3169 - Serial Transplantation of Pre-Leukemic T-ALL Stem Cells Requires EphA3

Abstract

T cell acute lymphoblastic leukaemia (T-ALL) represents 10–15% of all ALL, the most common pediatric leukemia. The abnormally self-renewing thymocytes of NUP98-HOXD13 (NHD13) transgenic mice, defined by their ability to engraft upon transplantation, are a novel model of T-ALL pre-leukemic stem cells, which self-renew but do not immediately cause leukaemia. We observed that NHD13 thymocytes inappropriately express the cell-surface tyrosine kinase receptor EphA3, which has been implicated in self-renewal in other cancers. To investigate the role of EphA3 in the abnormal self-renewal in NHD13 thymocytes, we generated a NHD13/EphA3-knockout mouse line. We compared the thymocytes of NHD13/EphA3-knockout mice with those from NHD13 mice and found similar abnormal clonal expansion based on the rearrangement of the TCR-B genes, and no differences in the expression of known abnormally-expressed self-renewal genes including HoxA5, HoxA7, HoxA9, HoxA10, Lmo2, Hhex and Lyl1. We performed thymocyte transplantations and found that NHD13/EphA3-knockout thymocytes are able to engraft similarly to NHD13 thymocytes, demonstrating similar abnormal self-renewal activity. Strikingly, while the NHD13 thymocytes engraft indefinitely upon serial transplantation, the NHD13/EphA3-knockout thymocytes fail to engraft by the third passage. This suggests that EphA3 is important for long- but not short-term self-renewal (as defined by the serial transplantation assay), and that this activity depends on molecular pathways other than those already identified in this and similar models. The data is also consistent with the potential involvement of EphA3 in a thymic import blockade effect, in which deletion of EphA3 may re-allow thymic progenitors to enter the thymus and out-compete the self-renewing NHD13 cells over time. Cell fate experiments are underway to examine this possibility. This study presents a new proposed mechanism for EphA3 in leukemic development, and the first oncogene-driven mechanism by which thymic progenitor deprivation may promote T-ALL.