Preparation and biodistribution of [18F]FP2OP as myocardial perfusion imaging agent for positron emission tomography

doi:10.1016/j.bmc.2009.12.022

Bioorganic & Medicinal Chemistry

Volume 18, Issue 3, 1 February 2010, Pages 1312-1320

https://doi.org/10.1016/j.bmc.2009.12.022 Get rights and content

Abstract

Myocardial extractions of pyridaben, a mitochondrial complex I (MC-I) inhibitor, is well correlated with blood flow. Based on the synthesis and characterization of pyridaben analogue 2-tert-butyl-5-[2-(2-[¹⁸F]fluroethoxy)ethoxy]benzyloxy]-4-chloro-2H-pyridazin-3-one ([¹⁸F]FP2OP), this study assessed its potential to be developed as myocardial perfusion imaging (MPI) agent.

Methods: The tosylate labeling precursor 2-(2-(4-(tert-butyl-5-chloro-6-oxo-1,6-dihydro-pyridazin-4-yloxymethyl)benzyloxy)ethoxy)ethyl ester (OTs-P2OP) and the nonradioactive 2-tert-butyl-5-[2-(2-[¹⁹F]fluroethoxy)ethoxy]benzyloxy]-4-chloro-2H-pyridazin-3-one ([¹⁹F]FP2OP) were synthesized and characterized by IR, ¹H NMR, ¹³C NMR and MS analysis. By substituting tosyl of precursor OTs-P2OP with ¹⁸F, the radiolabeled complex [¹⁸F]FP2OP was prepared and further evaluated for its in vitro physicochemical properties, in vivo biodistribution, the metabolic stability in mice, ex vivo autoradiography and cardiac PET/CT imaging.

Results: Starting with [¹⁸F]F⁻ Kryptofix 2.2.2./K₂CO₃ solution, the total reaction time for [¹⁸F]FP2OP was about 100 min, with final high-performance liquid chromatography purification included. Typical decay-corrected radiochemical yield stayed at 41 ± 5.3%, the radiochemical purity, 98% or more. Biodistribution in mice showed that the heart uptake of [¹⁸F]FP2OP was 41.90 ± 4.52%ID/g at 2 min post-injection time, when the ratio of heart/liver, heart/lung and heart/blood reached 6.83, 9.49 and 35.74, respectively. Lipophilic molecule was further produced by metabolized [¹⁸F]FP2OP in blood and urine at 30 min. Ex vivo autoradiography demonstrates that [¹⁸F]FP2OP may have high affinity with MC-I and that can be blocked by [¹⁹F]FP2OP or rotenone (a known MC-I inhibitor). Cardiac PET images were obtained in a Chinese mini-swine at 5, 15, 30 and 60 min post-injection time with high quality.

Conclusion: [¹⁸F]FP2OP was synthesized with high radiochemical yield. The promising biological properties of [¹⁸F]FP2OP suggest high potential as MPI agent for positron emission tomography in the future.

Graphical abstract

A pyridaben analogue as mitochondria complex I inhibitor was synthesized and radiolabeled with ¹⁸F ([¹⁸F]FP2OP) by one-step synthesis, then it was evaluated as a potential PET myocardial perfusion imaging agent.

Introduction

Coronary artery disease (CAD) is the leading cause of death in developed countries and is a serious health problem worldwide. Myocardial perfusion imaging (MPI), an aid in clinical diagnosis, plays an important role in noninvasive measurement of CAD. Currently, MPI is performed with six agents in clinic use: [²⁰¹Tl]TlCl, ^99mTc-sestamibi, ^99mTc-tetrofosmin, [⁸²Rb]RbCl, [¹³N]NH₃, and [¹⁵O]H₂O.¹ The first three of these tracers being single-photon-emitting agents, which serve as the mainstay of myocardial perfusion imaging (MPI) tests.² However, positron emission tomography (PET) imaging provides certain advantages versus single-photon emission computed tomography (SPECT), including higher spatial resolution, improved attenuation correction, and the capability to perform quantitative measurements at the peak of stress. Given this situation, there is a well-recognized need for the development of new perfusion tracers with more ideal properties for PET imaging. Cardiac PET, accordingly, has an opportunity to become the standard for evaluation of myocardial perfusion in coming years.³

