Small molecule screen reveals pathways that regulate C4 secretion in stem-cell derived astrocytes.

In the brain, the complement system plays a crucial role in the immune response and in synaptic elimination during normal development and disease. Here, we sought to identify pathways that modulate the production of complement component 4 (C4), recently associated with an increased risk of schizophrenia. To design a disease-relevant assay, we first developed a rapid and robust 3D protocol capable of producing large numbers of astrocytes from pluripotent cells. Transcriptional profiling of these astrocytes confirmed the homogeneity of this population of dorsal fetal-like astrocytes. Using a novel ELISA-based small molecule screen, we identified epigenetic regulators, as well as inhibitors of intracellular signaling pathways, able to modulate C4 secretion from astrocytes. We then built a Connectivity Map to predict and validate additional key regulatory pathways, including one involving c-Jun-kinase (JNK). This work provides a foundation for developing therapies for CNS diseases involving the complement cascade. Biological replicates of astrocytes differentited from human stem-cells (IPSCs, 1016A, n=3 ,ESCs, Hues8 n=3 and H1 n=2) were analyzed using scRNAseq