The possibility of a fully automated procedure for radiosynthesis of fluorine-18-labeled fluoromisonidazole using a simplified single, neutral alumina column purification procedure

https://doi.org/10.1016/j.apradiso.2010.04.006Get rights and content

Abstract

A novel fully automated radiosynthesis procedure for [18F]Fluoromisonidazole using a simple alumina cartridge-column for purification instead of conventionally used semi-preparative HPLC was developed. [18F]FMISO was prepared via a one-pot, two-step synthesis procedure using a modified nuclear interface synthesis module. Nucleophilic fluorination of the precursor molecule 1-(2′-nitro-1′-imidazolyl)-2-O-tetrahydropyranyl-3-O-toluenesulphonylpropanediol (NITTP) with no-carrier added [18F]fluoride followed by hydrolysis of the protecting group with 1 M HCl. Purification was carried out using a single neutral alumina cartridge-column instead of semi-preparative HPLC. The maximum overall radiochemical yield obtained was 37.49±1.68% with 10 mg NITTP (n=3, without any decay correction) and the total synthesis time was 40±1 min. The radiochemical purity was greater than 95% and the product was devoid of other chemical impurities including residual aluminum and acetonitrile. The biodistribution study in fibrosarcoma tumor model showed maximum uptake in tumor, 2 h post injection. Finally, PET/CT imaging studies in normal healthy rabbit, showed clear uptake in the organs involved in the metabolic process of MISO. No bone uptake was observed excluding the presence of free [18F]fluoride. The reported method can be easily adapted in any commercial FDG synthesis module.

Introduction

The oxygen-dependent covalent binding of the nitroimidazole, misonidazole (1-(2-nitroimidazolyl)-2-hydroxy-3-methoxy propane, MISO) in cells, multicellular spheroids, and tumors has stimulated interest in using this drug or a congener as an imaging agent for hypoxia in malignant tumors, myocardial infarct, or cerebral ischemia (Rasey et al., 1990, Wiebe, 2004, Hodgkiss, 1998, Startford and Workman, 1998, et al., 1999). In vivo demonstration of hypoxia requires tissue measurements with oxygen-electrodes, but the invasiveness of this technique has limited its application. Therefore, non-invasive assessment of tumor hypoxia with a specific radiotracer, prior to radiation therapy should provide a rational means for selecting patients for treatment with bioreductive drugs and chemical radiosensitizers. In addition, it is possible to differentiate radiation therapy modalities (neutron versus photon) by correlating results with labeled markers of hypoxic cells with tumor response. The potential advantage of neutron over conventional photon radiation is the former’s reduced dependence on oxygenation of the tumor and less variability of cell sensitivity to neutrons around the cell cycle (Yang et al., 1995). Consequently, the radiolabeled analogues of MISO and its derivatives are used as markers of hypoxic tissues (Jerabek et al., 1986; Rasey et al., 1987, Rasey et al., 1996; Koh et al., 1992, Martin et al., 1992, Varagnolo et al., 2000, Grönross et al., 2001, Lehtiö et al., 2003). PET-imaging with [18F]FMISO can help to estimate the oxygenation status of tumors in any part of the body (Iulina Toma-Dasu et al., 2009). The tracer has also been used to study the relative hypoxic volume of tumors during the course of radiation treatment. Recently, improvement in response to treatment with new selective experimental chemotherapy agents has been observed by using [18F]FMISO and PET (Eary and Krohn, 2000). Notwithstanding the disadvantages of [18F]FMISO, which, due its lypophilicity, is retained in the brain and is slowly cleared by the hepatobiliary route leading to high background and, hence, requiring delayed imaging; it is still the most used radiotracer in hypoxia studies in humans and it is in high demand for PET oncology studies (Koh et al., 1992: Valk et al., 1992; Rajendran et al., 2003, Rajendran et al., 2006; Couturier et al., 2004), though there are reports of the use of its more hydrophilic derivative, fluoroerythronitroimidazoze, FETNIM (Yang et al., 1995, Grönross et al., 2001). Additionally, a large number of [18F]-labeled analogues have been developed such as [18F]FETA (Rasey et al., 1999), [18F]EF1 (Kachur et al., 1999), [18F]EF5 (Couturier et al., 2004, Dolbier et al., 2001, Komar et al., 2008, Mahy et al., 2006) and [18F]FAZA (Kumar et al., 2002) for hypoxia imaging, although the data regarding their use in human studies is limited.

