Non-coding RNAs, including microRNAs (miRNAs), support the progression of glioma. miR-21 is a small, non-coding transcript involved in regulating gene expression in multiple cellular pathways, including the regulation of proliferation. High expression of miR-21 has been shown to be a major driver of glioma growth. Manipulating the expression of miRNAs is a novel strategy in the development of therapeutics in cancer. In this study we aimed to target miR-21. Using CRISPR genome-editing technology, we disrupted the miR-21 coding sequences in glioma cells. Depletion of this miRNA resulted in the upregulation of many downstream miR-21 target mRNAs involved in proliferation. Phenotypically, CRISPR-edited glioma cells showed reduced migration, invasion, and proliferation in vitro. In immunocompetent mouse models, miR-21 knockout tumors showed reduced growth resulting in an increased overall survival. In summary, we show that by knocking out a key miRNA in glioma, these cells have decreased proliferation capacity both in vitro and in vivo. Overall, we identified miR-21 as a potential target for CRISPR-based therapeutics in glioma.
Gliomas are primary, diffusely infiltrating brain tumors. Microglia are innate immune cells in the CNS and make up a substantial portion of the tumor mass. Glioma cells shape their microenvironment, communicating with and reprogramming surrounding cells, resulting in enhanced angiogenesis, immune suppression, and remodeling of the extracellular matrix. Glioma cells communicate with microglia, in part by releasing extracellular vesicles (EVs). Mouse glioma cells stably expressing a palmitoylated GFP to label EVs were implanted intracranially into syngeneic miR-21-null mice. Here, we demonstrate functional delivery of miR-21, regulating specific downstream mRNA targets in microglia after uptake of tumor-derived EVs. These findings attest to EV-dependent microRNA delivery as studied in an in vivo-based model and provide insight into the reprograming of microglial cells by tumor cells to create a favorable microenvironment for cancer progression.