pf-4708671 and Fragile-X-Syndrome

Transcriptional silencing of the FMR1 gene in fragile X syndrome (FXS) leads to the loss of the RNA-binding protein FMRP. In addition to regulating mRNA translation and protein synthesis, emerging evidence suggests that FMRP acts to coordinate proliferation and differentiation during early neural development. However, whether loss of FMRP-mediated translational control is related to impaired cell fate specification in the developing human brain remains unknown. Here, we use human patient induced pluripotent stem cell (iPSC)-derived neural progenitor cells and organoids to model neurogenesis in FXS. We developed a high-throughput, in vitro assay that allows for the simultaneous quantification of protein synthesis and proliferation within defined neural subpopulations. We demonstrate that abnormal protein synthesis in FXS is coupled to altered cellular decisions to favor proliferative over neurogenic cell fates during early development. Furthermore, pharmacologic inhibition of elevated phosphoinositide 3-kinase (PI3K) signaling corrects both excess protein synthesis and cell proliferation in a subset of patient neural cells.

Topics: Biological Assay; Cell Differentiation; Cell Lineage; Cell Proliferation; Class I Phosphatidylinositol 3-Kinases; Fragile X Mental Retardation Protein; Fragile X Syndrome; Gene Expression Regulation, Developmental; Humans; Imidazoles; Induced Pluripotent Stem Cells; Models, Biological; Morpholines; Neural Stem Cells; Neurogenesis; Organoids; Phosphoinositide-3 Kinase Inhibitors; Piperazines; Primary Cell Culture; Protein Biosynthesis; Pyrimidinones; RNA, Messenger; Signal Transduction

Aberrant neuronal translation is implicated in the etiology of numerous brain disorders. Although mTORC1-p70 ribosomal S6 kinase 1 (S6K1) signaling is critical for translational control, pharmacological manipulation in vivo has targeted exclusively mTORC1 due to the paucity of specific inhibitors to S6K1. However, small molecule inhibitors of S6K1 could potentially ameliorate pathological phenotypes of diseases, which are based on aberrant translation and protein expression. One such condition is fragile X syndrome (FXS), which is considered to be caused by exaggerated neuronal translation and is the most frequent heritable cause of autism spectrum disorder (ASD). To date, potential therapeutic interventions in FXS have focused largely on targets upstream of translational control to normalize FXS-related phenotypes. Here we test the ability of two S6K1 inhibitors, PF-4708671 and FS-115, to normalize translational homeostasis and other phenotypes exhibited by FXS model mice. We found that although the pharmacokinetic profiles of the two S6K1 inhibitors differed, they overlapped in reversing multiple disease-associated phenotypes in FXS model mice including exaggerated protein synthesis, inappropriate social behavior, behavioral inflexibility, altered dendritic spine morphology, and macroorchidism. In contrast, the two inhibitors differed in their ability to rescue stereotypic marble-burying behavior and weight gain. These findings provide an initial pharmacological characterization of the impact of S6K1 inhibitors in vivo for FXS, and have therapeutic implications for other neuropsychiatric conditions involving aberrant mTORC1-S6K1 signaling.

Topics: Animals; Autism Spectrum Disorder; Behavior, Animal; Brain; Dendritic Spines; Disease Models, Animal; Exploratory Behavior; Female; Fragile X Syndrome; Gene Expression Regulation; Imidazoles; Male; Mice, Inbred C57BL; Phenotype; Phosphorylation; Piperazines; Protein Kinase Inhibitors; Ribosomal Protein S6 Kinases, 70-kDa; Social Behavior