Massively parallel sequencing has revealed many de novo mutations in the etiology of developmental and epileptic encephalopathies (EEs), highlighting their genetic heterogeneity. Additional candidate genes have been prioritized in silico by their co-expression in the brain. Here, we evaluate rare coding variability in 20 candidates nominated with the use of a reference gene set of 51 established EE-associated genes. Variants within the 20 candidate genes were extracted from exome-sequencing data of 42 subjects with EE and no previous genetic diagnosis. We identified 7 rare non-synonymous variants in 7 of 20 genes and performed Sanger sequence validation in affected probands and parental samples. De novo variants were found only in SLC1A2 (aka EAAT2 or GLT1) (c.244G>A [p.Gly82Arg]) and YWHAG (aka 14-3-3g) (c.394C>T [p.Arg132Cys]), highlighting the potential cause of EE in 5% (2/42) of subjects. Seven additional subjects with de novo variants in SLC1A2 (n ¼ 1) and YWHAG (n ¼ 6) were subsequently identified through online tools. We identified a highly significant enrichment of de novo variants in YWHAG, establishing their role in early-onset epilepsy, and we provide additional support for the prior assignment of SLC1A2. Hence, in silico modeling of brain co-expression is an efficient method for nominating EE-associated genes to further elucidate the disorder's etiology and genotype-phenotype correlations.
Glioblastomas (GBM) are associated with high rates of relapse. These brain tumors are often resistant to chemotherapies like temozolomide (TMZ) and there are very few treatment options available to patients. We recently reported that polo-like kinase-1 (PLK1) is associated with the proliferative subtype of GBM; which has the worst prognosis. In this study, we addressed the potential of repurposing disulfiram (DSF), a drug widely used to control alcoholism for the past six decades. DSF has good safety profiles and penetrates the blood-brain barrier. Here we report that DSF inhibited the growth of TMZ resistant GBM cells, (IC90=100 nM), but did not affect normal human astrocytes. At similar DSF concentrations, self-renewal was blocked by ~100% using neurosphere growth assays. Likewise the drug completely inhibited the self-renewal of the BT74 and GBM4 primary cell lines. Additionally, DSF suppressed growth and self-renewal of primary cells from two GBM tumors. These cells were resistant to TMZ, had unmethylated MGMT, and expressed high levels of PLK1. Consistent with its role in suppressing GBM growth, DSF inhibited the expression of PLK1 in GBM cells. Likewise, PLK1 inhibition with siRNA, or small molecules (BI-2536 or BI-6727) blocked growth of TMZ resistant cells. Our studies suggest that DSF has the potential to be repurposed for treatment of refractory GBM.
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