Abstracts

Rationale: A ligand eliciting the same magnitude of response as an endogenous agonist is classified as a full agonist. When a ligand elicits an even greater magnitude of response than the endogenous agonist (i.e., higher receptor output signaling), the compound is classified as a superagonist. Preclinical and clinical data support 5-HT2 receptor agonists being efficacious treatments for a variety of motor seizures and seizure disorders. LP352 is a potent and selective 5-HT2C receptor agonist that was specifically designed to have increased selectivity for the 5-HT2C receptor vs. 5-HT2A and 5-HT2B compared to known serotonergic agonists such as fenfluramine and lorcaserin. LP352 displays a binding affinity (Ki) of 44 nM at the human 5-HT2C receptor and shows neither measurable affinity nor any functional agonism at 5-HT2A and 5-HT2B receptors. LP352 is being developed for the treatment of developmental and epileptic encephalopathies and other refractory epilepsies.

Methods: Human 5-HT2C DMR Assays: Dynamic mass redistribution (DMR) assays were performed using HEK293 cells expressing the human 5-HT2C receptor. Cells were plated in 384-well Corning Epic^® plates and allowed to incubate overnight in serum-free media. On the day of the assay, serially diluted test compounds were applied and DMR responses recorded on a Corning Epic BT reader. Data were analyzed by measuring the change in the DMR response (in picometers) from baseline at a timepoint that produced a maximal response – typically 30 to 60 minutes following compound addition. Rat Choroid Plexus Epithelial Cell Assays: Primary rat choroid plexus epithelial cells, a cell type known to express high levels of endogenous 5-HT2C, were harvested using standard methods and cultured for 24 or 48 hours. Cells were then plated in DMR assay plates or white, 96-well plates for use in classical inositol phosphate (IP) accumulation assays using [³H]myo-inositol. IP accumulation assays provide a more specific assessment of G protein activation by the test compounds. DMR assays were performed in the same manner as the HEK293 assays.

Results: Figure 1 shows dose-responses for lorcaserin, serotonin, and LP352 generated in the DMR assay. As the concentration of LP352 increases, its maximal cellular response exceeds that elicited by both lorcaserin and the endogenous ligand, serotonin. In both IP accumulation and DMR assays performed using primary rat choroid plexus epithelial cells, maximal LP352-induced cellular responses exceeded that of the endogenous ligand serotonin, consistent with classification as a superagonist at 5-HT2C receptors (Figure 2).

Conclusions: LP352 is an example of a 5-HT2C specific superagonist. Further clinical studies should be undertaken to determine if this highly targeted superagonism translates to safety and/or efficacy advantages in disorders likely to benefit from this unique pharmacology.

Funding: Sponsored by Longboard Pharmaceuticals