Preparation and biological evaluation of 3-[76Br]bromo-α-methyl-l-tyrosine, a novel tyrosine analog for positron emission tomography imaging of tumors

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Abstract

Introduction

3-[18F]fluoro-α-methyl-l-tyrosine ([18F]FAMT) is a useful amino acid tracer for positron emission tomography (PET) imaging of malignant tumors. FAMT analogs labeled with 76Br, a positron emitter with a long half-life (t1/2=16.1 h), could potentially be widely used as amino acid tracers for tumor imaging. In this study, 3-[76Br]bromo-α-methyl-l-tyrosine ([76Br]BAMT) was designed, and its usefulness was evaluated as a novel PET tracer for imaging malignant tumors.

Methods

In this study, both [76Br]BAMT and [77Br]BAMT were prepared. The in vitro and in vivo stability of [77Br]BAMT was evaluated by HPLC analysis. Cellular uptake and retention of [77Br]BAMT and [18F]FAMT were evaluated using LS180 colon adenocarcinoma cells. Biodistribution studies were performed in normal mice and in LS180 tumor-bearing mice, and the tumors were imaged with a small-animal PET scanner.

Results

[77Br]BAMT was stable in vitro but was catabolized after administration in mice. Cellular accumulation and retention of [77Br]BAMT were significantly higher than those of [18F]FAMT. In biodistribution studies, the tumor accumulation of [77Br]BAMT was higher than that of [18F]FAMT. However, some level of debromination was seen, which caused more retention of radioactivity in the blood and organs than was seen with [18F]FAMT. PET imaging with [76Br]BAMT enabled clear visualization of the tumor, and the whole-body image using [76Br]BAMT was similar to that using [18F]FAMT.

Conclusions

[77Br]BAMT showed high levels of tumor accumulation, and [76Br]BAMT enabled clear visualization of the tumor by PET imaging. Although an improvement in stability is still needed, 76Br-labeled FAMT analogs could potentially serve as PET tracers for the imaging of malignant tumors.

Introduction

Radiotracers that enable the visualization of abnormal tumor metabolism are useful tools for tumor diagnosis [1]. Since enhanced glucose metabolism is a well-known property of tumors, 2-[18F]fluoro-2-deoxy-d-glucose ([18F]FDG), a glucose analog labeled with a positron emitter, was developed and has been widely used for tumor imaging with positron emission tomography (PET) [1], [2]. However, high levels of accumulation in nontarget tissues such as the brain and inflammatory sites often impede the accurate diagnosis of tumors [2]. Other tracers that detect abnormal tumor metabolism have also been developed. These alternatives include amino acid tracers, which reflect the enhanced uptake of amino acid caused by excessive protein synthesis in tumors [3]. In contrast to [18F]FDG, amino acid tracers usually show low accumulation in nontarget tissues [1], [3]. At present, several amino acid tracers, including 2-amino-4-([11C]methylthio) butyric acid ([11C]methionine) and 18F-labeled l-tyrosine analogs, have already been introduced into clinical practice and have already been used to clearly visualize malignant tumors in patients with head and neck, lung and brain tumors [1], [3], [4], [5].

3-[18F]fluoro-α-methyl-l-tyrosine ([18F]FAMT), developed as a useful amino acid tracer, is incorporated into tumor cells solely via the amino acid transport system, the activity of which is increased in various human neoplasms [6], [7], [8], [9], [10]. Clinical studies have shown that [18F]FAMT could be used to differentiate between benign lesions and malignant tumors in cases involving brain tumors, lung cancers and lymphomas [11], [12]. In addition, it has been reported that [18F]FAMT has higher specificity than [18F]FDG with regard to the detection of maxillofacial tumors, thoracic tumors and lymph node metastases of non–small cell lung cancer [8], [13], [14]. However, the radiolabeling yield of [18F]FAMT is low because [18F]FAMT is synthesized by the radiolabeling of α-methyl-l-tyrosine with [18F]acetylhypofluoride. In addition, the half-life of 18F (t1/2=110 min) is not long. Thus, [18F]FAMT is not suitable for widespread use.

76Br is an attractive alternative. It is a positron emitter (β+=57%, electron capture=43%) and has a longer half-life (t1/2=16.1 h) than 18F, and substitution of 18F with 76Br would be easy because both bromine and fluorine belong to the halogen group. Several reports have demonstrated that PET imaging with 76Br-labeled tracers is feasible, not only for use in experimental models but also for clinical diagnosis [15], [16], [17]. Furthermore, the process of radiolabeling tyrosine with 76Br- and excess oxidant is easy, and the radiolabeling yield of that process is high [18]. Thus, 76Br use could potentially increase the availability of PET using an α-methyl-l-tyrosine analog in many hospitals. In this study, we designed 3-[76Br]bromo-α-methyl-l-tyrosine ([76Br]BAMT), a 76Br-substitued derivative of [18F]FAMT, and evaluated its usefulness as a novel PET tracer.

Section snippets

Generals

Enriched 76Se (99.67%) was purchased from Isoflex (San Francisco, CA, USA). Cu2Se, α-methyl-l-tyrosine and ethidium bromide were purchased from Sigma-Aldrich (St Louis, MO, USA). Dulbecco's modified Eagle's medium, penicillin, streptomycin and N-chlorosuccinimide were purchased from Wako Pure Chemical Industries (Osaka, Japan). Fetal bovine serum was purchased from Nichirei Biosciences (Tokyo, Japan). Male ddY mice were purchased from Japan SLC Inc. (Hamamatsu, Japan), and male BALB/c (nu/nu)

Radiolabeling

The radiolabeling yield of [76Br]BAMT and [77Br]BAMT was approximately 20%–30%, and radiochemical purity after purification by RP-HPLC (retention time: 14 min) was approximately 99% (Fig. 1). The specific activities of [76Br]BAMT and [77Br]BAMT were over 10 GBq/μmol.

Stability and lipophilicity of [77Br]BAMT

As shown in Fig. 2A, 95% of [77Br]BAMT remained intact over 6 h after incubation in vitro, indicating that [77Br]BAMT was stable in the serum. We further examined the in vivo stability of [77Br]BAMT by analyzing blood samples after

Discussion

Protein metabolism is more enhanced in malignant tumors than in normal tissues, and thus, PET imaging using radiolabeled amino acids is a valuable tool for imaging tumors. Various amino acid tracers, including [18F]FAMT, have been applied for clinical PET studies, and their potential to help visualize tumor lesions is well established [1], [3]. However, it is difficult to widely apply [18F]FAMT-PET because of its low radiolabeling yield as well as the half-life of 18F. Since 76Br is easily

Conclusion

In the present study, [77Br]BAMT showed a similar biodistribution profile to that of [18F]FAMT, and higher uptake into tumors was observed both in vitro and in vivo with [77Br]BAMT. Furthermore, [76Br]BAMT enabled clear visualization of transplanted tumors in mice imaged by PET. On the other hand, a retention of radioactivity in the body caused by debrominated free bromine was observed, which probably increased the overall radiation dose. Although an improvement in stability is still needed, 76

Acknowledgments

This work was supported by a Grant-in-Aid for Young Scientists (A) (22689035) from the Ministry of Health, Labor and Welfare. For operation of the AVF cyclotron, we are grateful to Mr. Hiroyuki Suto and the staff of Takasaki Ion Accelerators for Advanced Radiation Applications. We also would like to thank the staff of the Medical Radioisotope Application Group at the Japan Atomic Energy Agency as well as the Departments of Diagnostic Radiology and Nuclear Medicine at Gunma University Graduate

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