Tight-binding small molecule carboxylesterase 2 inhibitors reduce intracellular Irinotecan activation

As the primary enzyme responsible for activatable conversion of Irinotecan to SN-38, carboxylesterase 2 is a significant prognostic and predictive biomarker towards Irinotecan-based treatments for pancreatic ductal adenocarcinoma (PDAC). High SN-38 levels stemming from high activity levels of CES2 leads to harmful effects, including life-threatening diarrhea. While alternate strategies have been explored, inhibition of CES2 presents as an effective strategy to directly alter the pharmacokinetics of Irinotecan (CPT-11) conversion to ultimately control the amount of SN-38 produced. To address this, we conducted a high-throughput screen to discovery 18 small molecule inhibitors of CES2. The inhibitors are validated by dose-response and counter-screening and 16 of these inhibitors demonstrate selectivity for CES2. These 16 inhibitors inhibit CES2 in cells, indicating they are cell permeable. Furthermore, all 16 inhibitors showed inhibition of Irinotecan conversion with purified enzyme. The top five inhibitors prohibit cell death mediated by Irinotecan upon pre-incubation in PDAC cells. Three of these inhibitors were determined to display a tight-binding mechanism of action with strong binding affinity