC50 values for both conjugates (H2RGD-1 and H2RGD-2) and nonconjugated RGD were determined in a cell-based competitive binding assay with 125I-echistatin using U87MG cells, where enhanced specific binding was observed for the multivalent H2RGD-2 conjugate compared to the monovalent H2RGD-1 and nonconjugated cRGDyK. The U87MG cell line was also used to generate subcutaneous xenograft tumors on RAG2M mice, which were used to evaluate the in vivo properties of [68Ga(RGD-1)]+ and [68Ga(RGD-2)]+. After 2 h of dynamic imaging, both block and nonblock mice were sacrificed to collect select organs at the 2-h time point. Although the uptake is specific, as judged from the ratios of nonblock to block (2.36 with [67Ga(RGD-1)]+, 1.46 with [67Ga(RGD-2)]+), both conjugates display high uptake in blood.Conclusions: We have successfully synthesized and applied the first bifunctional versions of H2dedpa for conjugation to a targeting vector and subsequent imaging of the corresponding conjugates.

Synthesis of ApoSense compound [18F]2-(5-(dimethylamino)naphthalene-1-sulfonamido)-2-(fluoromethyl)butanoic acid ([18F]NST732) by nucleophilic ring opening of an aziridine precursor - Corrected Proof

2012-02-16

Abstract: Introduction: The small molecule 2-(5-(dimethylamino)naphthalene-1-sulfonamido)-2-(fluoromethyl)butanoic acid (NST732) is a member of the ApoSense family of compounds, capable of selective targeting, binding and accumulation within cells undergoing apoptotic cell death. It has application in molecular imaging and blood clotting particularly for monitoring antiapoptotic drug treatments. We are investigating a fluorine-18-radiolabeled analog of this compound for positron emission tomography studies.Methods: We prepared the tosylate precursor methyl 2-(5-(dimethylamino)naphthalene-1-sulfonamido)-2-(tosyloxymethyl)butanoate (4) to synthesize fluorine-18-labeled NST732. Fluorination reaction of the tosylate precursor in 1:1 acetonitrile:dimethylsulfoxide with tetrabutyl ammonium fluoride proceeds through an aziridine intermediate (4A) to afford two regioisomers: 2-(5-(dimethylamino)naphthalene-1-sulfonamido)-2-fluorobutanoate (5) and methyl 2-(5-(dimethylamino)naphthalene-1-sulfonamido)-2-(fluoromethyl)butanoate (6). Acid hydrolysis of the fluoromethylbutanoate (6) isomer produced NST732. As the fluorination reaction of the tosylate precursor proceeds through an aziridine intermediate (4A) and the fluorination conceivably could be done directly on the aziridine, we have separately prepared an aziridine precursor (4A). Fluorine-18 labeling of the aziridine precursor (4A) was performed with [18F]tetrabutyl ammonium fluoride to afford the same two regioisomers (5 and 6). The [18F]2-((5-dimethylamino)naphthalene-1-sulfonamido)methyl)-2-fluorobutanoic acid (NST732) was then obtained by the hydrolysis of corresponding [18F]-labeled ester (6) with 6 N hydrochloric acid.Results: Two regioisomers obtained from the fluorination reaction of aziridine were easily separated by high-performance liquid chromatography. The total radiochemical yield was 15%±3% (uncorrected, n=18) from the aziridine precursor in a 70-min synthesis time with a radiochemical purity >99%.Conclusion: Fluorine-18-labeled ApoSense compound [18F]NST732 is prepared in moderate yield by direct fluorination of an aziridine precursor.

Use of gated 13N-NH3 micro-PET to examine left ventricular function in rats - Corrected Proof

