Reinvestigation of the Synthesis and Evaluation of [N-methyl-11C]vorozole, a Radiotracer Targeting Cytochrome P450 Aromatase

Abstract

Introduction: We reinvestigated the synthesis of [N-methyl-11C]vorozole, a radiotracer for aromatase, and discovered the presence of an N-methyl isomer which was not removed in the original purification method. Herein we report the preparation and positron emission tomography (PET) studies of pure [N-methyl-11C]vorozole. Methods: Norvorozole was alkylated with [11C]methyl iodide as previously described and also with unlabeled methyl iodide. A high-performance liquid chromatography (HPLC) method was developed to separate the regioisomers. Nuclear magnetic resonance (NMR) spectroscopy (13C and 2D-nuclear Overhauser effect spectroscopy NMR) was used to identify and assign structures to the N-methylated products. Pure [N-methyl-11C]vorozole and the contaminating isomer were compared by PET imaging in the baboon. Results: Methylation of norvorozole resulted in a mixture of isomers (1:1:1 ratio) based on new HPLC analysis using a pentafluorophenylpropyl bonded silica column, in which vorozole coeluted one of its isomers under the original HPLC conditions. Baseline separation of the three labeled isomers was achieved. The N-3 isomer was the contaminant of vorozole, thus correcting the original assignment of isomers. PET studies of pure [N-methyl-11C]vorozole with and without the contaminating N-3 isomer revealed that only [N-methyl-11C]vorozole binds to aromatase. [N-methyl-11C]Vorozole accumulated in all brain regions with highest accumulation in the aromatase-rich amygdala and preoptic area. Accumulation was blocked with vorozole and letrozole consistent with reports of some level of aromatase in many brain regions. Conclusions: The discovery of a contaminating labeled isomer and the development of a method for isolating pure [N-methyl-11C]vorozole combine to provide a new scientific tool for PET studies of the biology of aromatase and for drug research and development.