Enhanced IGFBP7 expression or treatment with rhIGFBP7 induces sensitivity to ATRA in primary AML stem and progenitor cells. For all ex vivo experiments, cells were incubated with PBS (Control/Ctrl), 100 µg/mL rhIGFBP7, 0.5 µM ATRA, or the combination (Combi). For CFU progenitor and long-term liquid culture (stem cell) assays, samples (in duplicate) were incubated for 7 days or 4 weeks, respectively, and normalized against untreated controls. Data are plotted as mean ± SEM, unless stated otherwise. Patient sample characteristics are summarized in supplemental Table 1. (A-C) Primary AML samples were lentivirally transduced with control or IGFBP7-OE vectors. (A) CFU progenitor assays of primary AML samples, plotted as mean ± SD. (B) CFU plates, containing all colonies, were harvested, and flow cytometric analysis of AML cells derived from the colonies was performed, showing leukemic CD45^dim blasts in the transduced cell population for AML9 (LAIP = CD15; left panels) and AML29 (LAIP = CD56; right panels). (C) Percentage of CD45^dimLAIP⁺ blasts (left panel) and CD45^highLAIP⁺ blasts (middle panel) derived from the colonies, quantified, and normalized against untreated controls. Percentage of lymphocytes (right panel), measured using flow cytometry and plotted as mean ± SD. (D-E) CFU progenitor assays of primary AML samples, with AML cases not responding to combination therapies shown in the right lighter bars. Samples were incubated with rhIGFBP7 and ATRA (D) or with TCP (10 µM) and ATRA (E). (F) CFU stem cell assay (long-term liquid culture) of primary AML samples incubated with rhIGFBP7 and ATRA. (G) Schematic overview of the experiment (left panel). After injection of T-cell–depleted primary AML cells, NSG mice were treated with rhIGFBP7 (10 mg/kg) in week 4 (day 1) and week 8 (day 1-3) and/or ATRA (10 mg, 21-day-release pellet) in week 8 (day 3). Subsequently, equal numbers of human myeloid hCD45⁺CD33⁺ cells derived from the first transplant were injected into secondary recipients. At week 16, bone marrow cells from the mice were analyzed for the presence of hCD45⁺ cells (middle panel) and myeloid hCD45⁺CD33⁺ cells (right panel). *P < .05, **P < .01, ***P < .001, 1- or 2-way ANOVA with post hoc Tukey’s multiple-comparison test, unless stated otherwise.