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 µmol/L, similar to that of NE (3.36 ± 2.77 µmol/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 µmol/L and 6.05 ± 3.09 pmol/million cells per minute, comparable to NE (2.01 ± 0.85 µmol/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 (18)F imaging agent retained in the heart through the NET and allowing evaluation of the cardiac sympathetic neuronal function by PET imaging.

Synthesis and Cardiac Imaging of (18)F-Ligands Selective for ß1-Adrenoreceptors.

A series of potent and selective ß1-adrenoreceptor ligands were identified (IC50 range, 0.04-0.25 nM; ß1/ß2 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.

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 (18)F-labeled positron-emission tomography (PET) perfusion tracer, (18)F 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 (18)F 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 (18)F 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 (18)F 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.

(S)-N-(5-Chlorothiophene-2-sulfonyl)-beta,beta-diethylalaninol a Notch-1-sparing gamma-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 Alzheimer's 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.

Assessment of 18F-labeled mitochondrial complex I inhibitors as PET myocardial perfusion imaging agents in rats, rabbits, and primates.

PURPOSE: Myocardial extractions of mitochondria complex I (MC-I) inhibitors were high and well correlated with flow. This study assessed the potential of MC-I inhibitors to be developed as myocardial perfusion imaging (MPI) agents. METHODS: RP1003, RP1004, and RP1005 representing three classes of MC-I inhibitor were synthesized and radio-labeled with (18)F. These agents were evaluated for IC(50) values, tissue biodistribution, and cardiac PET imaging. (18)F-RP1004 was further examined for first-pass extraction and by imaging in non-human primates (NHP) and rats following coronary ligation. RESULTS: RP1003, RP1004, and RP1005 exhibited high MC-I inhibitory activity with IC(50) of 3.7, 16.7, and 14.4 nM. Heart uptakes in rats (percent injected dose per gram tissue) at 15 and 60 min after injection were 3.52 +/- 0.36 and 2.68 +/- 0.20 for (18)F-RP1003, 2.40 +/- 0.21 and 2.67 +/- 0.27 for (18)F-RP1004, and 2.28 +/- 0.12 and 1.81 +/- 0.17 for (18)F-RP1005. The heart to lung and liver uptake ratios were favorable for cardiac imaging with these agents. In isolated perfused rabbit hearts, the uptake of (18)F-RP1004 increased from 0.74 +/- 0.19 to 1.68 +/- 0.39 mL/min/g at flow rates of 1.66 to 5.06 mL/min/g. These values were higher than or similar to that of (99m)Tc-sestamibi. Cardiac imaging with these agents in rats and rabbits allowed visualization of the heart with minimal lung interference and rapid liver activity clearance. Imaging with (18)F-RP1004 also showed clear myocardium and marked liver activity washout in the NHP and clear detection of the perfusion-deficit area associated with left coronary artery ligation in the rat. CONCLUSION: MC-I inhibitors have the potential to be a new class of MPI agent.

A new 18F-labeled myocardial PET tracer: myocardial uptake after permanent and transient coronary occlusion in rats.

UNLABELLED: Conventional myocardial perfusion PET tracers require onsite tracer production because of their short radioactive half-lives. To investigate the potential of a new (18)F-labeled pyridazinone analog ((18)F-BMS-747158-02), we characterized this tracer in a rat model of permanent and transient coronary occlusion using small-animal PET. METHODS: Myocardial (18)F-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 (18)F-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.

Discovery of a novel series of Notch-sparing gamma-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.

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.

Initial characterization of an 18F-labeled myocardial perfusion tracer.

UNLABELLED: 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 microg/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.

Mechanism of uptake and retention of F-18 BMS-747158-02 in cardiomyocytes: a novel PET myocardial imaging agent.

BACKGROUND: BMS-747158-02 is a novel fluorine 18-labeled pyridazinone derivative designed for cardiac imaging. The uptake and retention mechanisms of F-18 BMS-747158-02 in cardiac myocytes were studied in vitro, and the biodistribution of F-18 BMS-747158-02 was studied in vivo in mice. METHODS AND RESULTS: Fluorine 19 BMS-747158-01 inhibited mitochondrial complex I (MC-I) in bovine heart submitochondrial particles with an IC(50) of 16.6 +/- 3 nmol/L that was comparable to the reference inhibitors of MC-1, rotenone, pyridaben, and deguelin (IC(50) of 18.2 +/- 6.7 nmol/L, 19.8 +/- 2.6 nmol/L, and 23.1 +/- 1.5 nmol/L, respectively). F-18 BMS-747158-02 had high uptake in monolayers of neonatal rat cardiomyocytes (10.3% +/- 0.7% of incubated drug at 60 minutes) that was inhibited by 200 nmol/L of rotenone (91% +/- 2%) and deguelin (89% +/- 3%). In contrast, an inactive pyridaben analog, P-070 (IC(50) value >4 micromol/L in MC-1 assay), did not inhibit the binding of F-18 BMS-747158-02 in cardiomyocytes. Uptake and washout kinetics for F-18 BMS-747158-02 in rat cardiomyocytes indicated that the time to half-maximal (t((1/2))) uptake was very rapid (approximately 35 seconds), and washout t((1/2)) for efflux of F-18 BMS-747158-02 was greater than 120 minutes. In vivo biodistribution studies in mice showed that F-18 BMS-747158-02 had substantial myocardial uptake (9.5% +/- 0.5% of injected dose per gram) at 60 minutes and heart-to-lung and heart-to-liver ratios of 14.1 +/- 2.5 and 8.3 +/- 0.5, respectively. Positron emission tomography imaging in the mouse allowed clear cardiac visualization and demonstrated sustained myocardial uptake through 55 minutes. CONCLUSIONS: F-18 BMS-747158-02 is a novel positron emission tomography cardiac tracer targeting MC-I in cardiomyocytes with rapid uptake and slow washout. These characteristics allow fast and sustained accumulation in the heart.

