Transforming growth factor beta(3) inhibits chronic myelogenous leukemia hematopoiesis by inducing Fas-independent apoptosis

Objective. Transforming growth factor beta(3) (TGF-beta(3)) is a potent sup pressor of human hematopoietic progenitor cells. In this article, we compar e the activity of TGF-beta(3) on highly purified CD34(+) cells and more imm ature CD34(+)DR(-) cells from chronic myelogenous leukemia (CML) patients i n chronic phase and normal donors. Materials and Methods. Primitive hematopoietic progenitors mere stimulated in liquid cultures and clonogenic assays by early-acting growth factors suc h as stem cell factor (SCF) and interleukin 11 (IL-11) and the intermediate -late-acting stimulating factors IL-3, granulocyte-macrophage colony-stimul ating factor, and erythropoietin. Molecular analysis of bcr/abl mRNA was pe rformed on single CML colonies by nested reverse transcriptase polymerase c hain reaction. Moreover, cell cycle analysis and assessment of apoptosis of normal and leukemic CD34(+) cells were performed by propidium iodide (PI) alone and simultaneous staining with annexin V and PI, respectively. Results. The colony-forming efficiency of CML CD34(+) cells was generally i nhibited by more than 90% regardless of whether the colony-stimulating fact ors were used alone or combined. When compared to normal CD34(+) cells, leu kemic cells mere significantly more suppressed in 6 of 8 culture conditions . The inhibitory effect of TGF-beta(3) on CD34(+) cells was exerted within the first 24 hours of incubation as demonstrated by short-term preincubatio n followed by IL-3- and SCF-stimulated colony assays. Evaluation of bcr/abl transcript on residual CML colonies incubated with TGF-beta(3) demonstrate d a small subset of neoplastic CD34(+) cells unresponsive to the inhibitory effect of the study cytokine, TGF-beta(3) demonstrated a greater inhibitor y activity on primitive CD34(+)DR(-) cells than on more mature CD34(+) cell s. Again, CML CD34(+)DR(-) cells were significantly more inhibited by TGF-b eta(3) than their normal counterparts in 3 of 8 culture conditions. Kinetic analysis performed on CD34(+) cells showed that TGF-beta induces cell cycl e arrest in G(1) phase. However, this mechanism of action is shared by norm al and leukemic cells. Conversely, TGF-beta(3) preferentially triggered the programmed cell death of CML CD34(+) cells without increasing the proporti on of leukemic cells coexpressing CD95 (Fas receptor), and this effect was not reversed by functional blockade of Fas receptor. Conclusion. We demonstrate that TGF-beta(3) exerts a potent suppressive eff ect on CML cells that is partly mediated hy Fas-independent apoptosis. (C) 2000 International Society for Experimental Hematology. Published by Elsevi er Science Inc.