Leukemia Prone B-Precursor Population in a p19ARF-/-RAG1-/- Mouse Model.

Abstract 3971

Poster Board III-907

Aberrant V(D)J-recombination may contribute to the oncogenic transformation of B-lymphoid precursor cells, suggesting that ongoing aberrant activity of recombinase activating gene (RAG) during lymphoid development is one of the potentially dangerous factors. Unexpectedly, deletions of RAG1 have been observed in a subset of cases of human B-progenitor acute lymphoblastic leukemia. Thus, the role of RAG1-mediated recombination in the pathogenesis of B-cell progenitor ALL remains unclear. The setting of RAG1-deficient severe combined immunodeficiency provides a unique opportunity to address this question. RAG1-deficient patients harbour an atypical CD34+CD19+ B-lymphoid precursor population in the bone marrow that is not present in healthy individuals. The role of these cells in the development of leukaemia has so far not been reported.

To analyze the role of RAG1 deficiency in the pathogenesis of leukaemia, we analyzed mice that lacked expression of p19ARF (and were thus tumour prone) and RAG1. p19ARF-/- (A) mice develop predominantly solid tumours, and in a minority of cases, T-lineage leukemia/lymphoma. p19ARF-/-RAG1-/- (AR) mice spontaneously developed B-precursor leukaemia (CD19+IgM-) at an incidence of 30% within 52 weeks of follow up, indicating that the simultaneous loss of both p19ARF and RAG1 promotes the development of B-lineage leukemia. Array CGH analysis of the resulting leukemia using customized microarrays incorporating genome wide coverage and dense tiling of genes involved in B-cell development identified recurring aneuploidy of chromosomes 1, 5 and 14, but a notable absence of RAG1-mediated DNA copy number alterations that are observed in RAG-sufficient mouse models for ALL. In support of this, we identified a leukemia-prone Sca1+CD19+ precursor cell population in the bone marrow of AR mice, which was not present in A mice. Transplantation of AR haematopoietic precursor cells (Sca1+) into irradiated wild type recipients accelerated leukaemia incidence to 100% in contrast to transplantation of AR Sca1+CD19- population, which resulted in a decreased incidence of B-precursor cell leukaemia. The leukaemia prone AR Sca1+CD19+ population showed a high BrdU uptake with an abnormal cell cycle progression and seems to be committed to the B cell lineage, as colony forming assays demonstrated. Co-culture of the AR Sca1+CD19+ population with OP9delta1 cells failed to differentiate these cells into T-precursor cells, however ongoing proliferation of B-lineage cells was observed.

Further characterisation of the RAG1-/- lymphoid precursor compartment revealed an equivalent Sca1+CD19+ population that was not leukaemia prone, and expressed p19ARF. This suggests that p19ARF is an important factor implied in tumour suppression in this B-precursor compartment.

Here we show for the first time in a mouse model that RAG1 loss of function in combination with inactivation of p19ARF contributes to leucemogenesis via accumulation of a leukaemia prone B-precursor cell population in the bone marrow.