Mitochondria, known as the powerhouse of the cell, are abundant in tissues with large energy requirements, such as the heart, where they take up 20–30% of the myocardial intracellular volume.⁴ Therefore, a couple of MPI agents, such as ¹⁸FTPP, [¹⁸F]FBnTP, BMS-747158–02, take mitochondria as their target.2, 5, 6 Four electron transport complexes are located in the inner mitochondrial membrane, of which MC-I is the first enzyme. Recent studies showed that ¹⁸F- or ¹²⁵I-labeled MC-I inhibitors was highly correlated with flow.7, 8, 9 Accordingly, we designed a new [¹⁸F]-labeled tracer based on pyridaben, one of MC-I inhibitors, for myocardial perfusion imaging.

Here we report the synthesis and characterization of pyridaben analogue 2-tert-butyl-5-[2-(2-[¹⁸F]fluroethoxy)ethoxy]benzyloxy]-4-chloro-2H-pyridazin-3-one ([¹⁸F]FP2OP) as a potential myocardial perfusion imaging (MPI) agent.

Section snippets

Materials

[¹⁸F]F⁻ was obtained from PET Center of Xuanwu Hospital (Beijing, China). No-carrier-added [¹⁸F]F⁻ was trapped on a QMA cartridges and eluted with 0.3 mL K₂CO₃ solution (10 mg/mL in H₂O) combined with 1 mL Kryptofix2.2.2. solution (Sigma–Aldrich) (13 mg/mL in acetonitrile).10, 11 Instant lyophilized Kit for ^99mTc-MIBI (MIBI = 2-methoxyisobutylisonitrile) preparation was donated by Beijing SHIHONG Pharmaceutical Co., Ltd (Beijing, China). ^99mTc-MIBI injection was prepared under Kit instruction.

Chemistry

The precursor OTs-P2OP was synthesized according to Figure 1 with good yield and characterized by ¹H NMR, ESI-MS and IR. The preparation procedure of nonradioactive [¹⁹F]FP2OP is also shown in Figure 1. The structure of the desired nonradioactive compound was confirmed by ¹H NMR, ¹³C NMR, ¹⁹F NMR, ESI-MS, and IR analysis. The HPLC chromatogram shows that [¹⁹F]FP2OP has a very good chemical purity (>98% at λ = 254 nm), with a retention time of 21.7 min (Fig. 2A). Based on these results mentioned

Discussion

In the early 1990s, several novel ^99mTc-labeled perfusion tracers (^99mTc-MIBI, ^99mTc-tetrofosmin and ^99mTcN-NOEt) had been developed for SPECT myocardial perfusion imaging. Among those agents, ^99mTc-MIBI has been accepted worldwide for evaluation of CAD with SPECT imaging. Thence forwards, several new MIBI complexes (^99mTcN-MIBI¹⁹ and ^99mTc(CO)₃(MIBI)₃²⁰) with different ^99mTc core have been reported for myocardial perfusion imaging. All of them have improved biodistribution properties in the

Conclusion

A novel ¹⁸F-labeled compound [¹⁸F]FP2OP has been successfully prepared in high yield and high radiochemical purity (>98%). Based on the results of the experiments reported here comes a conclusion that [¹⁸F]FP2OP is a lipophilic and neutral complex. The biodistribution studies in mice shows that this compound has a significant high heart uptake, and good heart/liver, heart/lung, heart/blood ratios. Cardiac PET imaging in a healthy Chinese mini-swine shows clear outline of myocardium at all time

Acknowledgments

This work was supported by the National Natural Science Foundation of China (20871020) and Beijing Natural Science Foundation (2092018). The authors wish to acknowledge the Beijing SHIHONG Pharmaceutical Co, Ltd for the donation of MIBI Kits. Dr. Yuanqun Jiang is acknowledged for her useful comments and suggestions on the language of our manuscript.

