Abstracts

Rationale: Fms-like tyrosine kinase 3 (FLT3) is a receptor tyrosine kinase expressed preferentially in blood and brain cells and is activated by the FLT3 ligand, a pleiotrophic cytokine abundant in blood and cortical spinal fluid. The FLT3 signaling pathway has been extensively studied in the context of blood cell development and leukemia, but its function in the brain remains largely unknown. Our group’s previous high-throughput drug screening work led to the unexpected discovery that a number of small molecule FLT3 inhibitor compounds (FLT3i), including KW-2449 and Sunitinib, stimulate the gene expression of chloride transporter KCC2 in neurons. KCC2 plays a pivotal role in brain development and functioning while dysregulation of KCC2 is strongly associated with epilepsy.

Methods: In this study, we undertook a functional genomics approach to examine the transcriptomic changes induced by pharmacological inhibition of the FLT3 pathway in brain cells, and tested the brain penetrance, target engagement, and anti-seizure efficacy of FLT3i drugs in a mouse model of epilepsy.


Results: Our results show that treating cultured human or mouse neurons with FLT3i drugs significantly stimulates the expression of genes that play important roles in brain development, while suppressing genes linked to neuroinflammation. On the contrary, FLT3i treatment of human microglia cells does not induce any significant transcriptomic change, highlighting the cell type specificity of FLT3 signaling in the brain. We performed a curated candidate transcription factor CRISPR screening to elucidate the molecular mechanism underlying FLT3-mediated gene regulation in neurons. The mRNA and protein expression levels of a number of epilepsy risk genes are enhanced in FLT3i-treated neurons, supporting potential therapeutic application of FLT3i to rescue disease risk gene haploinsufficiency. We further tested the in vivo efficacy of FLT3i drug in mice to demonstrate that KW-2449 can enter the mouse brain, induce KCC2 protein expression for up to three days via a single injection, and suppress seizure in a chemoconvulsant-induced mouse model of temporal lobe epilepsy.


Conclusions: Taken together, our results reveal previously unknown roles of FLT3 signaling in promoting neuronal functional maturation and suppressing neuroinflammation, indicating that FLT3 kinase signaling modulates a transcriptional program that plays critical roles in brain development and health, providing a promising novel therapeutic target for the treatment of epilepsy.


Funding: Simons Foundation Autism Research Initiative, Charles H. Hood Foundation, SYNGAP Research Fund, PTEN Foundation.