Synthesis and in vivo evaluation of F-18-labeled isoxazole analog for imaging of sigma-2 receptor status

Objectives: Sigma (σ) receptors are known to be overexpressed in most of solid tumors and differentiate proliferative cells from quiescent cells by density of population. Since expression of σ₂ are 8-10 times higher than σ₁ receptors, identification of σ₂ receptors status is helpful for the appropriate cancer treatment. In here, we synthesized F-18-labeled 5-(4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)butyl)-3-(4-fluorophenyl)isoxazole([¹⁸F]SL-2) and evaluated PET and biodistribution for σ₂ receptors status in EMT-6 tumor Xenograft BALB/c mice. Methods: Precursor was prepared from 4-BPin benzaldehyde via a 5 step synthesis. Fluorine-18 radiofluorination was performed at 110ºC for 20 min using copper catalyst and 4-DMAP (eluent and base) in a Synthera module. Purification was performed using preparative HPLC and the purified tracer was analyzed by analytical HPLC. Dynamic PET images of EMT-6 Xenograft BALB/c mice were obtained for 120 min (female, n=1) and Static PET images were obtained at 60-80 min (female, n=4) after intravenous injection of [¹⁸F]SL-2 (3.8±0.2 MBq). In vivo biodistribution was performed at 30 and 60 min post-injection of tracer (17.5 kBq) in EMT-6 Xenograft BALB/c mice (female, n=4) Results: Non-radioactive SL-2 showed high affinity of σ₂ receptors (0.05 nM) vs σ₁ receptors (769 nM). Total radiosynthesis time was 70 min including HPLC purification. Radiochemical yield was 2-5% (decay corrected to EOB). Radiochemical purity and molar activity were 98% and 1.65 Ci/µmol, respectively. In PET studies on Fig 1, the uptake of [¹⁸F]SL-2 accumulated slowly in tumor and showed relatively high uptake on the periphery (2.70±0.47 %ID/cc) than tumor core (1.47±0.77 %ID/cc). Tumor to muscle ratio was 1.2. In biodistribution studies (Fig 2), liver and pancreas showed high uptake. Tumor uptake was 0.86±0.22 %ID/g at 30 min and 1.17±0.29 %ID/g at 60 min, which was consistent with the PET data. Tumor : blood ratio was 2.14±0.79 and 3.79±3.52 and Tumor : muscle ratio was 0.89±0.17 and 1.86±1.44 at 30 min and 60 min, respectively. Conclusions: [¹⁸F]SL-2 was synthesized by copper mediated radiofluorination in automation module which was applicable for human and non-human research. In vitro binding assay suggested that SL-2 is a selective σ₂ receptors ligand with high affinity. In vivo studies demonstrated [¹⁸F]SL-2 may differentiate proliferative cells from quiescent cells by measuring σ₂ receptors status.