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Then, the partition coefficient, which is expressed as log P, was calculated. The in vitro stability was measured according to a previously published method [16]. Briefly, the radiolabeled compound was incubated in physiological saline at room temperature for 1, 2, and 4 h. To determine its serum stability, the radiotracer was incubated in mice serum at 37 °C for 4 h. To remove plasma proteins from the serum, 1 mL of acetonitrile was added, and the sample was centrifuged.
A novel radiotracer 1‑(2‑(2‑(2‑[¹⁸F]fluoroethoxy)ethoxy)ethyl)‑1H‑1,2,3‑triazole‑estradiol ([¹⁸F]FETE) was successfully synthesized, characterized and evaluated in mice for estrogen receptor (ER)-positive breast cancer targeting with positron emission tomography (PET) imaging.
The tosylate precursor 3 was radiolabeled with ¹⁸F and then reacted with 17α‑ethinyl‑estradiol to produce the final [¹⁸F]FETE. The physicochemical properties of [¹⁸F]FETE were tested in vitro, including determination of the octanol/water partition coefficient, stability and cellular uptake in MCF-7 (ER-positive) and MDA-MB-231 (ER-negative) cells. An ex vivo biodistribution study was performed in normal Sprague Dawley rats, and in vivo microPET imaging was performed on MCF-7 and MDA-MB-231 tumor-bearing mice. The results of biodistribution and PET imaging of [¹⁸F]FETE were compared with that of known 16α‑[¹⁸F]fuoro‑17β‑estradiol ([¹⁸F]FES). Radiation dose estimates for [¹⁸F]FETE were also analyzed.
[¹⁸F]FETE was obtained in high radiochemical yield (46.59 ± 8.06%) with high radiochemical purity (>99%) after HPLC purification and high molar activity (15.45 ± 3.15 GBq/μmol). [¹⁸F]FETE is a moderate lipophilic compound with good in vitro stability and the total synthesis time was 55 to 65 min. In biodistribution studies, [¹⁸F]FETE showed high uptake in the ER-abundant uterine tissue of normal immature SD rats (8.55 ± 1.21 and 6.83 ± 1.70%ID/g at 1 h after intravenous and intraperitoneal injection, respectively), and could be blocked with estradiol effectively (the uterus uptake was decreased to 0.63 ± 0.35%ID/g at 1 h after iv injection). MicroPET imaging of tumor-bearing mice with [¹⁸F]FETE at 1 h after iv injection revealed considerable uptake in ER-positive MCF-7 tumors (4.63 ± 0.73%ID/g) that could be inhibited (1.47 ± 0.29%ID/g) and low uptake in ER-negative MDA-MB-231 tumors (1.97 ± 0.36%ID/g). [¹⁸F]FES has relatively low uptake in ER-positive tumor (0.24 ± 0.19%ID/g) when compared with [¹⁸F]FETE. The adult female effective radiation dose of [¹⁸F]FETE in mice was estimated as 0.0022 mSv/MBq.
A novel 17α‑ethinyl‑estradiol-based ER probe [¹⁸F]FETE was developed with high molar activity and good in vitro stability. Based on the results of bio-evaluation in normal immature rats and tumor-bearing mice, it might be a promising candidate for specific PET imaging of ER-positive breast cancer.
18F-labeled rhodamines as potential myocardial perfusion agents: Comparison of pharmacokinetic properties of several rhodamines
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An additional advantage of 18F is that the 110 min half-life is long enough to allow centralized production of 18F-labeled radiopharmaceuticals and distribution to clinical centers without an on-site cyclotron, while still being short enough to allow repeated MPI studies of a patient on the same day. In recent years, several 18F-labeled compounds have been evaluated as possible myocardial perfusion agents including quaternary ammonium salts [3]; tetraphenylphosphonium compounds [4–7]; rotenone derivatives [8,9]; and pyridazinone analogs, such as BMS-747158-02 (Flurpiridaz F18; Lantheus Medical Imaging, Inc.) [10–17]. Flurpiridaz is currently in Phase 3 clinical trials [18–22], and BFPET (Fluoropharma) has recently completed phase 1 clinical trials [7,23].
