Overexpression of APC Reduces Stem Cell Capacity and Leukemogenic Potential of PLZF/RAR-Positive Leukemic Stem Cells.

Abstract

Acute myeloid leukemia (AML)-associated fusion proteins (AAFP), such as PML/RAR, PLZF/RAR or AML1/ETO have leukemogenic potential in vivo which is related to their capacity to induce a differentiation block and aberrant self renewal in early hematopoietic progenitors. The potential of the AAFP to induce the leukemic phenotype is strictly related to their capacity to oligomerize and to form high molecular weight complexes (HMW). Furthermore we recently showed that the deregulation of the Wnt-signaling seems to be a key event for the leukemogenesis by the above mentioned AAFP because it is indispensable for the aberrant self renewal of leukemic stem cells (LSC). A „tandem affinity purification” (TAP) screen of KG1 cells expressing PLZF/RAR, PML/RAR or AML-1/ETO for elucidating the composition of the related HMW revealed the „adenomatous polyposis coli” protein (APC) as an interaction partner of PLZF/RAR. APC is a key inhibitory regulator of the Wnt-signaling and is frequently inactivated by mutations in colon carcinomas. Therefore the role of the interaction between APC and PLZF/RAR for the leukemogenesis was further investigated. Here we report that i.) only PLZF/RAR strongly interacted with APC as confirmed by co-immunoprecipitation experiments; ii.) the overexpression of APC reverted the aberrant activation of the Wnt-signaling by PLZF/RAR as revealed by a reduction of the TCF/LEF mediated transcriptional activity; iii.) the overexpression of APC also reduced the self renewal potential of PLZF/RAR-positive HSC as revealed by a reduced replating efficiency of these cells in semi solid medium, as well as by a reduction of the colony number in colony forming units spleen (CFU-S) assays; iv.) APC was able to revert the leukemogenic potential of PLZF/RAR-positive LSC as revealed by the retroviral overexpression of APC in bone marrow isolated from mice with PLZF/RAR-induced AML which were inoculated into secondary recipients. In fact until now the presence of APC significantly increased the survival of these secondary recipients with respect to mock-infected controls. Taken together these data strongly suggest that the direct interaction with PLZF/RAR is an important mechanism for the functional inactivation of APC which contributes to the aberrant activation of the Wnt-signaling in PLZF/RAR-positive cells. Furthermore our data provide further evidence that the aberrantly activated Wnt-signaling pathway represents a valid target for molecular therapy approaches.