OBJECTIVES/GOALS: Glioblastomas (GBMs) are heterogeneous, treatment-resistant tumors that are driven by populations of cancer stem cells (CSCs). In this study, we perform an epigenetic-focused functional genomics screen in GBM organoids and identify WDR5 as an essential epigenetic regulator in the SOX2-enriched, therapy resistant cancer stem cell niche. METHODS/STUDY POPULATION: Despite their importance for tumor growth, few molecular mechanisms critical for CSC population maintenance have been exploited for therapeutic development. We developed a spatially resolved loss-of-function screen in GBM patient-derived organoids to identify essential epigenetic regulators in the SOX2-enriched, therapy resistant niche. Our niche-specific screens identified WDR5, an H3K4 histone methyltransferase responsible for activating specific gene expression, as indispensable for GBM CSC growth and survival. RESULTS/ANTICIPATED RESULTS: In GBM CSC models, WDR5 inhibitors blocked WRAD complex assembly and reduced H3K4 trimethylation and expression of genes involved in CSC-relevant oncogenic pathways. H3K4me3 peaks lost with WDR5 inhibitor treatment occurred disproportionally on POU transcription factor motifs, required for stem cell maintenance and including the POU5F1(OCT4)::SOX2 motif. We incorporated a SOX2/OCT4 motif driven GFP reporter system into our CSC cell models and found that WDR5 inhibitor treatment resulted in dose-dependent silencing of stem cell reporter activity. Further, WDR5 inhibitor treatment altered the stem cell state, disrupting CSC in vitro growth and self-renewal as well as in vivo tumor growth. DISCUSSION/SIGNIFICANCE: Our results unveiled the role of WDR5 in maintaining the CSC state in GBM and provide a rationale for therapeutic development of WDR5 inhibitors for GBM and other advanced cancers. This conceptual and experimental framework can be applied to many cancers, and can unmask unique microenvironmental biology and rationally designed combination therapies.

Metsulfuron is used for POST control of spotted spurge in many warm-season turfgrasses. A suspected resistant (R) biotype of spotted spurge was collected from turfgrass in Georgia with a history of exclusive metsulfuron use. Research was conducted to evaluate the resistance level of this biotype to metsulfuron, efficacy of other mechanisms of action for control, and the molecular basis for resistance. Compared with a susceptible (S) biotype, the R biotype required >90 and >135 times greater metsulfuron rates to reach 50% injury and reduce biomass 50% from the nontreated, respectively. The R biotype was also resistant to trifloxysulfuron but was injured equivalent to the S biotype from dicamba, glyphosate, and triclopyr. Gene sequencing of the R biotype revealed a Trp574 to Leu substitution that has conferred resistance to acetolactate synthase (ALS) inhibitors in previous research. This is the first report of ALS resistance in spotted spurge. More importantly, this is the first report of a herbicide-resistant broadleaf weed from a turfgrass system in the United States.