David S. Casebier

Evaluation of the Novel Myocardial Perfusion Positron-Emission Tomography Tracer 18F-BMS-747158-02 Comparison to 13N-Ammonia and Validation With Microspheres in a Pig Model

Background— Positron-emission tomography (PET) tracers for myocardial perfusion are commonly labeled with short-lived isotopes that limit their widespread clinical use. 18F-BMS-747158-02 (18F-BMS) is a novel pyridaben derivative that was evaluated for assessment of myocardial perfusion by comparison with 13N-ammonia (13NH3) and with radioactive microspheres in a pig model. Methods and Results— Fourteen pigs injected with 500 MBq of 13NH3 or 100 to 200 MBq of 18F-BMS underwent dynamic PET at rest and during pharmacological stress. In 8 of these pigs, 18F-BMS was injected during stress combined with transient, 2.5-minute constriction of the left anterior descending coronary artery. Radioactive microspheres were coinjected with 18F-BMS. Ratios of myocardial tracer uptake to surrounding tissues were determined, and myocardial blood flow was quantified by compartmental modeling. Both tracers showed high and homogeneous myocardial uptake. Compared with 13NH3, 18F-BMS showed higher activity ratios between myocardium and blood (rest 2.5 versus 4.1; stress 2.1 versus 5.8), liver (rest 1.2 versus 1.8; stress 0.7 versus 2.0), and lungs (rest 2.5 versus 4.2; stress 2.9 versus 6.4). Regional myocardial blood flow assessed with 18F-BMS PET showed good correlation (r=0.88, slope=0.84) and agreement (mean difference −0.10 [25th percentile −0.3, 75th percentile 0.1 mL · min−1 · g−1]) with that measured with radioactive microspheres over a flow range from 0.1 to 3.0 mL · min−1 · g−1. The extent of defects induced by left anterior descending coronary artery constriction measured by 18F-BMS and microspheres also correlated closely (r=0.63, slope=1.1). Conclusions— 18F-BMS-747158-02 is a very attractive new PET perfusion tracer that allows quantitative assessment of regional myocardial perfusion over a wide flow range. The long half-life of 18F renders this tracer useful for clinical PET/CT applications in the workup of patients with suspected or proven coronary artery disease.

Initial Characterization of an 18F-Labeled Myocardial Perfusion Tracer

PET allows for quantitative, regional myocardial perfusion imaging. The short half-lives of the perfusion tracers currently in use limit their clinical applicability. Here, the biodistribution and imaging quality of a new 18F-labeled myocardial perfusion agent (18F-BMS-747158-02) in an animal model are described. Methods: The biodistribution of 18F-BMS-747158-02 was determined at 10 and 60 min after injection. The first-pass extraction fraction of the tracer was measured in isolated rat hearts perfused with the Langendorff method. Small-animal PET imaging was used to study tracer retention. Results: The biodistribution at 10 min after injection demonstrated high myocardial uptake (3.1 percentage injected dose per gram [%ID/g]) accompanied by little activity in the lungs (0.3 %ID/g) and liver (1.0 %ID/g). The tracer showed a high and flow-independent myocardial first-pass extraction fraction, averaging 0.94 (SD = 0.04). PET imaging provided excellent delineation of myocardial structures. The heart-to-lung activity ratio increased from 4.7 to 10.2 between 1 and 15 min after tracer injection (at rest). Adenosine infusion (140 μg/kg/min) led to a significant increase in myocardial tracer retention (from 1.68 [SD = 0.23]) s−1 to 3.21 [SD = 0.92] s−1; P = 0.03). Conclusion: The observation of a high and flow-independent first-pass extraction fraction promises linearity between tracer uptake and myocardial blood flow. Sustained myocardial tracer uptake, combined with high image contrast, will allow for imaging protocols with tracer injection at peak exercise followed by delayed imaging. Thus, 18F-BMS-747158-02 is a promising new tracer for the quantitative imaging of myocardial perfusion and can be distributed to imaging laboratories without a cyclotron.

Evaluation of LMI1195, a Novel 18F-Labeled Cardiac Neuronal PET Imaging Agent, in Cells and Animal Models

