Overexpression of BHLHB1 (OLIG 2) Causes Precursor T-Cell Lymphoblastic Lymphoma/Leukemia (pre-T LBL) in the Presence of LMO1 Overexpression.

The human BHLHB1 (basic domain, helix-loop-helix protein, class B, 1; also known as OLIG 2) was initially cloned by virtue of its activation in pre-T lymphoblastic lymphoma/leukemia (pre-T LBL) with a t(14; 21)(q11.2; q22). BHLHB1, whose expression is normally restricted to neural tissues, is activated in this translocation by its juxtaposition with the TCRA locus. A survey of patients with pre-T LBL (Ferrando et al., Lancet 363:535) indicated that rare patients without a known t(14;21) overexpressed BHLHB1. We generated transgenic mice that expressed a full-length BHLHB1 cDNA driven by the lck promoter. As anticipated, Northern blot analysis showed ectopic BHLHB1 expression in the thymus and bone marrow. We followed a total of 69 transgenic offspring from 3 independent founder lines, all of which showed expression of BHLHB1 in the thymus by Northern blot, for one year. None of these mice developed evidence of T-cell malignancy. Since SCL, like BHLHB1 a class B bHLH protein cooperates with LMO1 to induce pre-T LBL in mice, we considered the possibility that BHLHB1 might also cooperate with LMO1 to induce T-cell tumors. Therefore, we crossed the lck-BHLHB1 mice with lck-LMO1 mice. Approximately 47% (8/47) of BHLHB1-LMO1 double positive mice have developed pre-T LBL by 14 months, while 19% (3/16) of BHLHB1 single positive mice and 7% (1/15) of LMO1 single positive have developed pre-T LBL or died unexpectedly within this time period. The leukemia is highly aggressive, and displays massive thymic enlargement, diffuse lymphadenopathy, and organ infiltration (spleen, liver, kidney, and lung). The immunophenotype of the leukemic cells is CD3-positive and CD4/8-double positive, and Southern blot analysis of these tumors demonstrated clonal TCRB gene rearrangements. We have been able to culture cells from these tumors for over 6 months in the absence of supplemental cytokines, suggesting that these cells are immortal.

We compared the gene expression pattern of the T-cell tumors from BHLHB1-LMO1 double transgenic mice with that of normal thymus tissue using a long oligonucleotide set containing >20, 000 features. Several genes were consistently upregulated at least 4-fold in the tumor tissue, including several translation initiation/elongation factors, transferrin receptor, CCL8 (a chemokine ligand), BST1 (an antigen that lymphocyte growth), granzyme A (a product of cytolytic T-cells), and SAA3 (an acute phase reactant).

Several studies have shown that the LMO1 and LMO2 each collaborate with SCL to induce pre-T LBL in almost 100% of mice within 6 months. Similar to the situation seen with SCL, we have now demonstrated that LMO1 and BHLHB1 collaborate to induce an aggressive pre-T LBL in mice. Since BHLHB1 overexpression can inhibit E2A function, it seems reasonable to suggest that BHLHB1 is exerting its leukemogenic effect through an inhibition of E-proteins important for T-cell differentiation such as HEB or E2A.