2012-02-16

Abstract: Introduction: Myocardial perfusion gating techniques offer the possibility of measurement of left ventricular end-systolic (ESV) and end-diastolic volume (EDV) and left ventricular ejection fraction (LVEF) in clinical and preclinical trials. The aim of this study was to evaluate left ventricular volumes (LVV) and LVEF with 13N-NH3 in comparison with the reference 18F-FDG in different rat models.Methods: In this study, 18 male Wistar rats, 12 control rats and 6 rats with myocardial infarction (MI) were imaged with micro-PET. The ratswere scanned with gated 13N-NH3 and 18F-FDG sequentially for the assessment of LVV and LVEF. A validated three-dimensional segmentation algorithm was used to calculate LVV and LVEF.Results: Mean LVEF measured with 13N-NH3 was 45.6±8.9 and 75.3±9.4%, mean ESV was 0.40±0.12 and 0.14±0.11 ml, and mean EDVwas 0.53±16 and 0.75±0.18 ml for MI and control rats, respectively. Moderate to good correlations were observed between values of 13N-NH3 and 18F-FDG for calculation of ESV [r=0.80, P<.0001, standard error of estimate (SEE)=0.10], EDV (r=0.63, P=.005, SEE=0.14) and LVEF (r=0.84, P<.0001, SEE=9.5). LVEF measured with 13N-NH3 was significantly lower in MI rats in comparison to measurement with 18F-FDG (45.6±8.9 vs 54.9±9.3 %; P=.04).Conclusion: Correlations were moderate to good for the assessment of ESV, EDV and LVEF between gated 13N-NH3 and 18F-FDG. LVEF was underestimated with gated 13N-NH3 in rats with myocardial infarction. In healthy rats, LV volumes and LVEF can be measured reproducibly with either approach.

Robust labeling and comparative preclinical characterization of DOTA-TOC and DOTA-TATE - Corrected Proof

Irina Velikyan, Hui Xu, Manoj Nair, Håkan Hall — 2012-02-16

Abstract: Objectives: Various radionuclide-labeled somatostatin analogues are used currently for diagnosis and therapy of neuroendocrine tumors. In particular, [68Ga]Ga-DOTA-TOC is commonly used for diagnosis, while [177Lu]Lu-DOTA-TATE is used for therapy. With the development of theranostics and personalized medicine where the imaging diagnosis is tailored to the subsequent radiotherapy, it is of paramount importance to investigate the relevance of the ligand exchange. The aim of this study was to compare binding capacity of [67/68Ga]Ga-DOTA-TOC ([67/68Ga]Ga-N-(4,7,10-(tris(carboxymethyl)-1,4,7,10-tetraazacyclododecan-1-yl)acetyl-D-Phe-c[Cys-D-Tyr-Trp-Lys-Thr-Cys]-Thr(ol)) and [67/68Ga]Ga-DOTA-TATE ([67/68Ga]Ga-N-(4,7,10-(tris(carboxymethyl)-1,4,7,10-tetraazacyclododecan-1-yl)acetyl-D-Phe-c[Cys-D-Tyr-Trp-Lys-Thr-Cys]-Thr) in vitro in monkey brain cryosections and in vivo in the rat, where, in contrast to transfected cell lines, there is a heterogeneous distribution of somatostatin receptor (SSTR) subtypes. The influence of various production methods of [68Ga]Ga-DOTA-TOC and [68Ga]Ga-DOTA-TATE on the biological performance of the tracers was also studied.Material and Methods: [67Ga]Ga-DOTA-TOC, [68Ga]Ga-DOTA-TOC, [67Ga]Ga-DOTA-TATE and [68Ga]Ga-DOTA-TATE were synthesized including preconcentration and purification of the generator eluate. The binding of the radioligands was assessed in vitro using autoradiography on cryosections of Rhesus monkey brains and in vivo/ex vivo using organ distribution studies in rats.Results and Discussion: The tracer production method was improved in terms of higher robustness, simplification and good manufacturing practice (GMP) relevance. The synthesis variation did not influence the biological performance of the tracers. There was no statistically significant difference observed in the binding of [67/68Ga]Ga-DOTA-TOC and [67/68Ga]Ga-DOTA-TATE either in brain cortex in vitro or in rat biodistribution and uptake in SSTR-positive tissues such as pancreas, adrenals and pituitary. The uptake in these organs was precluded by the excess of octreotide (Sandostatin). The 10-fold higher affinity to SSTR2 of DOTA-TATE as compared to DOTA-TOC known from studies in transfected cells was reflected in a slightly more intense binding of [67/68Ga]Ga-DOTA-TATE than of [67/68Ga]Ga-DOTA-TOC in the monkey brain sections in vitro, but not in vivo in the rat.Conclusion: A robust 68Ga-labeling method was introduced. The difference in the uptake of [67/68Ga]Ga-DOTA-TOC and [67/68Ga]Ga-DOTA-TATE in SSTR2-positive organs was not statistically significant either in vitro in tissue studies or in vivo/ex vivo in rat experiments. The results indicate that the more complex environment in vitro and in vivo diminishes the difference observed in transfected cell line binding.

