Functional analysis of ETV6/ARG gene develop differentiation therapy for leukemias

ETV6/TEL基因是一种转录因子，是ETS家族成员之一。该基因已被报道与PDGFRb、MDS1/EVI1、BTL、ACS2、STL、JAK2、ABL、CDX2、TrkC、AML1、MN1、Pax5等多种基因融合。其中，PDGFRb、ABL、JAK2和TrkC是酪氨酸激酶(TK)。我们认为，ETV6融合蛋白的功能分析将有助于了解白血病的发生机制。我们已经在AML-M3患者的AML-M3细胞系中发现了一个新的ETV6伴侣基因ARG(ABL相关基因或ABL2)，另一个TK基因是从AML-M3患者的AT(15；17)(q22；q11.2)和at(1；12)(q25；p13)建立的。我们对该融合蛋白的功能进行了分析，并证明该融合蛋白对BA/F3细胞具有IL3非依赖性生长，对大鼠1细胞具有锚定非依赖性生长活性，表明该蛋白具有与其他酪氨酸激酶/TETV6蛋白相似的致癌活性。进一步的研究表明，ETV6/Arg癌蛋白…MORE通过类似的信号分子包括PI3K、SHC、ras-GAP和CRK-L，触发与激活的ABL癌基因相关的一些相同的信号通路。虽然ABL与多种人类恶性肿瘤有关，但尽管ARG与ABL有很高的同源性，但尚未有与人类恶性肿瘤相关的报道。因此，这是第一个表明ARG参与人类白血病的报道，ETV6/ARG蛋白具有致癌活性。到目前为止，ETV6/ARG融合基因已在三种类型的白血病(M3，M4eo和T-ALL)中被报道。在从M3患者建立的HT93细胞系中，在全反式维甲酸和细胞因子(IL3、GCSF、GM-CSF)的培养下，观察到向中性粒细胞、嗜酸性粒细胞和嗜碱性粒细胞的独特分化。在T-ALL细胞系中，观察到在细胞因子(IL3、G-CSF和GM-CSF)的培养下向髓系细胞的惊人分化。综上所述，ETV6/ARG蛋白可能在某些细胞因子的作用下，在特定条件下具有分化能力。因此，我们开始检测ETV6/ARG蛋白的分化能力。将ETV6/ARG载体导入K562或HL60细胞后，继续观察细胞形态的变化。当ETV6/ARG构建完整后，未观察到细胞形态的改变。现在，我们正在用ETV6 A 5/ARG构建的缺失ETV6 EX的载体进行类似的实验。5.更少

The ETV6/TEL gene is a transcription factor, one of the ETS family members. This gene has been reported to fuse to a variety of genes, PDGFRB, MDSl/EVI1, BTL, ACS2, STL, JAK2, ABL, CDX2, TRKC, AML1, MN1, Pax5 and so on. Among them, PDGFRB, ABL, JAK2 and TRKC are tyrosine kinases (TK). We believe that function analysis of ETV6 fusion proteins should contribute much to understanding mechanism of leukemogenesis. We already identified a novel ETV6 partner gene, ARG (ABL related gene or ABL2), another TK gene in a cell line established from an AML-M3 patient with at(15 ; 17)(q22 ; q1 1.2) and at(1 ; 12)(q25 ; p13). We have analysed function of this tyrosine kinase/ETV6 fusion protein, and have demonstrated that this protein confers IL3 independent growth to Ba/F3 cells, and anchorage independent growth activity to Rat- 1 cells, indicating that this protein possesses oncogenic activity similar to the other tyrosine kinase/TETV6 proteins. Further study has shown that the ETV6/ARG oncoprotein … More triggers some of the same signaling pathways associated with activated ABL oncogenes through similar signaling molecules including PI3K, SHC, ras-GAP and CRK-L. Although the ABL is well known to be involved in various human malignancies, ARG has not been reported to be involved in human malignancies despite of its high homology to ABL. Thus, this is the first report showing involvement of ARG in human leukemia, and that ETV6/ARG protein has oncogenic activity.ETV6/ARG fusion gene has been reported in three types of leukemias (M3, M4eo and T-ALL), so far. In the HT93Acell line established from a M3 patient, unique differentiation to neutrophils, eosinophils and basophils is observed in culture with all-trans retinoic acid and cytokines (IL3, GCSF, GM-CSF). In T-ALL cell line, a surprising differentiation to myeloid cells is observed in culture with cytokines (IL3, G-CSF, and GM-CSF). Taken together, ETV6/ARG protein is likely to possess differentiation capacity under specific condition with some cytokines. Therefore, we started to examine differentiation capacity of ETV6/ARG protein. After transfection of ETV6/ARG constructs to K562 or HL60, we continued to observe change of cell morphology. With full ETV6/ARG construct transfection, cell morphological change has not been observed. Now, we are doing the similar experiment with ETV6 A 5/ARG construct which has deletion of ETV6 ex. 5. Less