We recently reported the development of the [¹⁸F]fluorodiethylene glycol ester of rhodamine B as a potential positron emission tomography (PET) tracer for myocardial perfusion imaging (MPI). This compound was developed by optimizing the ester moiety on the rhodamine B core, and its pharmacokinetic properties were found to be superior to those of the prototype ethyl ester. The goal of the present study was to optimize the rhodamine core while retaining the fluorodiethyleneglycol ester prosthetic group.
A series of different rhodamine cores (rhodamine 6G, rhodamine 101, and tetramethylrhodamine) were labeled with ¹⁸F using the corresponding rhodamine lactones as the precursors and [¹⁸F]fluorodiethylene glycol ester as the prosthetic group. The compounds were purified by semipreparative HPLC, and their biodistribution was measured in rats. Additionally, the uptake of the compounds was evaluated in isolated rat cardiomyocytes.
As was the case with the different prosthetic groups, we found that the rhodamine core has a significant effect on the in vitro and in vivo properties of this series of compounds. Of the rhodamines evaluated to date, the pharmacologic properties of the ¹⁸F-labeled diethylene glycol ester of rhodamine 6G are superior to those of the ¹⁸F-labeled diethylene glycol esters of rhodamine B, rhodamine 101, and tetramethylrhodamine. As with ¹⁸F-labeled rhodamine B, [¹⁸F]rhodamine 6G was observed to localize in the mitochondria of isolated rat cardiomyocytes.
Based on these results, the ¹⁸F-labeled diethylene glycol ester of rhodamine 6G is the most promising potential PET MPI radiopharmaceutical of those that have evaluated to date, and we are now preparing to carry out first-in-human clinical studies with this compound.
Characterization of the four isomers of 123I-CMICE-013: A potential SPECT myocardial perfusion imaging agent
2014, Bioorganic and Medicinal Chemistry
Citation Excerpt :
Mitochondria are abundant in high energy consuming tissues such as in the heart, where they comprise 20–30% of the myocardial intracellular volume.31 Three well-known potent MC-I inhibitors are rotenone,16–19,22,32 fenazaquin,21 and pyridaben,20,23–30 and have been radiolabeled with 99mTc, 125I/123I and 18F. The 18F pyridaben analog (18F-BMS-747158-02) is the most extensively studied tracer with promising results in preclinical studies using small and large animal models and on-going clinical studies.20,23–27
Myocardial perfusion imaging (MPI) with single photon emission computed tomography (SPECT) is widely used in the assessment of coronary artery disease (CAD). We have developed ¹²³I-CMICE-013 based on rotenone, a mitochondrial complex I (MC-1) inhibitor, as a promising new MPI agent. Our synthesis results in a mixture of four species of ¹²³I-CMICE-013 A, B, C, D. In this study, we separated the four species and evaluated their biodistribution and imaging properties. The cold analogs ¹²⁷I-CMICE-013 A, B, C, D were isolated and characterized and their chemical structures proposed. Methods: ¹²³I-CMICE-013 was synthesized by radiolabeling rotenone with Na¹²³I in trifluoroacetic acid (TFA) with iodogen as the oxidizing agent at 60 °C for 45 min, and the four species were separated by RP-HPLC. The cold analogs ¹²⁷I-CMICE-013 A, B, C and D were isolated with a similar procedure and characterized by NMR and mass spectrometry. Biodistribution and microSPECT imaging studies were carried out on normal rats. Results: We propose the mechanism