LAUSR

NUP98-HOXA9 (“NA9”) is a fusion gene commonly found, together with other mutations, in human acute myeloid leukemia cells, and occasionally in blast-phase (BP) cells of patients with chronic myeloid leukemia (CML) in whom disease progression has occurred. These findings underscore interest in elucidating the role of NA9 in contributing to an aggressive leukemic program. Previous studies addressing this issue reported biological and molecular consequences of NA9 alone in human cells, including normal human hematopoietic cells, and evidence of a co-operative role of NA9 with BCR-ABL1 in experimentally-induced models of leukemia in mouse cells.We now report the results of assessing the biological and molecular consequences of lentivirally-introduced expression of NA9 in freshly isolated CD34 CML cells from patients with chronic phase (CP) disease (for a general outline and technical details, see Online Supplementary Figure S1A-E). Samples from patients with CP CML are known to contain variable ratios of normal and BCR-ABL1/Philadelphia chromosome-positive cells, particularly in the most primitive cell compartments. Therefore, for this study, we selected samples from three CP-CML patients for whom we had previously shown the content of any type of normal CD34 cells to be very low based on assessments of their progeny genotypes produced for >30 weeks (Online Supplementary Table S1). Initial in vitro experiments with these three CML patients’ CP cells showed that NA9-transduced cells (compared to control-transduced cells) produced particularly large expansions of granulopoietic colony-forming cells in growth factor-supplemented liquid suspension cultures, with somewhat reduced effects on expanding the outputs of these progenitors in 6-week long-term cultures on human growth factor-producing stroma, or in serial replating assays of the transduced cells cultured in semi-solid medium (Online Supplementary Figure S1F-H). However, in all cases, the progeny of the NA9-transduced CML cells ultimately underwent terminal differentiation (Online Supplementary Figure S1I), thus recapitulating results reported previously for NA9-transduced normal human CD34 hematopoietic cells. Outputs of NA9and control-transduced CD34 cells from the same three CP-CML patients’ samples were also assessed in sublethally irradiated NOD-PrkdcIL2Rgc mice engineered to produce human interleukin 3, granu-