Wnt Signaling Is Associated with Anti-Apoptosis in the Interaction Between Acute Myeloid Leukemia Cells and Stromal Cells

Abstract 1366

Recent studies have revealed that strength of the Wnt signaling pathway regulates normal hematopoiesis including hematopoietic stem cells, and aberrant activation of the pathway is involved in the development of several types of leukemias. In the bone marrow microenvironment, stromal cells are rich sources of cytokines and some secreted cytokines have been observed to block induction of cell death in myeloid leukemia cells exposed to chemotherapy. Here, we examined the role of the Wnt signaling pathway on cell-adhesion, proliferation and survival of the stroma-dependent human AML cell line, TRL-01 cells, which we previously established (Ninomiya, et al. Cancer Gen Cyto, 2006). TRL-01 cells were maintained in the co-culturing with the hTERT-transduced human bone marrow stroma cell line, HTS cells, and cell death of TRL-01 cells was induced after removal of HTS cells. Treatment with the Wnt-receptor competitor, secreted Frizzled related protein (sFRP)-1, or the Rho kinase inhibitor, Y29632 (previously reported as an inhibitor of the non-canonical Wnt downstream pathway), but not with the specific inhibitor of the canonical Wnt pathway (DKK-1) induced apoptosis in dose-dependent manners in TRL-01 cells co-cultured with HTS cells. These results suggested that the non-canonical pathway of Wnt signaling might regulate survival of TRL-01 on the stromal cells. Next, we comprehensively investigated transcripts of the Wnt pathway components (10 Frizzleds, 2 LRPs, and 18 Wnts) in TRL-01 cells and HTS cells using RT-PCR. Transcripts of Wnt5A and Wnt9A were expressed in TRL-01 cells, but not in HTS cells. Moreover, the cell death of TRL-01 cells after removal of HTS cells was partially prevented by additional treatment with Wnt5A or Wnt9A not by other Wnt molecules such as Wnt5B. On the other hand, treatment with Wnt3A induced activated nuclear beta-catenin using Western-blotting, however, did not contribute to the survival of TRL-01 cells without the stromal co-culturing. Moreover, we are investigating regulation of the anti-apoptotic downstream pathway molecules in TRL-01 cells as well as effects of other inhibitors targeted for the Wnt signaling. We are also examining expressions of Wnt pathway components in primary AML bone marrow cells and other human bone marrow stroma cell lines such as HS-5 cells. These results imply association of Wnt5A and Wnt9A with anti-apoptosis in the interaction between AML cells and stromal cells, and a possible therapeutic target of AML for overcoming the resistance to chemotherapy in the bone marrow microenvironment.