Pd2(dba)3/P(MeNCH2CH2)3N·HCl-mediated Stille cross-coupling of [1-11C]acetyl chloride with aryl and heteroaryl stannanes - Corrected Proof

Takuya Arai — 2012-02-16

Abstract: Introduction: Developments in the Stille cross-coupling of [1-11C]acetyl chloride with tributylphenylstannane mediated by the Pd2(dba)3/P(MeNCH2CH2)3N·HClsystem are reported.Methods: The reaction conditions for the cross-coupling of [1-11C]acetyl chloride with tributylphenylstannane were optimized, and the reaction scope was investigated.Results: The cross-couplings of [1-11C]acetyl chloride with a range of aryl and heteroaryl stannanes were performed with good to excellent radiochemical conversions using the developed method.Conclusions: A highly efficient method for the Stille cross-coupling of [1-11C]acetyl chloride with organostannanes was demonstrated. It is anticipated that our method will be an attractive addition to the carbon-11-labeling procedures available for the synthesis of positron emission tomography probes.

A multimodality reporter gene for monitoring transplanted stem cells - Corrected Proof

2012-02-16

Abstract: Introduction: The aim of this study is to explore the feasibility of a triple-fused reporter gene, termed TGF [herpes simplex virus type 1 thymidine kinase (HSV1-tk), enhanced green fluorescent protein (eGFP) and firefly luciferase (Fluc)], to monitor stem cells using multimodality molecular imaging.Methods: A recombinant adenovirus vector carrying the triple-fused reporter gene (Ad5-TGF) was constructed. Bone marrow mesenchymal stem cells (BMSCs) were transfected with different virus titers of Ad5-TGF [multiplicities of infection (MOIs) were 0, 50, 100, 150, 200 and 250]. The mRNA and protein expressions of HSV1-tk, eGFP and Fluc in the transfected BMSCs were evaluated using polymerase chain reaction and Western blot. After the transfection of the BMSCs with different virus titers of Ad5-TGF (MOIs were 25, 50, 75, 100 and 125), their uptake rates of 131I-FIAU were measured. Whole-body fluorescence, bioluminescence and micro-positron emission tomography (PET) images were acquired 1 day after the transfected BMSCs were injected into the left forelimb of rats.Results: After the transfection with different titers of Ad5-TGF, the positive transfection rate reached a peak (70%) when the MOI was 100. HSV1-tk, eGFP and Fluc mRNA and protein were detected in the Ad5-TGF-transfected BMSCs, which implies their successful transfection and expression. The BMSCs uptake of 131I-FIAU increased with the adenovirus titer and incubation time and reached a plateau (approximately 5.3%) after 3 h. Strong signals were observed in the injected left forearms in the fluorescence, bioluminescence and micro-PET images.Conclusions: A triple-fused reporter gene, TGF, can be used as a multifunctional molecular probe for multimodality imaging.

Radiolabelling and evaluation of novel haloethylsulfoxides as PET imaging agents for tumor hypoxia - Corrected Proof

2012-02-16

Abstract: The significance of imaging hypoxia with the PET ligand [18F]FMISO has been demonstrated in a variety of cancers. However, the slow kinetics of [18F]FMISO require a 2-h delay between tracer administration and patient scanning. Labelled chloroethyl sulfoxides have shown faster kinetics and higher contrast than [18F]FMISO in a rat model of ischemic stroke. However, these nitrogen mustard analogues are unsuitable for routine production and use in humans. Here we report on the synthesis and in vitro and in vivo evaluation of two novel sulfoxides which we synthesised from a single precursor molecule via either 2-[18F]fluoroethyl azide click chemistry or conventional nucleophilic displacement of a chloride leaving group. The yields of the click chemistry approach were 90±5% of [18F] based on 2-[18F]fluoroethyl azide, and the yields for the SN reaction were 15±5% of [18F] based on K[18F]F. Both radiotracers underwent metabolism in an in vitro assay using S9 liver fractions with biological half-lives of 32.39 and 43.32 min, respectively. Imaging studies using an SK-RC-52 tumor model in BALB/c nude mice have revealed that only [18F] is retained in hypoxic tumors, whereas [18F] is cleared from those tumors at a rate similar to that of muscle tissue. [18F] has emerged as a promising new lead structure for further development of sulfoxide-based hypoxia imaging agents. In particular, the mechanism of uptake needs to be elucidated and changes to the chemical structure need to be made in order to reduce metabolism and improve radiotracer kinetics.

