A NOTCH1-Independant Pathway of c-Myc Oncogenesis in TAL1⁺ Human T-ALL.

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignancy of developing thymocytes afflicting both children and adults. Although the outcome has significantly improved over the past decades, further advances in targeted therapy will require an accurate stratification of T-ALL subtypes based on the precise understanding of the mechanisms involved. TAL1 is among the most frequently deregulated oncogenes in T-ALL. Studies on TAL1 transgenic animals have recently shown that leukemic growth and survival is largely dependent on the NOTCH1-controlled c-Myc pathway. We recently identified a particular aggressive paediatric T-ALL with t(1;14)(p32;q11)-mediated TAL1 overexpression and, in agreement with a central role of c-Myc in human T-ALL, displaying high levels of c-Myc transcripts. However, the absence of NOTCH1/FBW7 mutations in this case suggested an alternative c-Myc deregulation process. Multicolour FISH analysis indeed revealed a cryptic t(7;8)(q34;q24) within the same t(1;14)⁺ T-ALL clone, and breakpoint cloning confirmed the juxtaposition of c-Myc to the TCRβ locus. To further investigate the likely scenario of neoplastic development in this patient, we undertook a detailed geno/phenotyping of the tumor sample. In good agreement with the proposed oncogenic role of TAL1 in blocking thymocytes at late stages of maturation, immunophenotypic, transcriptional and genotype profiles clearly converged towards a maturation arrest of the malignant clone at the late cortical CD8 SP stage. By contrast, analysis of translocation breakpoints indicated that both events resulted from V(D)J recombination mistakes, and congruently occurred during the early stages of TCRδ and TCRβ rearrangements (DN2/DN3). Thus, the activation of both c-Myc and TAL1 was clearly decoupled from maturation arrest. As thymocyte progression from DN3 to SP includes extensive proliferation at the ISP/DP transition before rearrangement of the TCRα chain, the tumor clone was expected to show polyclonal TCRα rearrangements. Instead, LR-LMPCR and TCRα multiplex PCR revealed the presence of a monoclonal TCRα rearrangement. Thus, out of hundreds of thymocytes carrying t(1;14) and t(7;8), only one progressed towards malignant transformation after the onset of TCRα rearrangements, suggesting the necessity of yet a “3^rd hit”. Most importantly, this indicates that in absence of NOTCH1, TAL1 and c-Myc cooperation was not sufficient to trigger aggressive proliferation. One likely candidate for this 3^rd hit is LMO2, which we also found significantly over-expressed in the tumour clone. As previously underlined in animal models, the synergy between LMO2, TAL1 and c-MYC oncogenes likely accounts for the aggressive tumor phenotype in this patient, and illustrates the complex cooperative oncogenic pathways in human T-ALL pathogenesis. Intriguingly, the lack of major mediastin involvement in a clone with such a fulgurating growth raises the alternative possibility that proliferation was actually fired in the periphery through cognate triggering of the expressed TCRαβ, and thus that proliferation is uncoupled from differentiation block.