Effects of polar κ receptor agonists designed for the periphery on ATP-induced Ca2+ release from keratinocytes

In order to obtain polar κ agonists, which cannot pass the blood–brain barrier, secondary amines 3a and 3b were reductively alkylated with pyridine-3-carbaldehyde to give pyridylmethyl substituted perhydroquinoxalines 5a and 5b, respectively. Both 5a and 5b show very high κ-opioid receptor affinity with Ki values of 0.13 nM and 3.8 nM, respectively. Moreover, they are very selective for the κ receptor. In the [35S]GTPγS assay, both quinoxalines reached full agonistic activity. With an EC50 value of 34 nM, 5a is only slightly less potent than the prototypical κ agonist U-69,593. For the determination of the logD values, a shake-flask method was developed, making use of quantification by mass spectrometry which requires only very small amounts of the compound (<0.8 mg). According to this method, the logD7.4 and logD5.4 values of 5a were 1.1 and −0.45, indicating very high polarity. The logD5.4 value was recorded due the acidic milieu of the skin. Perhydroquinoxalines 3–5 reduced the release of Ca2+ ions into the cytoplasm after stimulation of HaCaT cells with ATP, thereby proving biological activity in human skin cells.