Pretargeting vs. direct targeting of human betalox5 islet cells subcutaneously implanted in mice using an anti-human islet cell antibody - Corrected Proof

2012-02-10

Abstract: Introduction: We previously demonstrated MORF/cMORF pretargeting of human islets and betalox 5 cells (a human beta cell line) transplanted subcutaneously in mice with the anti-human islet antibody, HPi1. We now compare pretargeting with direct targeting in the beta cell transplant model to evaluate the degree to which target/non-target (T/NT) ratios may be improved by pretargeting.Methods: Specific binding of an anti-human islet antibody HPi1 to the beta cells transplanted subcutaneously in mice was examined against a negative control antibody. We then compared pretargeting by MORF-HPi1 plus 111In-labeled cMORF to direct targeting by 111In-labeled HPi1.Results: HPi1 binding to betalox5 human cells in the transplant was shown by immunofluorescence. Normal organ 111In backgrounds by pretargeting were always lower, although target accumulations were similar. More importantly, the transplant to pancreas and liver ratios was, respectively, 26 and 10 by pretargeting as compared to 9 and 0.6 by direct targeting.Conclusions: Pretargeting greatly improves the T/NT ratios, and based on the estimated endocrine to exocrine ratio within a pancreas, pretargeting may be approaching the sensitivity required for successful imaging of human islets within this organ.

Synthesis and evaluation of nucleoside radiotracers for imaging proliferation - Corrected Proof

2012-02-10

Abstract: Introduction: Uncontrolled proliferation is a fundamental characteristic of cancer, and consequently, imaging of tumor proliferative status finds interest clinically both as a diagnostic tool and for evaluation of response to treatment. Positron emission tomography (PET) radiotracers based on a nucleoside core, such as 3′-[18F]fluoro-3′-deoxythymidine ([18F]FLT), have been extensively studied for this purpose. However, [18F]FLT suffers from poor DNA incorporation leading to occasional poor correlation of [18F]FLT tumor uptake with other proliferation indicators such as Ki-67 immunostaining.Methods: N3-((1-(2-[18F]fluoroethyl)-1H-[1,2,3]-triazol-4-yl)methyl)thymidine ([18F]2) and N3-((1-(2-[18F]fluoroethyl)-1H-[1,2,3]-triazol-4-yl)methyl)-4′-thio-β-thymidine ([18F]3) were synthesized by click chemistry from [18F]fluoroethyl azide and by direct nucleophilic substitution of a tosylate precursor. Metabolic stability and phosphorylation potential of the radiotracers were evaluated in vitro and compared to [18F]FLT. Further, metabolic stability and biodistribution analysis of [18F]2 and [18F]3 were evaluated in vivo.Results: Stable isotope standards and radiochemistry precursors were synthesized by modification of existing literature procedures. [18F]2 and [18F]3 were synthesized in a radiochemical yield of 8%–12% (end of synthesis, non-decay corrected). Both nucleosides were stable to metabolic degradation by thymidine phosphorylase, and in vivo stability analysis showed only one metabolite for [18F]3. No phosphorylation of [18F]2 could be detected in HCT116 cell homogenates, and in the same assay, only minor (∼8%) phosphorylation of [18F]3 was observed. Biodistribution in Balb/c mice indicated rapid clearance for [18F]2 and [18F]3 to a lesser extent.Conclusions: The favorable biodistribution and metabolic profile of [18F]3 warrant further investigation as a next-generation PET proliferation marker.

