Inhibition of STAT3 leads to selective eradication of LSCs compared with HSPCs. (A) Viability of LSCs isolated from 6 primary human AML samples, 3 at the time of diagnosis and 3 at the time of relapse, after 24-hour treatment with 5 µM of STAT3i compared with vehicle control. (B) Percent number of colonies from 3 primary AML samples treated with 5µM of STAT3i or vehicle control for 24 hours and plated in human methylcellulose media for up to 14 days. (C) Western blot showing protein level of STAT3 after treatment with shRNA against STAT3 or scramble control (shSCR). (D) Percent number of colonies in 3 primary AML samples treated with either shSTAT3 or shSCR. (E) Viability of CD34⁺ cells (HSPCs) isolated from 3 human cord blood samples and treated with 5 µM of STAT3i vs vehicle control after 24 hours. NS, not significant. (F) Percent colonies of HSPCs isolated from 3 cord blood samples and treated with either 5 µM of STAT3i or vehicle control for 24 hours and plated in human methylcellulose media for up to 14 days. (G) Percent engraftment at 8 to 10 weeks of 3 primary AML samples pretreated with either 5 μM of STAT3i or vehicle control overnight and injected into busulfan-treated NSG-S mice. The sample in bracket was used for secondary transplant, which shows complete eradication of LSCs. (H) Percent human blasts in mice engrafted with a primary AML samples and treated in vivo with 30 mg/kg of STAT3i or vehicle control intraperitoneally daily for 6 days. Statistical analyses were performed using the Student t-test. P values are represented as follows: *P ≤ .05; **P ≤ .01; ***P ≤ .001; ****P ≤ .0001.