The most commonly used route for the synthesis of [18F]FMISO, to date, is the nucleophilic substitution of the tosylate-leaving group by [18F]fluoride on the tetrahydropyranyl-protected precursor 1-(2′-nitro-1′-imidazolyl)-2-O-tetrahydropyranyl-3-O-toluenesulphonylpropanediol (NITTP) followed by the hydrolysis of the protecting group (Kämäräinen et al., 2004, Patt et al., 1999). The synthesis procedure is summarized in Scheme 1. Recently, a fully automated synthesis of [18F]FMISO using HPLC purification with a radiochemical yield more than 60% in a synthesis time of approximately 60 min has been reported (Patt et al., 1999). [18F]FMISO radiosynthesis using Sep-Pak® purification with more than 40% radiochemical yield (without decay correction) in about 40 min has also been reported (Tang et al., 2005). Since HPLC purification is quite complicated for routine operation and significantly adds to the synthesis time, we report a fully automated radiosynthesis procedure of [18F]FMISO through nucleophilic fluorination and employing a neutral alumina cartridge-column purification using a Nuclear Interface synthesis module configured for FDG synthesis. A similar study using a modified Explora FDG4 module is reported by Wang et al. (2009) with a decay corrected yield of 55% within a total synthesis time of 50 min. Systematic investigation of the dependence of radiochemical yield on the amount of precursor initially taken has also been investigated.

Section snippets

Reagents and apparatus

NITTP, FMISO reference standard, 75 mM TBAHCO3 solution, molecular-grade acetonitrile, 10% NaCl, 1 M HCl, 1 M NaH2PO4 buffer, sterile and pyrogen-free water for injection, and pharmaceutical grade ethanol were procured from ABX Advanced Biochemical Compounds, Germany. Fluorine-18 separation cartridge, Chromafix 45-PS-HCO3, was obtained from Marcherey-Nagel, Germany. Aluminum oxide active (neutral, Brockmann grade I–II) for column chromatography was procured from Merck, India. Evacuated 10 mL vials

Synthesis and quality control

Starting from the precursor, NITTP, [18F]FMISO was prepared in a synthesis module configured for [18F]FDG synthesis. Purification was achieved using a single neutral alumina cartridge-column. The radiochemical yield expressed as the percentage of radioactivity finally obtained as [18F]FMISO compared with the starting 18F activity, without decay correction, was 23.40±0.65% (n=3), 37.49±1.68% (n=3), and 34.1±3.2 (n=5) with 5, 10, and 25 mg of NITTP, respectively (Table 1). Hence, the highest yield

Conclusion

We have developed a novel, fully automated radiosynthesis procedure for [18F]FMISO achieving satisfactory chemical and radiochemical purity using single neutral alumina cartridge-column for purification instead of semi-preparative HPLC. Further, we could do this in a general purpose fluorination module configured for FDG synthesis with moderately good yield 37.49±1.68% with 10 mg NITTP, without decay correction, in 40±1 min. The synthesis procedure is fast, reliable, and very similar to [18F]FDG

Acknowledgements

We would like to thank Dr. P.S.Soni, Head, Medical Cyclotron Facility, Board of Radiation and Isotope Technology (BRIT), Department of Atomic Energy, Mumbai for help related to the cyclotron production of 18F. The support from Dr. V. Rangarajan and technologists from Bio-imaging Unit, Tata Memorial Hospital, Mumbai, India for PET/CT imaging of rabbits is sincerely acknowledged. Help from Quality Control and Microbiology Staff of BRIT in determining the Al3+ ion concentration and analyzing the

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