Radiolabeled Zn-DPA as a potential infection imaging agent - Corrected Proof

2012-02-10

Abstract: Introduction: A zinc-dipicolylamine analog (Zn-DPA) conjugated with a fluorophore (PSVue®794) has been shown to image bacterial infections in mice. However, radiolabeled Zn-DPA has not previously been considered for nuclear imaging of infection.Methods: Both 111In-labeled DOTA-biotin and Zn-DPA-biotin were combined using streptavidin (SA) as a noncovalent linker. Mice injected intramuscularly with Streptococcus pyogenes (infection model) or with lipopolysaccharide (LPS) (inflammation model) were coinjected intravenously with 6 μg of DPA as PSVue794 and as 111In-DOTA-biotin/SA/biotin-Zn-DPA. Periodic fluorescent and SPECT (single photon emission computed tomography)/CT (computed tomography) images were acquired, and biodistributions were obtained at 22 h.Results: Histological examination confirmed the validity of both the infection and inflammation animal models. Both the whole-body optical and nuclear images showed obvious accumulations in the target thigh in both models at all time points. At 22 h, the average target thigh accumulation of 111In was 1.66%ID/g (S.D. 0.15) in the infection mice compared to 0.58%ID/g (S.D. 0.07) in the inflammation mice (P<.01), and the 111In target/normal thigh ratio was 2.8 fold higher in the infection animals compared to the inflammation animals.Conclusions: These preliminary results show that Zn-DPA within streptavidin targets S. pyogenes-infected mice similarly to its free fluorescent analogue. The significantly higher accumulation in the live bacterial infection thigh compared to that of the LPS-induced inflammation thigh suggests that Zn-DPA may be a promising imaging agent to distinguish between bacterial infections and sterile inflammations.

Radiation dosimetry of the translocator protein ligands [18F]PBR111 and [18F]PBR102 - Corrected Proof

2012-02-02

Abstract: Introduction: The translocator protein (TSPO) ligands [18F]PBR111 and [18F]PBR102 show promise for imaging neuroinflammation. Our aim was to estimate the radiation dose to humans from primate positron emission tomography (PET) studies using these ligands and compare the results with those obtained from studies in rodents.Methods: [18F]PBR111 and [18F]PBR102 PET–computed tomography studies were carried out in baboons. The cumulated activity in the selected source organs was obtained from the volume of interest time–activity curves drawn on coronal PET slices and adjusted for organ mass relative to humans. Radiation dose estimates were calculated in OLINDA/EXM Version 1.1 from baboon studies and compared with those calculated from Sprague–Dawley rat tissue concentration studies, also adjusted for relative organ mass.Results: In baboons, both ligands cleared rapidly from brain, lung, kidney and spleen and more slowly from liver and heart. For [18F]PBR111, the renal excretion fraction was 6.5% and 17% for hepatobiliary excretion; for [18F]PBR102, the renal excretion was 3.0% and 15% for hepatobiliary excretion. The estimated effective dose in humans from baboon data was 0.021 mSv/MBq for each ligand, whilst from rat data, the estimates were 0.029 for [18F]PBR111 and 0.041 mSv/MBq for [18F]PBR102.Conclusion: Biodistribution in a nonhuman primate model is better suited than the rat model for the calculation of dosimetry parameters when translating these ligands from preclinical to human clinical studies. Effective dose calculated from rat data was overestimated compared to nonhuman primate data. The effective dose coefficient for both these TSPO ligands determined from PET studies in baboons is similar to that for [18F]FDG.

A fast chemoenzymatic synthesis of [11C]-N5,N10-methylenetetrahydrofolate as a potential PET tracer for proliferating cells - Corrected Proof

Muhammad Saeed, Timothy J. Tewson, Colbin E. Erdahl, Amnon Kohen — 2012-02-02

Abstract: Introduction: Thymidylate synthase and folate receptors are well-developed targets of cancer therapy. Discovery of a simple and fast method for the conversion of 11CH3Ito[11C]-formaldehyde (11CH2O) encouraged us to label the co-factor of this enzyme. Preliminary studies conducted on cell lines have demonstrated a preferential uptake of [11-14C]-(R)-N5,N10-methylene-5,6,7,8-tetrahydrofolate (14CH2H4folate) by cancerous cell vs. normal cells from the same organ (Saeed M., Sheff D. and Kohen A. Novel positron emission tomography tracer distinguishes normal from cancerous cells. J Biol Chem 2011;286:33872–33878), pointing out 11CH2H4folate as a positron emission tomography (PET) tracer for cancer imaging. Herein we report the synthesis of 11CH2H4folate, which may serve as a potential PET tracer.Methods: In a remotely controlled module, methyl iodide (11CH3I) was bubbled into a reaction vial containing trimethylamine N-oxide in N,N-Dimethylformamide (DMF) and heated to 70°C for 2 min. Formaldehyde (11CH2O) formed after the completion of reaction was then mixed with a solution of freshly prepared tetrahydrofolate (H4folate) by using a fast chemoenzymatic approach to accomplish synthesis of 11CH2H4folate. Purification of the product was carried out by loading the crude reaction mixture on a SAX cartridge, washing with water to remove unbound impurities and finally eluting with a saline solution.Results: The synthesis and purification of 11CH2H4folate were completed within 5 min. High-performance liquid chromatography analysis of the product after SAX purification indicates that more than 90% of the radioactivity that was retained on the SAX cartridge was in 11CH2H4folate, with minor (<10%) radioactivity due to unreacted 11CH2O.Conclusion: We present a fast (∼5 min) synthesis and purification of 11CH2H4folate as a potential PET tracer. The final product is received in physiologically compatible buffer (100 mM sodium phosphate, pH 7.0 containing 500 mM NaCl) and ready for use in vivo.

