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  • While several C terminal peptides


    While several C-terminal peptides have been reported to have modest (astressin) or substantial (astressin2-B, antisauvagine-30) subtype selectivity for the CRF2 receptor (), [125I]YP20 is the first reported small analog of the C-terminus of CRF exhibiting subtype selectivity for the CRF1 receptor vs. CRFR2α receptor. With respect to the nature of the peptide binding site on the ECDs of class B G protein-coupled receptors, NMR chemical-shift perturbation experiments have revealed detailed structural information about the binding interface of astressin with the isolated soluble ECD of the CRF2β receptor (). While comparable structural detail has not been reported for the CRF1 receptor, a pharmacophore model based on NMR spectroscopic studies with isolated soluble CRF1receptor ECD predicted that ECD binding site was relatively long (≈15Ǻ), suggesting that small molecule antagonists may have difficulty blocking large peptide ligands (particularly agonists which bind via the two-site mechanism) from binding to the holo-receptor (). Radioligand selection may be the critical factor for detecting CRF1 receptor antagonists that bind to the ECD. Compounds with this site of action could be useful starting points for developing therapeutic agents with improved physical properties over J-site antagonists. However, it remains to be seen whether non-peptidic ECD antagonists effective at blocking CRF\'s function and adhering to the rule of five for CNS drugs (Lipinski et al., 1997) can be identified from screening small molecule Bax channel blocker collections. Although [125I]astressin could also be utilized as a radioligand for screening compound collections for small molecule ECD-specific antagonists of the CRF1 receptor, the complete lack inhibition of [125I]YP20 binding by J-site antagonists is a key advantage of [125I]YP20 for identifying ECD-specific antagonists. Limitations of the radioligand filtration binding technique for screening compound collections include the moderate number of compounds that can be screened per assay due to the filter washing step, the short (60days) half-life of 125I, lack of suitability for multiplexing, and the high costs of monitoring and disposing of radioactivity. However, the [125I]YP20 binding assay can be further exploited to develop and cross validate alternative ECD binding assays that lack these limitations. If a fluorescently tagged analog of YP20 retaining high affinity for the ECD can be identified, techniques compatible with homogenous assay formats, such as fluorescence polarization or time-resolved fluorescence, could be utilized (). For example, a high through put screening assay for the β2 adrenergic receptor was recently described using the time-resolved fluorescence technique of QRET (quenching resonance energy transfer) (). In summary, [125I]YP20 is the first ECD-specific radioligand for the CRF1 receptor and is thus an important new tool for probing drug interactions at this pharmacologically important drug target.
    Acknowledgement All financial support provided by AstraZeneca, Pharmaceuticals LP.
    Introduction Drug addiction can be considered a multifactorial disorder of chronic relapse as a result of the interaction of biological and environmental factors, characterized by a loss of control over use of the drug (Kalivas and Volkow, 2005, Koob, 2010, Koob and Volkow, 2009, Volkow et al., 2015). It has been repeatedly demonstrated that adverse life experiences can render individuals more prone to addictive substances of abuse and make them more vulnerable to relapse after periods of detoxification (Le Moal, 2009, vington et al., 2008, Sinha et al., 2011). Thus, stress is considered a risk factor that can influence all stages of drug addiction, as it plays a role in initiation, maintenance, escalation of intake and relapse (Burke and Miczek, 2015, Kalivas and Volkow, 2005, Koob, 2010, Sinha et al., 2011, Logrip et al., 2011, Logrip et al., 2012).