Regulation of apoptosis in AMI, MDS and normal hematopoiesis

Members of the Bcl-2 family of genes are believed to regulate apoptosis in hematopoietic cells. We examined, by RT-PCR, Western blotting and quantitative flow cytometry (Q-FCM) the expression of Bcl-2 related genes in normal hematopoiesis and AML. In early normal progenitors (CD34 + 33-13-1, Bcl-XL but not Bcl-2 was found to be expressed. Normal promyelocytes (CD34- 33 + ) had no detectable Bcl-2 or Bcl-X while leukemic promyelocytes expressed Bcl-2 and Bcl-XL. Bcl-XL was highly, but variably expressed in the majority of AML. Bcl-2 was expressed in all AML samples and AML CD34 cells studied, but the level of Bcl-2 did not predict for response to FLAG-lda (Fludarabine-ARA-C-ldarubicin-G-CSF) induction chemotherapy (p = 0.79). However, in patients failing to achieve CR, small subpopulations of preexisting cells expressing very high Bcl-2 levels were Identified by Q-FCM which had a survival advantage compared to cells with lower Bcl-2. Cytokines did not affect Bcl-2 or Bax levels in AML blasts in vitro, but exerted their anti-apoptotic effect by decreasing expression of pro-apoptotic Bax. Downregulation of Bcl-2 and Bci-X, by ATRA in H L-60 and NB-4 cells was associated with increased cell kill by ARA-C. The effects of ATRA on genes regulating apoptosis in AML IQ vivo are presently under investigation (FLAIRG protocol). Other Bcl-2 family genes are also being studied. While d_g noyo AML undergo little spontaneous apoptosis, a high degree of apoptosis was found in MDS. We are presently investigating the expression of apoptotic genes in AML vs. MDS progenitor cells and expect that a better understanding of the mechanisms regulating apoptosis will help in the development of new therapeutic strategies.