Quantification of regional cerebral blood flow in rats using an arteriovenous shunt and micro-PET - Corrected Proof

2012-01-20

Abstract: Introduction: Measurement of regional cerebral blood flow (rCBF) in rodents can provide knowledge of pathophysiology of the cerebral circulation, but generally requires blood sampling for analysis during positron emission tomography (PET). We therefore tested the feasibility of using an arteriovenous (AV) shunt in rats for less invasive blood analysis.Methods: Six anesthetized rats received [15O]H2O and [15O]CO PET scans with their femoral artery and vein connected by an AV shunt, the activity within which was measured with a germanium ortho-oxysilicate scintillation detector. The [15O]H2O was intravenously injected either at a faster or slower injection rate, while animals were placed either with their head or heart centered in the gantry. The time–activity curve (TAC) from the AV shunt was compared with that from the cardiac ventricle in PET image. The rCBF values were calculated by a nonlinear least-square method using the dispersion-corrected AV-shunt TAC as an input.Results: The AV-shunt TAC had higher signal-to-noise ratio, but also had delay and dispersion compared with the image-derived TAC. The delay time between the AV-shunt TAC and image-based TAC ranged from 11 to 21 s, while the dispersion was estimated to be ∼5 s as a time constant of the dispersion model of exponential function, and both were properly corrected. In a steady-state condition of [15O]CO PET, the blood activity concentration by AV-shunt TAC was also comparable in height with the image-based TAC corrected for partial volume. Whole-brain CBF values measured by [15O]H2O were 0.37±0.04 (mean±S.D.) ml/g/min, partition coefficient was 0.73±0.04 ml/g, and the CBF varied in a linear relationship with partial pressure of carbon dioxide during each scan.Conclusions: The AV-shunt technique allows less invasive, quantitative and reproducible measurement of rCBF in [15O]H2O PET studies in rats than direct blood sampling and radioassay.

A historical perspective on the specific activity of radiopharmaceuticals: what have we learned in the 35 years of the ISRC? - Corrected Proof

Suzanne E. Lapi, Michael J. Welch — 2012-01-20

Abstract: Specific activity (SA), defined as the amount of radioactivity per unit mass of a compound, is arguably one of the most important parameters in radiopharmaceutical development, particularly in quality control of carbon-11- and fluorine-18-labeled compounds. This review article will outline the progression of improvements in SA over the last few decades. The International Symposium of Radiopharmaceutical Chemistry abstracts were an excellent source of materials for this review and will be referenced throughout.

Varenicline increases in vivo striatal dopamine D2/3 receptor binding: an ultra-high-resolution pinhole [123I]IBZM SPECT study in rats - Corrected Proof

2012-01-20

Abstract: Introduction: Ex vivo storage phosphor imaging rat studies reported increased brain dopamine D2/3 receptor (DRD2/3) availability following treatment with varenicline, a nicotinergic drug. However, ex vivo studies can only be performed using cross-sectional designs. Small-animal imaging offers the opportunity to perform serial assessments. We evaluated whether high-resolution pinhole single photon emission computed tomography (SPECT) imaging in rats was able to reproduce previous ex vivo findings.Methods: Rats were imaged for baseline striatal DRD2/3 availability using ultra-high-resolution pinhole SPECT (U-SPECT-II) and [123I]IBZM as a radiotracer, and randomized to varenicline (n=7; 2 mg/kg) or saline (n=7). Following 2 weeks of treatment, a second scan was acquired.Results: Significantly increased striatal DRD2/3 availability was found following varenicline treatment compared to saline (time⁎treatment effect): posttreatment difference in binding potential between groups corrected for initial baseline differences was 2.039 (P=.022), indicating a large effect size (d=1.48).Conclusions: Ultra-high-resolution pinhole SPECT can be used to assess varenicline-induced changes in DRD2/3 availability in small laboratory animals over time. Future small-animal studies should include imaging techniques to enable repeated within-subjects measurements and reduce the amount of animals.

