CRISPRi screen to identify functional long noncoding RNAs in pediatric acute myeloid leukemia

Long noncoding RNAs (lncRNAs) have emerged as new regulators of gene expression. Large-scale studies highlight their precise expression patterns, and many are implicated in pathophysiological processes, but the vast majority still lack functional characterization. We previously profiled lncRNA expression in the human blood system and in pediatric acute myeloid leukemia (AML) samples, and found stem cell and subtype-specific signatures in AML blasts. To identify functional candidates, we screened 619 of these lncRNAs in a CRISPRi-based dropout approach. Of 26 hits, 10 were validated via proliferation assays and qPCR. One candidate stood out as essential in 6 of the 8 cell lines tested – LNC666, a nuclear-enriched transcript whose knockdown is marked by megakaryocytic M-07es. It is flanked by two coding genes, both of which seem dispensable for AML cells as shown by CRISPR-Cas9 knockout. Meanwhile, shRNAs against LNC666 and gene excision both led to a growth disadvantage, with similar results in patient-derived xenografts in vivo. In healthy CD34+ cells, LNC666 knockdown reduced erythroid differentiation. Our results suggest LNC666 as an important lncRNA for the pathogenesis of AML.