Psychostimulant abuse and vulnerability to dependence (addiction) and relapse during abstinence linger as major public health problems in the United States and effective, safe pharmacotherapeutic approaches are needed. The 5-HT2CR receptor (5-HT2CR) has been identified as an important mediator of the behavioral effects of psychostimulants and selective 5-HT2CR agonists have shown efficacy in suppression of the discriminative stimulus effects of stimulants as well as drug-taking and drug-seeking in self-administration models. Thus, strategies to maximize signaling through the 5-HT2CR may prove therapeutically useful to inhibit relapse in stimulant abusers. The recent discovery that the 3rd intracellular loop of the 5-HT2CR binds protein phosphatase and tensin homologue deleted on chromosome 10 to control activation states of the receptor provides a new medication development opportunity. The 5- HT2CR:PTEN complex is disrupted by a small peptide fragment of the 5-HT2CR which evokes 5- HT2CR agonist-like properties in vivo with a limited side effect profile, suggesting that either brain-penetrant peptides or small molecules that inhibit the 5-HT2CR:PTEN association will be a novel method to enhance 5-HT2CR function. In the present proposal, submitted in response to RFA-DA-10-018, we will examine the 5-HT2CR:PTEN association as a novel pharmacological target using a multidisciplinary approach involving chemistry, molecular modeling, live cell assays in vitro and preclinical models of addiction in vivo. We will design and synthesize new peptide and peptidomimetic inhibitors of the 5-HT2CR:PTEN interaction based on the modeled peptide interface between the 5-HT2CR and PTEN, and convert these into small molecules optimized for high activity and drug-like properties. These novel inhibitors will be evaluated in live cell assays for their efficacy to disrupt the 5-HT2CR:PTEN complex and enhance signaling through the 5-HT2CR. The potency, efficacy and selectivity of optimized inhibitors of the 5- HT2CR:PTEN interaction will be assessed in vivo in using rat drug discrimination models. The most promising candidates will be evaluated in cocaine self-administration and reinstatement assays. The proposed studies will provide novel peptides, small molecules and cellular assays to probe the neurobiological function of the 5-HT2CR macromolecular complex and potentially discover novel therapeutic agents for the treatment of psychostimulant addiction in humans.