Background—Heart failure has been associated with impaired cardiac sympathetic neuronal function. Cardiac imaging with radiolabeled agents that are substrates for the neuronal norepinephrine transporter (NET) has demonstrated the potential to identify individuals at risk of cardiac events. N-[3-Bromo-4-(3-[18F]fluoro-propoxy)-benzyl]-guanidine (LMI1195) is a newly developed 18F-labeled NET substrate designed to allow cardiac neuronal imaging with the high sensitivity, resolution, and quantification afforded by positron emission tomography (PET). Methods and Results—LMI1195 was evaluated in comparison with norepinephrine (NE) in vitro and 123I-meta-iodobenzylguanidine (MIBG) in vivo. The affinity (Ki) of LMI1195 for NET was 5.16±2.83 &mgr;mol/L, similar to that of NE (3.36±2.77 &mgr;mol/L) in a cell membrane–binding assay. Similarly, LMI1195 uptake kinetics examined in a human neuroblastoma cell line had Km and Vmax values of 1.44±0.76 &mgr;mol/L and 6.05±3.09 pmol/million cells per minute, comparable to NE (2.01±0.85 &mgr;mol/L and 6.23±1.52 pmol/million cells per minute). In rats, LMI1195 heart uptake at 15 and 60 minutes after intravenous administration was 2.36±0.38% and 2.16±0.38% injected dose per gram of tissue (%ID/g), similar to 123I-MIBG (2.14±0.30 and 2.19±0.27%ID/g). However, the heart to liver and lung uptake ratios were significantly higher for LMI1195 than for 123I-MIBG. In rabbits, desipramine (1 mg/kg), a selective NET inhibitor, blocked LMI1195 heart uptake by 82%, which was more effective than 123I-MIBG (53%), at 1 hour after dosing. Sympathetic denervation with 6-hydroxydopamine, a neurotoxin, resulted in a marked (79%) decrease in LMI1195 heart uptake. Cardiac PET imaging with LMI1195 in rats, rabbits, and nonhuman primates revealed clear myocardium with low radioactivity levels in the blood, lung, and liver. Imaging in rabbits pretreated with desipramine showed reduced heart radioactivity levels in a dose-dependent manner. Additionally, imaging in sympathetically denervated rabbits resulted in low cardiac image intensity with LMI1195 but normal perfusion images with flurpiridaz F 18, a PET myocardial perfusion imaging agent. In nonhuman primates pretreated with desipramine (0.5 mg/kg), imaging with LMI1195 showed a 66% decrease in myocardial uptake. In a rat model of heart failure, the LMI1195 cardiac uptake decreased as heart failure progressed. Conclusions—LMI1195 is a novel 18F imaging agent retained in the heart through the NET and allowing evaluation of the cardiac sympathetic neuronal function by PET imaging.

A New 18F-Labeled Myocardial PET Tracer: Myocardial Uptake After Permanent and Transient Coronary Occlusion in Rats

Conventional myocardial perfusion PET tracers require onsite tracer production because of their short radioactive half-lives. To investigate the potential of a new 18F-labeled pyridazinone analog (18F-BMS-747158-02), we characterized this tracer in a rat model of permanent and transient coronary occlusion using small-animal PET. Methods: Myocardial 18F-BMS-747158-02 distribution in healthy rats (n = 7), rats with transient (3-min) left coronary artery occlusion (n = 11), and rats with permanent left coronary occlusion (n = 11) was analyzed with a dedicated small-animal PET scanner. Results: Normal hearts demonstrated intense and almost homogeneous tracer uptake throughout the left ventricle for more than 2 h. During permanent coronary occlusion, PET demonstrated perfusion defects, which remained unchanged (37.6% ± 8.8%, 37.4% ± 10.2%, and 36.2% ± 9.8% left ventricle at 15, 45, and 115 min, respectively, after tracer injection). After transient ischemia, the induced defect size decreased significantly after reperfusion (16.2% ± 9.3%, 6.0% ± 6.5%, and 1.4% ± 1.3% left ventricle). Tracer reinjection after transient ischemia resulted in normalization of the induced defect. Conclusion: Coronary occlusion yielded distinct myocardial 18F-BMS-747158-02 uptake defects in the area of ischemia, which demonstrated normalization of activity after reperfusion and reinjection. These promising kinetic parameters may allow for assessment of flow using exercise–rest protocols similar to those used in combination with exercise and rest perfusion SPECT.

Evaluation of a Novel 18 F-Labeled Positron-Emission Tomography Perfusion Tracer for the Assessment of Myocardial Infarct Size in Rats

Background—The goal of this study was to evaluate a new 18F-labeled positron-emission tomography (PET) perfusion tracer, 18F BMS747158-02, for the assessment of myocardial infarct (MI) size. Methods and Results—Wistar rats were studied 24 hours after ligation of the left coronary artery either permanently (n=15) or transiently (n=16) for 30 minutes. Seven nonoperated rats were studied as controls. The rats were injected with 37 MBq of 18F BMS747158-02 and imaged with a small animal PET scanner for 20 minutes. Polar maps were generated for measurement of PET defect size, and left ventricular systolic and diastolic volumes were assessed in gated images. As a reference, MI size was determined by 2,3,5-triphenyltetrazolium chloride staining of left ventricular tissue samples. Permanent or transient ligation of the left coronary artery produced transmural or subendocardial MI of variable sizes, respectively. In normal rats, PET imaging demonstrated intense and homogeneous uptake of 18F BMS747158-02 throughout the myocardium. After ligation, sharply defined perfusion defects were present. Throughout the imaging period, the defect size correlated closely with the MI size either after permanent (r=0.88; P<0.01; mean difference, 1.86%) or transient (r=0.92; P<0.01; mean difference, 2.16%) ligation of the left coronary artery. Moreover, reduction of left ventricular systolic function measured with PET correlated with the MI size (r=−0.81; P<0.01; n=23). Conclusions—Myocardial 18F BMS747158-02 PET imaging provides excellent image quality and uptake properties, enabling accurate evaluation of MI size and left ventricular function in rats. It is a promising technique for evaluation of MI size in clinical trials.