of the rotenone iodination and the structures of the four species. First, I⁺ forms an intermediate three-membered ring with 6′ and 7′ carbons. Second, the lone electron pair of the water molecule attacks the 6′ or 7′-carbon, following by the formation of 6′-OH, and 7′-I bonds as in major products C and D, or 6′-I and 7′-OH bonds as in minor products A and B. The weaker 6′-I bond in the intermediate prompts the nucleophilic attachment of water at the favorable 6′-carbon to generate C and D. MicroSPECT images of ¹²³I-CMICE-013 A, B, C, D in rats showed clear visualization of myocardium and little interference from lung and liver. The imaging time activity curves and biodistribution data showed complex profiles for the four isomers, which is not expected from the structure activity relationship theory. Conclusion: ^123/127I-CMICE-013 A and B are constitutional isomers with C and D, while A and C are diastereomers of B and D, respectively. Overall, the biological characteristics of the four species are not correlated perfectly with their molecular structures.
Synthesis and 18F-labeling of the analogues of Omecamtiv Mecarbil as a potential cardiac myosin imaging agent with PET
2013, Nuclear Medicine and Biology
Cardiac myosin is a potential molecular target for heart failure imaging since its changes can be used to assess the function of heart. In this study, two analogues of Omecamtiv Mecarbil, which is the first selective activator of cardiac myosin, were synthesized and radio-labeled with ¹⁸F. Then the radio-compounds were evaluated as potential cardiac myosin imaging agent.
The labeling precursor and the nonradioactive compounds were synthesized and characterized by IR, ¹H NMR, ¹³C NMR and MS analysis. By substituting bromo of precursors with ¹⁸F, the radiolabeled compounds [¹⁸F]8 and [¹⁸F]10 were prepared and further evaluated for their in vitro physicochemical properties, stabilities, protein binding assay and ex vivo biodistribution.
Starting with [¹⁸F]F^- Kryptofix 2.2.2./K₂CO₃ solution, the total reaction time for [¹⁸F]8 and [¹⁸F]10 was about 40 min respectively, with final high-performance liquid chromatography purification included. Typical decay-corrected radiochemical yield stayed at 12.47% ± 3.30% (n = 8), the radiochemical purity, 98% or more. Their specific activity was estimated as 50 GBq/μmol. Both [¹⁸F]8 and [¹⁸F]10 could be stable after incubation in water at room temperature and in serum or binding buffer at 37 °C for 3 h. Biodistribution in normal mice showed that both [¹⁸F]8 and [¹⁸F]10 have good heart uptake at 2 min post-injection time. Compound [¹⁸F]10 has better heart retention and higher heart to background ratios than those of [¹⁸F]8. In vitro protein binding assay demonstrates that [¹⁸F]10 may have high affinity with myosin from bovine heart.
[¹⁸F]8 and [¹⁸F]10 were synthesized with good radiochemical yield and high radiochemical purity (> 98%). One of the compounds ([¹⁸F]10) has higher bovine heart myosin binding affinity and better heart/liver ratio. It will be further evaluated as a potent cardiac myosin imaging agent in normal and systolic heart failure model with positron emission tomography in the future.
Synthesis and characterization of 123I-CMICE-013: A potential SPECT myocardial perfusion imaging agent
2013, Bioorganic and Medicinal Chemistry
Citation Excerpt :
Preliminary phase I clinical study findings about the safety and biodistribution profile of 18F-BMS-747158-02 illustrate its potential in myocardial perfusion PET imaging showing favorable myocardial to background ratios.17 Another group of pyridaben based 18F MPI tracers, 18F-FP1OP, 18F-FP2OP, and 18F-FP3OP has also been developed recently.18,19 These lipophilic and neutral compounds show significant heart uptake and good heart-to-nontarget ratios in mice.
