Figure 3
Figure 3. Eltrombopag does not increase long-term self-renewal of normal or MDS/AML patient-derived BM-MNC. Serial replating assays of BM-MNC were performed in methylcellulose containing hrIL-3, hrSCF, hrIL-6, insulin, and transferrin. Cultures were supplemented with 0, 3, or 10 μg/mL Eltrombopag or 100 ng/mL hrTPO. After 12 days of culture, cells were isolated from the methylcellulose medium and replated onto the next plate. Serial replatings were carried out for 4 rounds of replating or until colony formation exhausted. (A) Serial relating of BM-MNC from healthy donors (n = 5). (B) Serial replating of BM-MNC from patients with MDS/AML (n = 13). (C) Xenotransplantation assays of BM-MNC from MDS/AML patients (n = 3). BM-MNC were transplanted into NOG mice after sublethal irradiation. Successfully engrafted mice were either treated with Eltrombopag (0.3 mg/mL) in the drinking water or left untreated. After 12 weeks of Eltrombopag treatment, all mice were killed and analyzed for human CD45-expressing cells by FACS. (Left) Representative dot plot of human CD45 analysis. (Right) Chimerism of human CD45+ cells in xenografted mice (n = 20 total) with or without Eltrombopag treatment. Averages and SDs of mouse cohorts of each of the 3 transplanted samples are shown. None of the cohorts showed higher engraftment upon treatment with Eltrombopag.

Eltrombopag does not increase long-term self-renewal of normal or MDS/AML patient-derived BM-MNC. Serial replating assays of BM-MNC were performed in methylcellulose containing hrIL-3, hrSCF, hrIL-6, insulin, and transferrin. Cultures were supplemented with 0, 3, or 10 μg/mL Eltrombopag or 100 ng/mL hrTPO. After 12 days of culture, cells were isolated from the methylcellulose medium and replated onto the next plate. Serial replatings were carried out for 4 rounds of replating or until colony formation exhausted. (A) Serial relating of BM-MNC from healthy donors (n = 5). (B) Serial replating of BM-MNC from patients with MDS/AML (n = 13). (C) Xenotransplantation assays of BM-MNC from MDS/AML patients (n = 3). BM-MNC were transplanted into NOG mice after sublethal irradiation. Successfully engrafted mice were either treated with Eltrombopag (0.3 mg/mL) in the drinking water or left untreated. After 12 weeks of Eltrombopag treatment, all mice were killed and analyzed for human CD45-expressing cells by FACS. (Left) Representative dot plot of human CD45 analysis. (Right) Chimerism of human CD45+ cells in xenografted mice (n = 20 total) with or without Eltrombopag treatment. Averages and SDs of mouse cohorts of each of the 3 transplanted samples are shown. None of the cohorts showed higher engraftment upon treatment with Eltrombopag.

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