Development of an In Vivo Method to Estimate Effective Drug Doses and Quantify Fatty Acid Amide Hydrolase in Rodent...

Fatty acid amide hydrolase (FAAH) is a key enzyme in the endocannabinoid system. N-(3,4-Dimethylisoxazol-5-yl)piperazine-4-[4-(2-fluoro-4-[¹¹C]methylphenyl)thiazol-2-yl]-1-carboxamide ([¹¹C]DFMC) was developed as an irreversible-type positron emission tomography (PET) tracer for FAAH. Here, we attempted to noninvasively estimate rate constant k₃ (rate of transfer to the specifically-bound compartment) as a direct index for FAAH in the rat brain. First, the two-tissue compartment model analysis including three parameters [K₁-k₃, two-tissue compartment model for the irreversible-type radiotracer (2TCMi)] in PET study with [¹¹C]DFMC was conducted, which provided 0.21 ± 0.04 ml·cm^-3·min^-1 of the net uptake value (K_i), an indirect index for FAAH, in the FAAH-richest region (the cingulate cortex). Subsequently, to noninvasively estimate K_i value, the reference model analysis (Patlak graphical analysis reference model) was tried using a time-activity curve of the spinal cord. In that result, the noninvasive K_i value (K^REF) was concisely estimated with high correlation (r > 0.95) to K_i values based on 2TCMi. Using estimated K^REF value, we tried to obtain calculated-k₃ based on previously defined equations. The calculated k₃ was successfully estimated with high correlation (r = 0.95) to direct k₃ in 2TCMi. Finally, the dose relationship study using calculated k₃ demonstrated that in vivo ED₅₀ value of [3-(3-carbamoylphenyl)phenyl] N-cyclohexylcarbamate, a major inhibitor of FAAH, was 66.4 µg/kg in rat brain. In conclusion, we proposed the calculated k₃ as an alternative index corresponding to regional FAAH concentrations and suggested that PET with [¹¹C]DFMC enables occupancy study for new pharmaceuticals targeting FAAH. SIGNIFICANCE STATEMENT: In the present study, we proposed calculated k₃ as an alternative index corresponding with fatty acid amide hydrolase concentration. By using calculated k₃, in vivo ED₅₀ of [3-(3-carbamoylphenyl)phenyl] N-cyclohexylcarbamate was successfully estimated to be 66.4 µg/kg for rats. Thus, we demonstrated the pharmacological utility of positron emission tomography with N-(3,4-dimethylisoxazol-5-yl)piperazine-4-[4-(2-fluoro-4-[¹¹C]methylphenyl)thiazol-2-yl]-1-carboxamide.