Methylthioadenosine phosphorylase as target for chemoselective treatment of T-cell acute lymphoblastic leukemic cells
Methylthioadenosine phosphorylase as target for chemoselective treatment of T-cell acute lymphoblastic leukemic cells
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DOI：
10.1006/bcmd.2002.0483
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发表时间：
2002-01-01
影响因子：
2.3
通讯作者：
Gebhart, E
中科院分区：
文献类型：
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作者：
Efferth, T;Miyachi, H;Gebhart, E
我们分析了甲硫腺苷磷酸化酶（MTAP）在T细胞急性淋巴细胞白血病（T-ALL）化疗选择性治疗中的作用。MTAP将甲硫腺苷转化为腺嘌呤，其用作替代嘌呤源，如果从头嘌呤生物合成被抗代谢物抑制（即，甲氨蝶呤）。化疗选择性概念的想法是，在染色体9 p21处缺失MTAP的肿瘤比没有这种缺失的正常细胞更容易对抗代谢物敏感。首先，我们通过比较基因组杂交筛选了13个T-ALL细胞系的9 p21缺失。5个细胞系显示9号染色体短臂dim（9 p21 pter）缺失。进一步的分析用CEM细胞进行，其中9 p21缺失通过荧光原位杂交证实。用MTAP表达载体转染CEM细胞。共转染绿色荧光蛋白（GFP）质粒，流式细胞仪检测转染效率。使用生长抑制试验，与模拟对照转染子相比，MTAP转染细胞对抗代谢物甲氨蝶呤（MTX）、曲美蝶呤（TMX）和L-丙氨酸（ALA）的反应降低。多柔比星（DOX），这是不参与DNA生物合成的活性没有改变MTAP转染。由于p16（INK 4a）肿瘤抑制基因也位于9 p21，我们用p16 INK 4a表达载体转染CEM细胞。这些转染细胞对所有四种药物都具有更强的抗性，表明p16（INK 4a）并不特异性地影响抗代谢物。然而，在MTAP缺失的肿瘤细胞中，抗代谢物的化学选择性作用可以通过耐药性的发展来补偿。为了证明这种可能性，我们分析了MTX耐药亚系CEM/MTX 1500 LV，其中MTX耐药赋予二氢叶酸还原酶（DHFR）基因被扩增。TMX在CEM/MTX 1500 LV细胞中表现出相当大的交叉耐药性，而ALA则没有。因此，ALA可以在9 p21/MTAP缺失的细胞中表现出化学选择性，即使发生DHFR扩增。我们的结论是，ALA可能比MTX或TMX更适合MTAP介导的T-ALL的化学选择性治疗。治疗前检测9 p21和MTAP缺失可能有助于开发T-ALL的预测性分子化疗敏感性试验。(C)2002 Elsevier Science（美国）。
We analyzed the role of methylthioadenosine phosphorylase (MTAP) for chemoselective treatment of T-cell acute lymphoblastic leukemia (T-ALL). MTAP converts methylthioadenosine into adenine which serves as an alternative purine source, if de novo purine biosynthesis is inhibited by antimetabolites (i.e., methotrexate). The idea of the chemo selectivity concept is that tumors with MTAP deletion at chromosome 9p21 are more susceptible to antimetabolites than normal cells without such a deletion. First, we screened 13 T-ALL lines for 9p21 deletions by comparative genomic hybridization. Five cell lines revealed deletions at the short arm of chromosome 9, dim(9p21pter). Further analyses were performed with CEM cells in which the 9p21 deletion was corroborated by fluorescence in situ hybridization. CEM cells were transfected with an MTAP expression vector. A green fluorescent protein (GFP) plasmid was cotransfected, to monitor the transfection efficacy by flow cytometry. The response of MTAP-transfected cells to the antimetabolites methotrexate (MTX), trimetrexate (TMX), and L-alanosine (ALA) was decreased compared to mock control transfectants using growth inhibition assays. The activity of doxorubicin (DOX) which is not involved in DNA biosynthesis was not changed in MTAP transfectants. As the p16(INK4a) tumor suppressor gene resides also at 9p21, we transfected CEM cells with a p16INK4a expression vector. These transfectant cells were more resistant to all four drugs indicating that p16(INK4a), did not specifically affect antimetabolites. The chemoselective effect of antimetabolites in MTAP-deleted tumor cells may, however, be compensated by the development of drug resistance. To prove this possibility, we analyzed an MTX-resistant subline, CEM/MTX1500LV, in which the MTX-resistance conferring dihydrofolate reductase (DHFR) gene was amplified. While TMX exhibited considerable cross-resistance in CEM/MTX1500LV cells, ALA did not. Thus, ALA could exhibit chemoselectivity in 9p21/MTAP-deleted cells, even if DHFR amplification occurs. We conclude that ALA may be more suitable than MTX or TMX for MTAP-mediated chemoselective treatment of T-ALL. Pretherapeutical detection of 9p21 and MTAP deletion may be helpful in developing a predictive molecular chemosensitivity test for T-ALL. (C) 2002 Elsevier Science (USA).