Evaluation of the Anti-Inflammatory Effects of Novel Fatty Acid-Binding Protein 4 Inhibitors in Microglia

Abstract Fatty acid-binding protein 4 (FABP4) is a key lipid binding protein expressed in microglia, which has been demonstrated to play a critical role in microglial-mediated neuroinflammation, a component of many neurodegenerative diseases. Compounds able to inhibit the function of FABP4 have shown promise in reducing microglial-mediated neuroinflammation, however, their physicochemical properties would prevent their ability to be easily formulated and traverse the blood–brain barrier (BBB) in order to access microglial FABP4. To this end, this study assessed the ability of a series of FABP4 inhibitors, with more desirable physicochemical properties, to attenuate microglial inflammation in an in vitro setting. Four inhibitors with varying affinity to FABP4, as measured by isothermal titration calorimetry (MFP-0011462, MFP-0012314, MFP-0012318, and MFP-0012328), were assessed for their ability to induce toxicity and attenuate reactive oxygen species (ROS) generation and tumour necrosis factor-α (TNF-α) release from lipopolysaccharide (LPS)-activated BV-2 microglia. All FABP4 inhibitors were determined to be soluble in the aqueous buffers at the highest concentration used in the assays (100 µM). Isothermal titration calorimetry demonstrated that the compounds had varying affinities for FABP4 (KD values of 316 nM to > 100 µM). The ability of FABP4 inhibitors to reduce LPS-mediated ROS production aligned with their KD for FABP4, with the most effective inhibitor (MFP-0012328) also able to reduce TNF-α production (by RT-qPCR) and TNF-α release from LPS-activated BV-2 cells by 17% and 25%, respectively. These studies have demonstrated that a series of FABP4 inhibitors with more appropriate physicochemical properties for BBB penetration are able to reduce microglial-mediated inflammation, which may be of benefit in diseases where overactivation of microglia leads to neurodegeneration.