A novel PU.1 inhibitor for the treatment of Alzheimer’s disease

Abstract Background In 2015, we identified the microglia‐specific transcription factor PU.1, encoded for by SPI1 , as a master regulator of neuroinflammation in a mouse model of Alzheimer’s Disease and in human brain tissue from deceased Alzheimer’s Disease patients 1 . In 2017, a GWAS SNP that delays onset of Alzheimer’s Disease, was shown to reduce PU.1 expression levels 2 . Here, we describe a high‐throughput screen for small molecules that inhibit PU.1 activity. Our goal is to develop novel agents that can rein in the abnormal neuroimmune response associated with Alzheimer’s dementia. Method We used a stable immortalized mouse microglia (BV2) reporter line that expresses luciferase in a PU.1‐dependent manner. We screened a total of 58’100 molecules from 3 separate chemical libraries and obtained 264 hits. We further validated these hits using multiple assays in human stem cell derived microglia. Finally, we administered our top hit (A11) to transgenic mouse models of Alzheimer’s Disease. Result A11 exhibits anti‐inflammatory effects at a concentration of 20 nM in activated microglia, as assessed by RTqPCR and ELISA for IL1β, morphological analyses and lipid accumulation quantification. In mice, A11 preferentially distributed to brain tissues after systemic (intraperitoneal) application and reached a peak concentration at 4 hours. When dosing mice that develop severe neurodegeneration (CK‐p25 transgenic mice), we found a reduction of microglial proliferation, astrogliosis and retention of p25‐positive neurons in the hippocampus. No abnormalities were seen in the treated control littermates. Conclusion Our novel PU.1 inhibitor (A11) shows nanomolar potency and efficacy in multiple assays and cell lines. Mouse experiments demonstrate suitability for systemic application. Administration of A11 in CK‐p25 transgenic mice show an amelioration of neurodegeneration‐associated hallmarks, such as microglial and astrocytic activation and loss of p25‐positive neurons. References: (1) Gjoneska,E, A R Pfenning, H Mathys, G Quon, A Kundaje, L H Tsai and M Kellis. 2015. Conserved epigenomic signals in mice and humans reveal immune basis of Alzheimer’s disease. Nat. 518(7539)365‐9. (2) Huang,K L, E Marcora, A A Pimenova, A F Di Narzo et al. 2017. A common haplotype lowers PU.1 expression in myeloid cells and delays onset of Alzheimer’s disease. Nat Neurosci. 20(8) 1052‐1061.