Functional properties and mechanism of action of PPTQ, an allosteric agonist and low nanomolar positive allosteric modulator at GABA A receptors.

The former sedative-hypnotic and recreational drug methaqualone (Quaalude) is a moderately potent, non-selective positive allosteric modulator (PAM) at GABAA receptors (GABAA Rs) (Hammer et al., 2015). In the present study, we have identified a novel methaqualone analog, 2-phenyl-3-(p-tolyl)quinazolin-4(3H)-one (PPTQ), in a screening of 67 analogs at five αβ2 γ2S GABAA R subtypes and delineated its functional properties and mechanism of action at wild-type and mutant GABAA Rs expressed in Xenopus laevis oocytes by two-electrode voltage clamp electrophysiology. PPTQ was found to be an allosteric agonist and a PAM (ago-PAM) at human α1 β2 γ2S and α4 β2 δ GABAA Rs, exhibiting intrinsic activity at micromolar concentrations and potentiating the GABA-evoked signaling through the receptors at concentrations down to the low-nanomolar range. Whereas PPTQ exclusively increased the potency of GABA at the α1 β2 γ2S receptor, it increased both GABA potency and efficacy at α4 β2 δ and displayed modest potency-based preference for α4 β2 δ over α1 β2 γ2S . In elaborate mutagenesis and competition experiments PPTQ was found to act through the same or an overlapping site as etomidate in the transmembrane β(+) /α(-) subunit interfaces, whereas it did not seem to target the other three transmembrane interfaces in the GABAA R. Finally, the PPTQ site was shown to be allosterically linked with sites targeted by neurosteroids and barbiturates but not with the high-affinity benzodiazepine site in the α1 β2 γ2S receptor. In conclusion, the development of a highly potent, bioavailable GABAA R ago-PAM by subtle modifications to the methaqualone scaffold demonstrates that derivatization of this infamous drug from the past can lead to modulators with distinct functional characteristics at the receptors.