Activating mutations in the NOTCH1 gene have been found in about 60% of patients with T-cell acute lymphoblastic leukemia (T-ALL). In order to study the molecular mechanisms by which altered Notch signaling induces leukemia, a zebrafish model of human NOTCH1-induced T-cell leukemia was generated. Seven of sixteen mosaic fish developed a T-cell lymphoproliferative disease at about 5 months. These neoplastic cells extensively invaded tissues throughout the fish and caused an aggressive and lethal leukemia when transplanted into irradiated recipient fish. However, stable transgenic fish exhibited a longer latency for leukemia onset. When the stable transgenic line was crossed with another line overexpressing the zebrafish bcl2 gene, the leukemia onset was dramatically accelerated, indicating synergy between the Notch pathway and the bcl2-mediated antiapoptotic pathway. Reverse transcription-polymerase chain reaction analysis showed that Notch target genes such as her6 and her9 were highly expressed in NOTCH1-induced leukemias. The ability of this model to detect a strong interaction between NOTCH1 and bcl2 suggests that genetic modifier screens have a high likelihood of revealing other genes that can cooperate with NOTCH1 to induce T-ALL.

Keywords: Notch; bcl2; T-ALL; zebrafish

Introduction

T-cell acute lymphoblastic leukemia (T-ALL) comprises 10-25% of all pediatric and adult ALL. Unlike the more common B-cell lineage ALL, T-ALL is characterized by the expression of stage-related T-cell surface antigens, higher leukocyte counts, older age at presentation and a male predominance. Even though cure rates for adult and children with T-ALL have reached 50 and 65-75%, respectively, (1-5) the prospects for additional improvements in therapy are dependent upon a more complete understanding of the molecular aspects of T-ALL pathogenesis and how they influence prognosis.

Notch receptors are a family of cell surface proteins that regulate cell fate decisions and growth control. (6,7) Dysregulated Notch signal transduction can contribute to several types of cancers, in addition to other human diseases. Notch signaling was first linked to tumorigenesis through identification of a recurrent t(7;9)(q34;q34.3) chromosomal translocation that is found in an uncommon subset of human T-ALL. (8) This chromosome translocation juxtaposes the NOTCH1 intracellular domain (ICN1) with the T-cell receptor-β chain promoter/enhancer region, resulting in the constitutive activation of Notch signaling in the T cells. Mice reconstituted with hematopoietic progenitor cells expressing the truncated human NOTCH1 gene develop T-cell leukemia. (9,10) Immature, double-positive T cells accumulate in the bone marrow with simultaneous inhibition of B-cell development, indicating that Notch1 signaling drives common lymphoid progenitor cells into the T-cell lineage. T-cell neoplasia can also be caused by proviral integration into the Notch gene, which leads to aberrant Notch signaling. Proviral integration of the Moloney murine leukemia virus or feline leukemia virus into Notch1 or Notch2, respectively, causes T-cell leukemia. (11,12) Although the t(7;9)(q34;q34.3) chromosomal translocation is uncommon, it was recently found that about 60% of adult and childhood T-ALL patients carry deletions and/or mutations in the NOTCH1 gene. (13-16) In vitro Notch reporter assays indicated that these mutations and/or deletions lead to aberrant NOTCH1 activation. Both γ-secretase inhibitors and dominant-negative Mastermind (MAML1) induce G1 cell-cycle arrest of...