Development of gene expression signatures for TLX1 overexpression in human thymus C34+ T-cell progenitors cultured on an OP9-DLL1 feeder layer

The T-cell leukemia homeobox 1 (TLX1, HOX11) transcription factor is critically involved in the multistep pathogenesis of T-cell acute lymphoblastic leukemia (T-ALL) and often cooperates with NOTCH1 activation during malignant T-cell transformation. However, the exact molecular mechanisms by which these T-cell specific oncogenes cooperate during transformation remain to be established. Here, we used an integrative genomics approach to show that the oncogenic properties of TLX1 are mediated by genome-wide interference with the ETS1 and RUNX1 transcription factors. Partial disruption of ETS1 and RUNX1 activity by ectopic TLX1 expression in immature thymocytes drives repression of T-cell specific super-enhancers and mediates an unexpected transcriptional antagonism with NOTCH1 signaling. These phenomena coordinately trigger a TLX1 driven pre-leukemic phenotype in human thymic precursor cells, which corresponds with the in vivo thymic regression observed in murine TLX1 tumor models, and creates a strong genetic pressure for acquiring activating NOTCH1 mutations as a prerequisite for full leukemic transformation. In conclusion, our results uncover a functional antagonism between cooperative oncogenes during the earliest phases of tumor development and provide novel insights in the multistep pathogenesis of TLX1 driven human leukemia. Gene expression was measured after TLX1 overexpression in human CD34+ T-cell progenitors cultured on an OP9-DLL1 feeder layer. Cells were collected after 72h of co-culture. This was performed for 2 independent thymus CD34+ donors.

T细胞白血病同源框1（TLX1, HOX11）转录因子在T细胞急性淋巴细胞白血病（T-ALL）的多步骤发病机制中发挥关键作用，且在恶性T细胞转化过程中常与NOTCH1激活协同发挥功能。然而，这类T细胞特异性癌基因在转化过程中协同作用的确切分子机制仍有待阐明。本研究借助整合基因组学方法证实，TLX1的致癌特性是通过在全基因组范围内干扰ETS1与RUNX1转录因子的功能而介导的。在未成熟胸腺细胞中异位表达TLX1，可部分削弱ETS1与RUNX1的活性，进而抑制T细胞特异性超级增强子（super-enhancers）的表达，并意外介导与NOTCH1信号通路的转录拮抗作用。上述现象协同触发了人胸腺前体细胞中TLX1驱动的白血病前期表型，该表型与小鼠TLX1肿瘤模型中观察到的体内胸腺退化现象一致，同时施加了强大的遗传选择压力，促使细胞获得激活型NOTCH1突变——这是实现完全白血病转化的必要前提。综上，本研究揭示了协同癌基因在肿瘤发生最早期阶段的功能拮抗作用，为阐明TLX1驱动的人类白血病多步骤发病机制提供了全新视角。本研究在培养于OP9-DLL1饲养层的人CD34+ T细胞祖细胞中过表达TLX1后检测基因表达水平，共培养72小时后收集细胞，实验针对2名独立的胸腺CD34+供体完成。