Figure 2. Normal and leukemic human HSPC engraftment in vivo following MYXV treatment.

(A) Normal BM-derived MNCs or CD34+ selected cells were exposed to MYXV at 10 MOI for 3-hours ex vivo and transplanted into irradiated NOG mice. Analysis of mouse BM after 6 to 8-weeks demonstrated normal human cell engraftment with CD45⁺/HLA-abc⁺ cells observed in both MYXV treated (n=9) and mock treated (n=10) groups. Shown are representative flow cytometry plots. (B) Similar engraftment frequencies were observed between mock and MYXV treated cells. As expected, the percent engraftment values varied between mice; however, the overall levels of engraftment were similar between mice receiving mock or MYXV treated cells (p=0.41). Symbols representing mice transplanted with MNCs or CD34+ cells are shown in open and filled shapes (circle: Mock treated, square: MYXV purged), respectively. (C) AML cells were mock treated or exposed to MYXV at 10 MOI for 3-hours ex vivo and subsequently transplanted into irradiated NOG mice. Mock treated AML cells demonstrated leukemic engraftment in all mice tested (n=7). MYXV treated AML cells resulted in leukemic engraftment in only 10% of transplanted mice as analyzed by flow cytometry (n=10). Shown are representative flow cytometry plots. (D) Leukemic engraftment was also demonstrated using PCR for the FLT3 ITD mutation. Leukemic human cells in mouse bone marrow all exhibited the AML FLT3 ITD genotype. The FLT3 ITD mutation in AML cells created the larger FLT3 band, compared to normal controls. Only engrafted samples are shown: one representative of 7 from mock-treated and the single FACS-positive engrafted sample from the 10 MYXV-purged mice. Normal BM cells and AML MNCs were used as PCR controls. (E) MYXV treatment resulted in significantly lower percent engraftment levels in comparison to mock treated controls (p<0.05). Values represent mean±SEM. Gating was established using appropriate isotype controls.