Advancing mGlu1 positive allosteric modulators as therapeutics to facilitate abstinence in cocaine use disorder

Project Summary A major challenge in treating cocaine use disorder is that vulnerability to cue-induced craving and relapse persist even after long periods of abstinence. There is presently no FDA-approved medication to lessen cocaine craving. The goal of Eleutheria Pharmaceuticals LLC is to develop metabotropic glutamate receptor 1 (mGlu1) positive allosteric modulators (PAMs) as therapeutic agents to reduce craving and thereby facilitate abstinence in persons with cocaine use disorder. The target, mGlu1, has been thoroughly validated by the lab of the PI and Company Founder, and by other work. Our studies have used the ‘incubation of drug craving’ model, in which cue-induced craving in rats progressively intensifies (‘incubates’) during abstinence and then remains high for months. Incubation of craving also occurs in humans. We showed that incubation depends on strengthening of glutamate synapses in a key brain region for drug craving (nucleus accumbens) via incorporation of atypical high conductance AMPA-type glutamate receptors (CP-AMPARs). mGlu1 PAMs reverse this plasticity by eliciting a form of long-term depression (LTD) that is expressed by CP-AMPAR internalization and, through this mGlu1- LTD, reduce cocaine craving. In 2020, the PI obtained a NIDA U18 grant that has taken our project to the Assay Development stage. In this Phase I STTR application, we propose two Aims that will result in the identification of novel chemotypes and position us to apply for Phase II STTR funding. Classic in vitro screens for mGlu1 activation rely on detecting Ca2+ mobilization in cell lines. We have developed a novel assay to selectively detect activation of the signaling pathway downstream of mGlu1 that mediates craving reduction, namely the rapid translation of the LTD protein oligophrenin-1 (OPHN1). Briefly, HiBiT-tagged OPHN1 (stably transfected into an mGlu1 over-expressing cell line) generates a luminescent signal upon mGlu1 stimulation. Aim 1 will optimize this assay so it can be used in high throughput screening in Aim 2. The assay will be optimized by reduction of baseline signal to achieve an optimal signal window and Z’ ≥ 0.6 as a benchmark for success. Reproducibility at scale will be evaluated during a pilot screen of 1430 unique compounds tested in triplicate at a single dose. In Aim 2, the optimized assay will be used for a quantitative high throughput primary screen of 4382 bioactive compounds at Oregon State University’s High-Throughput Screening Services Laboratory. Hits from this screen and additional compounds chosen for similarity to our mGlu1 PAM tool compound will be sourced fresh for dose curves and cytotoxicity assessment, and medicinal chemistry prioritization conducted. If we are successful in achieving our Phase I goals (optimize and execute the primary screen, identify and validate hit scaffolds), we will submit a Phase II application to assess prioritized compounds in secondary assays, optimize their activity and PK properties during iterative SAR studies, and conduct in vivo testing (‘incubation of craving’ model) to identify lead compounds. To achieve these goals, the PI has assembled a strong team with expertise in drug screening, medicinal chemistry, business development, and the needs of substance use disorder patients.Narrative A major challenge in treating cocaine use disorder is that vulnerability to cue-induced craving and relapse persists even after long periods of abstinence, and there is presently no FDA-approved medication to lessen craving and thereby facilitate abstinence. We have shown, in a translationally relevant rat model, that mGlu1 positive allosteric modulators (PAMs) reduce cocaine craving after prolonged abstinence by reversing cocaine- induced synaptic plasticity. The goals of this proposal are to optimize a novel assay for mGlu1 activation and use it to screen a library of biologically active compounds, setting the stage for future efforts to develop new mGlu1 PAMs as therapeutic agents.