Synthesis and characterization of novel fluorescent sigma-2 receptor ligands

Sigam-2 (σ₂) receptors are overexpressed in a variety of human and rodent tumors and play a pivotal role in cancer biology. Furthermore, it has proved to be a unique biomarker of cell proliferation in solid tumors. With the aim of providing more potent and reliable tools to investigate the σ₂ receptor biology, we report herein on the synthesis and characterization of novel fluorescent σ₂ receptor ligands designed via an integrated approach by using N-(4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)butyl)-2-(2- [¹⁸F]fluoroethoxy)-5-methylbenzamide ([¹⁸F]ISO-1, the only σ₂ receptor radioligand used in clinical trials up to date) as the lead compound.
All the synthesized compounds possess the pharmacophore (6,7-dimethoxy-1,2,3,4-tetrahdyroxyisoquinoline) and a fluorophore directly connected via various carbon linkers as shown in Figure 1. The clog P values were calculated by On-line Lipophilicity in Virtual Computational Chemistry Laboratory as shown in Table 1 (compounds 2-5 vs SW116 and SW120, the clog P 4.24–5.05 vs 6.74 and 7.29). We measured the excitation/emission wavelength, the quantum yields (QY) and the absorbance in phosphate-buffered saline (PBS). The emission maxima of these compounds are about 450 nm. High QYs were observed for compounds 2–5 with 5.95%, 8.11%, 7.52% and 5.48%, respectively.
The affinities of ligands for the σ₂ and σ₁ receptors were determined with radioligand competition experiments. [^3H]DTG in the presence of 10 μM dextrallorphan was used for the σ2 receptors and (+)-[³H]pentazocine for σ₁ receptors. The results are presented in Table 1. In general, the fluorescent compounds preferred to bind to σ₂ receptors. Compounds 2–5, containing a barbituric acid moiety as electron acceptor, possessed high affinity (10.9–22.3 nM) and subtype selectivity for σ₂ receptors (K_i(σ₁)/K_i(σ₂) = 14–57).
In conclusion, fluorescent σ₂ receptor ligands with high affinity and subtype selectivity have been developed and warrant further evaluation.