HOXA10-AS is required for the maintenance of KMT2A-rearranged AML cells. (A) Percentage of cells transduced with shRNAs against HOXA10-AS after 21 days in culture compared with day 5 and a nontargeting shRNA (ctrl; n = 3; unpaired Student t test; presented as mean ± SEM). (B) Cell counts on day 21 following treatment with LNA-GapmeRs against HOXA10-AS compared with day 2 and the nontargeting control (ctrl; n = 2; unpaired Student t test; presented as mean ± SEM). (A-B) EOL-1, MOLM-13, and MV4-11 were used as KMT2A-r cell lines, while NB-4 was used as the non-KMT2A-r cell line. (C-D) 5-Bromo-2′-deoxyuridine cell cycle (C) and Annexin V apoptosis (D) assays in EOL-1 and NB-4 cells after shRNA-mediated HOXA10-AS knockdown using 3 different shRNAs. The percentage of cells is shown as mean ± SD (n = 2; *P < .05; **P < .01). (E) Schematic of the genomic locus and of the regions that are excised by the sgRNA pairs. (F) Percentage of cells transduced with HOXA10-AS excision sgRNAs (GFP⁺) after 21 days in culture compared with day 2 and nontargeting sgRNAs (n ≥ 3; unpaired Student t test; presented as grand mean). EOL-1, MOLM-13 and MV4-11 were used as KMT2A-r cell lines, while NB-4 was used as the non-KMT2A-r cell line. (G-H) 5-Bromo-2′-deoxyuridine cell cycle (G) and Annexin V apoptosis (H) assays in EOL-1 and NB-4 cells after sgRNA-mediated excision using 4 different sgRNA pairs. The percentage of cells is shown as mean ± SD (n = 2; *P < .05; **P < .01).