BMS-747158-02: a novel PET myocardial perfusion imaging agent.

BACKGROUND: BMS-747158-02 is a fluorine 18-labeled pyridaben derivative designed as a new myocardial perfusion imaging agent for use with positron emission tomography (PET). This study evaluated BMS-747158-02 in animal models of cardiac perfusion and compared it with established single photon emission computed tomography agents. METHODS AND RESULTS: In a rat biodistribution study, BMS-747158-02 (15 microCi) had substantially higher myocardial uptake than technetium 99m sestamibi (100 microCi) at 15 minutes (3.5% +/- 0.3% %ID/g vs 1.9% +/- 0.1% %ID/g) and 120 minutes (3.2% +/- 0.4% of injected dose per gram vs 1.8% +/- 0.0% of injected dose per gram) after intravenous administration. Uptake ratios of heart to lung and liver at 60 minutes were also higher for BMS-747158-02 (12.7 +/- 1.4 and 3.7 +/- 0.2, respectively) than Tc-99m sestamibi (5.9 +/- 0.5 and 2.4 +/- 0.4, respectively). In an isolated rabbit heart model at flow rates of 1.66 to 5.06 mL x min(-1).g(-1) wet left ventricular weight, the net BMS-747158-02 heart uptake increased proportionally (0.93 +/- 0.15 to 2.44 +/- 0.40 mL.min(-1) x g(-1)) and to a greater extent than that of thallium 201 (0.76 +/- 0.02 to 1.11 +/- 0.02 mL x min(-1) x g(-1)) or Tc-99m sestamibi (0.49 +/- 0.03 to 0.77 +/- 0.08 mL x min(-1) x g(-1)). PET imaging with BMS-747158-02 showed a clear and sustained cardiac uptake in rats, rabbits, and nonhuman primates with minimal lung interference and rapid liver clearance. Myocardial perfusion deficit zones created by either permanent left coronary ligation or reperfusion after ligation in rats were both clearly identified on PET cardiac images of BMS-747158-02 and had good agreement with in vitro histology. CONCLUSIONS: BMS-747158-02 exhibited high and sustained cardiac uptake that was proportional to blood flow, and it represents a new class of PET myocardial perfusion imaging agent.

Amino Acid derived enamides: synthesis and aminopeptidase activity.

Recently developed copper-catalyzed coupling methodology has been applied to the synthesis of amino acid derived enamides. Bond formation proved to be strongly influenced by protection strategy and vinyl iodide substitution while tolerant of limited side chain functionality. Assessment of aminopeptidase activity revealed a preference for (E)-1,2-disubstituted constructs.

Synthesis and biological evaluation of the mitochondrial complex 1 inhibitor 2-[4-(4-fluorobutyl)benzylsulfanyl]-3-methylchromene-4-one as a potential cardiac positron emission tomography tracer.

A series of fluorinated chromone analogs with IC50 values ranging from 9 to 133 nM for the mitochondrial complex 1 (MC-I) has been prepared. A structure-activity relationship (SAR) study of the most potent fluorinated chromone analog 10 demonstrated the linkage heteroatom preference of the side chain region of the molecule while maintaining potent MC-I inhibitory activity. Tissue distribution studies 30 min after [(18)F]10 administration to Sprague-Dawley (SD) rats demonstrated high uptake of the radiotracer from the blood pool into the myocardium (2.24% ID/g), kidney (1.93% ID/g), and liver (2.00% ID/g). After 2 h about 66% of the activity in the myocardium at 30 min had been retained, whereas approximately 70% had been cleared from the liver and kidney. MicroPET images of SD rats after [(18)F]10 administration allowed easy assessment of the myocardium through 60 min with minimal lung or liver interference.

Quinazoline derivatives as MC-I inhibitors: evaluation of myocardial uptake using Positron Emission Tomography in rat and non-human primate.

Several quinazoline derivatives were made as mitochondrial complex 1 inhibitors. Compound 4 showed an IC(50) of 11.3 nM and was the most potent compound of this series. The (18)F analog of 4, [(18)F] 4, was injected in the rat and showed high and rapid heart uptake, fast liver clearance, and low blood uptake. Images obtained using a microPET showed clear delineation of the myocardium in normal rats and perfusion deficit in ischemic rats. In the non-human primate, [(18)F] 4 showed rapid uptake and clearance from the myocardium and high liver uptake.