Coronary artery disease (CAD) is a major cause of death in Canada and the United States. Single photon emission computed tomography (SPECT) myocardial perfusion imaging (MPI) is a useful diagnostic test in the management of patients with CAD. The widely used SPECT MPI agents, ^99mTc sestamibi and ^99mTc tetrofosmin, exhibit less than ideal pharmacokinetic properties with decreasing uptake with higher flows. ¹²³I has a similar energy as ^99mTc, an ideal half life, and is readily available from cyclotrons. The objective of this study was to develop an ¹²³I labeled MPI agent based on rotenone, a mitochondrial complex I inhibitor, as an alternative to currently available SPECT MPI agents. Methods: ¹²³I-CMICE-013 was synthesized by radiolabeling rotenone with ¹²³I in trifluoroacetic acid (TFA) with iodogen as the oxidizing agent at 60 °C for 45 min, followed by RP-HPLC purification. The product was formulated in 5% EtOH in 10 mM NaOAc pH 6.5. The inactive analog ¹²⁷I-CMICE-013 was isolated and characterized by NMR and mass spectrometry, and the structure determined. Micro-SPECT imaging studies were carried out in normal and infarcted rats. Biodistribution studies were performed in normal rats at 2 h (n = 6) and 24 h (n = 8) post injection (p.i.). Results: ¹²³I-CMICE-013 was isolated with >95% radiochemical purity and high specific activity (14.8–111 GBq/μmol; 400–3000 mCi/μmol). Structural analysis showed that rotenone was iodinated at 7′-position, with removal of the 6′,7′-double bond, and addition of a hydroxy group at 6′-position. MicroSPECT images in normal rats demonstrated homogeneous and sustained myocardial uptake with minimal interference from lung and liver. Absent myocardial perfusion was clearly identified in rats with permanent left coronary artery ligation and ischemia-reperfusion injury. In vivo biodistribution studies in normal rats at 2 h p.i. showed significant myocardial uptake (2.01 ± 0.48%ID/g) and high heart to liver (2.98 ± 0.93), heart to lung (4.11 ± 1.04) and heart to blood (8.37 ± 3.97) ratios. At 24 h p.i., the majority of ¹²³I-CMICE-013 was cleared from tissues, and a significant amount of tracer was found in the thyroid, indicating in vivo deiodination of the tracer. Conclusion: ¹²³I-CMICE-013 is a promising new radiotracer for SPECT MPI with high myocardial uptake, very good target to background ratios and favorable biodistribution characteristics.
2-tert-Butyl-4-chloro-5-[4-(2-fluoroethoxy)benzyloxy]pyridazin-3(2H)-one
2012, Acta Crystallographica Section E: Structure Reports Online

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Bioorganic & Medicinal Chemistry

Abstract

Graphical abstract

Introduction

Section snippets

Materials

Chemistry

Discussion

Conclusion

Acknowledgments

References and notes (23)

Curr. Probl. Cardiol.

Am. J. Cardiol.

J. Nucl. Cardiol.

Bioorg. Med. Chem.

J. Nucl. Med.

Hebei Chem. Eng. Ind.

Tetrahedron

Appl. Radiat. Isot.

Appl. Radiat. Isot.

J. Label. Compd. Radiopharm.

J. Nucl. Cardiol.

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<sup>18</sup> F–labeled estradiol derivative for targeting estrogen receptor-expressing breast cancer

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Synthesis and <sup>18</sup>F-labeling of the analogues of Omecamtiv Mecarbil as a potential cardiac myosin imaging agent with PET

Synthesis and characterization of <sup>123</sup>I-CMICE-013: A potential SPECT myocardial perfusion imaging agent

2-tert-Butyl-4-chloro-5-[4-(2-fluoroethoxy)benzyloxy]pyridazin-3(2H)-one