In vitro and in vivo evaluation of a 64Cu-labeled NOTA-Bn-SCN-Aoc-bombesin analogue in gastrin-releasing peptide receptor expressing prostate cancer - Corrected Proof

2012-01-20

Abstract: Introduction: Bombesin (BN) is an amphibian peptide that binds to the gastrin-releasing peptide receptor (GRPR). It has been demonstrated that BN analogues can be radiolabeled for potential diagnosis and treatment of GRPR-expressing malignancies. Previous studies have conjugated various chelators to the eight C-terminal amino acids of BN [BN(7-14)] for radiolabeling with 64Cu. Recently, (1,4,7-triazacyclononane-1,4,7-triacetic acid) (NOTA) has been evaluated as the five-coordinate 64Cu complex, with results indicating GRPR-specific tumor uptake. This study aimed to conjugate S-2-(4-isothiocyanatobenzyl)-NOTA (p-SCN-Bn-NOTA) to BN(7-14) such that it could form a six-coordinate complex with 64Cu and to evaluate the resulting peptide.Methods: p-SCN-NOTA was conjugated to 8-aminooctanoic acid (Aoc)-BN(7-14) in solution to yield NOTA-Bn-SCN-Aoc-BN(7-14). The unlabeled peptide was evaluated in a cell binding assay using PC-3 prostate cancer cells and 125I-Tyr4-BN to determine the IC50 value. The peptide was radiolabeled with 64Cu and evaluated for internalization into PC-3 cells and for tumor uptake in mice bearing PC-3 xenografts using biodistribution and micro-positron emission tomography imaging studies.Results: The binding assay demonstrated that NOTA-Bn-SCN-Aoc-BN(7-14) bound with high affinity to GRPR with an IC50 of 1.4 nM. The radiolabeled peptide demonstrated time-dependent internalization into PC-3 cells. In vivo, the peptide demonstrated tumor-specific uptake and imaging that were comparable to those of previously reported 64Cu-labeled BN analogues.Conclusions: These studies demonstrate that 64Cu-NOTA-Bn-SCN-Aoc-BN(7-14) binds to GRPR-expressing cells and that it can be used for imaging of GRPR-expressing prostate cancer.

Evaluation of the relationship between [18F]FDG and P-glycoprotein expression: an experimental study - Corrected Proof

Chunjing Yu, Weixing Wan, Bin Zhang, Shengming Deng, Tzu-Chen Yen, Yiwei Wu — 2012-01-20

Abstract: Introduction: P-glycoprotein (P-gp) is a cell-membrane-associated protein that transports a variety of drug substrates. We sought to evaluate the relationship between 2-[18F]fluoro-2-deoxy-d-glucose ([18F]FDG) and P-gp expression using breast carcinoma Bcap37/multidrug resistant (MDR1) and Bcap37 in vitro and in vivo.Methods: The function of P-gp expressed in Bcap37/MDR1 cells was evaluated using verapamil (VER), a classical inhibitor of P-gp. The accumulation of 99mTc-methoxyisobutylisonitrile ([99mTc]MIBI) in vitro was measured. In vivo imaging of severe combined immune deficiency (SCID) mice implanted with Bcap37 and Bcap37/MDR1 cells was performed by scintigraphy and micro-positron emission tomography (PET).Results: The uptake of [99mTc]MIBI was 0.62%±0.05% in the Bcap37/MDR1 cells and 2.02%±0.28% in the Bcap37 cells. VER significantly increased the uptake of [99mTc]MIBI in the Bcap37/MDR1 cells (1.90%±0.09%) but not in the Bcap37 cells (2.15%±0.27%). In vivo, neither the Bcap37 nor Bcap37/MDR1 tumors grown in the SCID mice could be detected by [99mTc]MIBI scintigraphy. Both the Bcap37 and Bcap37/MDR1 tumors were visible by micro-PET. The mean standardized uptake value (SUV) was significantly higher in the Bcap37 tumors (1.00±0.06) than in the Bcap37/MDR1 (0.67±0.11) tumors. VER significantly increased the mean SUV in the Bcap37/MDR1 tumors (1.02±0.16) but not in the Bcap37 tumors (1.09±0.22).Conclusions: [18F]FDG combined with VER may be an effective noninvasive method of determining P-gp expression in tumors.