Evaluation of a Novel 18F-Labeled PET Perfusion Tracer for the Assessment of Myocardial Infarct Size in Rat

Background—The goal of this study was to evaluate a new 18F-labeled positron-emission tomography (PET) perfusion tracer, 18F BMS747158-02, for the assessment of myocardial infarct (MI) size. Methods and Results—Wistar rats were studied 24 hours after ligation of the left coronary artery either permanently (n=15) or transiently (n=16) for 30 minutes. Seven nonoperated rats were studied as controls. The rats were injected with 37 MBq of 18F BMS747158-02 and imaged with a small animal PET scanner for 20 minutes. Polar maps were generated for measurement of PET defect size, and left ventricular systolic and diastolic volumes were assessed in gated images. As a reference, MI size was determined by 2,3,5-triphenyltetrazolium chloride staining of left ventricular tissue samples. Permanent or transient ligation of the left coronary artery produced transmural or subendocardial MI of variable sizes, respectively. In normal rats, PET imaging demonstrated intense and homogeneous uptake of 18F BMS747158-02 throughout the myocardium. After ligation, sharply defined perfusion defects were present. Throughout the imaging period, the defect size correlated closely with the MI size either after permanent (r=0.88; P<0.01; mean difference, 1.86%) or transient (r=0.92; P<0.01; mean difference, 2.16%) ligation of the left coronary artery. Moreover, reduction of left ventricular systolic function measured with PET correlated with the MI size (r=−0.81; P<0.01; n=23). Conclusions—Myocardial 18F BMS747158-02 PET imaging provides excellent image quality and uptake properties, enabling accurate evaluation of MI size and left ventricular function in rats. It is a promising technique for evaluation of MI size in clinical trials.

Synthesis and Biological Evaluation of Pyridazinone Analogues as Potential Cardiac Positron Emission Tomography Tracers

A series of fluorinated pyridazinone derivatives with IC50 values ranging from 8 to 4000 nM for the mitochondrial complex 1 (MC1) have been prepared. Structure-activity relationship (SAR) assessment indicated preference of the fluorine label to be incorporated on an alkyl side chain rather than directly on the pyridazinone moiety. Tissue distribution studies of a series of analogues ([18F] 22-28) in Sprague-Dawley (SD) rats identified [18F]27 as the most promising radiotracer with high uptake in cardiac tissue (3.41%ID/g; 30 min post injection) in addition to favorable heart to nontarget organ distribution ratios. MicroPET images of SD rats and nonhuman primates after [18F]27 administration allowed easy assessment of the myocardium through 60 min with minimal lung or liver interference.

Discovery of a novel series of Notch-sparing γ-secretase inhibitors

Using a cell-based assay, we have identified a new series of Notch-sparing gamma-secretase inhibitors from HTS screening leads 2a and 2e. Lead optimization studies led to the discovery of analog 8e with improved gamma-secretase inhibitory potency and Notch-sparing selectivity.

(S)-N-(5-Chlorothiophene-2-sulfonyl)-β,β-diethylalaninol a Notch-1-sparing γ-secretase inhibitor

Accumulation of beta-amyloid (Abeta), produced by the proteolytic cleavage of amyloid precursor protein (APP) by beta- and gamma-secretase, is widely believed to be associated with Alzheimers disease (AD). Research around the high-throughput screening hit (S)-4-chlorophenylsulfonyl isoleucinol led to the identification of the Notch-1-sparing (9.5-fold) gamma-secretase inhibitor (S)-N-(5-chlorothiophene-2-sulfonyl)-beta,beta-diethylalaninol 7.b.2 (Abeta(40/42) EC(50)=28 nM), which is efficacious in reduction of Abeta production in vivo.

Synthesis and Cardiac Imaging of 18F-Ligands Selective for β1-Adrenoreceptors

A series of potent and selective β1-adrenoreceptor ligands were identified (IC50 range, 0.04-0.25 nM; β1/β2 selectivity range, 65-450-fold), labeled with the PET radioisotope fluorine-18 and evaluated in normal Sprague-Dawley rats. Tissue distribution studies demonstrated uptake of each radiotracers from the blood pool into the myocardium (0.48-0.62% ID/g), lung (0.63-0.97% ID/g), and liver (1.03-1.14% ID/g). Dynamic μPET imaging confirmed the in vivo dissection studies.