Fig. 4. mTOG-Ψ treatment improves differentiation and engraftment of malignant MDS-HSPCs.
a, Experimental approach employed to examine the effects of mTOG-Ψ and PAIP1 on the CFU potential of PUS7-KD MDS-L cells and patient-derived HR-MDS-HSPCs. b, Number of colonies obtained from HSPCs from two patients with HR-MDS (MDS135 and MDS304) 15 d following treatment with SCR-Ψ, mTOG-Ψ, siPAIP1, and mTOG-Ψ and siPAIP1. MDS135, *P = 0.0129; and MDS304, **P = 0.0016; one-way ANOVA with multiple comparison; n = 2–5 independent biological experiments, indicated as individual dots, subject to material availability. c, Number of colonies in shRNA control (shCTRL)-treated and PUS7-KD MDS-L cells on day 15 following transduction with SCR-Ψ, mTOG-Ψ, siPAIP1, and mTOG-Ψ and siPAIP1. Data are the mean ± s.d. of n = 3 independent biological replicates. **P = 0.0093; ***P = 0.0009; *P = 0.0348 (shPUS7 + siPAIP1) and *P = 0.0197 (shPUS7 + mTOG-Ψ + siPAIP1); two-tailed Student’s t-test. d, Schematic showing the experimental conditions used for HSPC differentiation in the presence of erythropoietin (EPO) or granulocyte colony-stimulating factor (G-CSF). e, Number of differentiated erythroid and myeloid cells, relative to the SCR-Ψ control, obtained following different treatments. The experiments were performed in duplicate or triplicate; n = 4 patients, except for mTOG-Ψ, where n = 5 patients; each dot represents a patient. Erythoid, *P = 0.047 (mTOG-Ψ), ***P = 0.0003 (siPAIP1), **P = 0.0016 (siPAIP1 + mTOG-Ψ); myeloid, ****P = 0.000012 (mTOG-Ψ), **P = 0.0011 (siPAIP1), **P = 0.0036 (siPAIP1 + mTOG-Ψ); one-tailed Student’s t-test. f,g, Schematic of the HR-MDS-HSPC xenotransplantation experiment (f) and representative fluorescence-associated-cell-sorting plots showing the expression levels of human CD45 (hCD45) in the BM of the NSG-S mice (h). g, The percentages of hCD45 cells in the red gates are shown. h, Percentage of human engraftment (hCD45+) in mice transplanted with HSPCs from three patients with HR-MDS (MDS275, MDS135 and MDS272). *P = 0.0299 and *P = 0.0283; two-tailed Student’s t-test; n = 2–5 mice, each dot represents a transplanted mouse. i, Changes in the percentage of myeloid (CD33+) and lymphoid (CD19+) cells in each mTOG-Ψ-treated patient-derived xenotransplant; one-tailed Student’s t-test. j, Flow cytometric analysis of human CD123 in hCD45+hCD34+hCD45RA+ cells from littermates transplanted with HR-MDS-HSPCs ± SCR-Ψ or mTOG-Ψ (left). The levels of CD123 in the same cell population from an NSG-S mouse transplanted with LR-MDS-HSPCs are shown for comparison (dashed black line). Mean fluorescence intensity (MFI) of CD123 in hCD45+hCD34+hCD45RA+ cells from the patient-derived xenotransplantation experiments (right). **P = 0.0047; two-tailed Student’s t-test. b,c,h,i, Data are the mean ± s.d.