Nasedkina Tatyana V, Zharinov Vladislav S, Isaeva Ekaterina A, Mityaeva Olga N, Yurasov Roman N, Surzhikov Sergei A, Turigin Alexander Y, Rubina Alla Y, Karachunskii Alexander I, Gartenhaus Ronald B, Mirzabekov Andrei D
Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russia.
Clin Cancer Res. 2003 Nov 15;9(15):5620-9.
Currently, many forms of leukemia are considered potentially curable, with prognosis and clinical outcome strongly dependent on the underlying molecular pathophysiology. A substantial number of leukemia patients harbor nonrandom karyotypic abnormalities that define subgroups with unique biological and clinical features. For detection of these types of gene rearrangements, a combination of multiplex RT-PCR with hybridization on oligonucleotide gel array was presented previously, which identified five chromosomal translocations with fusion variants. In the present study, additional clinically relevant translocations were included in our analysis using a second generation of microarrays. We also expanded significantly on the clinical correlation of our findings.
An oligonucleotide microarray was designed for hybridization with products of a multiplex RT-PCR to identify the following translocations: t(9;22)p190, t(4;11), t(12;21), t(1;19), typical for acute lymphoblastic leukemia; t(9;22)p210 for chronic myeloid leukemia; and t(8;21), t(15;17), inv16, typical for acute myeloblastic leukemia.
To demonstrate the potential clinical application of the method, 247 cases of childhood leukemia were screened, and the above-mentioned gene rearrangements were found in 30% of cases. The sensitivity and specificity of the assay is comparable with the RT-PCR technique, so that it can be used to follow minimal residual disease. The feasibility of an additional refinement of the method, on-chip-multiplex PCR, has been successfully demonstrated by identifying a common translocation, t(9;22), in chronic myeloid leukemia.
Our data suggest that the microarray-based assay can be an effective and reliable tool in the clinical screening of leukemia patients for the presence of specific gene rearrangements with important diagnostic and prognostic implications. The method is amenable for automation and high-throughput analysis.
目前，许多形式的白血病被认为具有潜在的可治愈性，其预后和临床结果在很大程度上取决于潜在的分子病理生理学。大量白血病患者存在非随机核型异常，这些异常定义了具有独特生物学和临床特征的亚组。为了检测这些类型的基因重排，先前提出了多重逆转录聚合酶链反应（RT-PCR）与寡核苷酸凝胶阵列杂交相结合的方法，该方法鉴定出了五种带有融合变体的染色体易位。在本研究中，使用第二代微阵列在我们的分析中纳入了其他临床相关的易位。我们还在很大程度上扩展了研究结果的临床相关性。
设计了一种寡核苷酸微阵列，用于与多重RT-PCR产物杂交，以鉴定以下易位：急性淋巴细胞白血病典型的t(9;22)p190、t(4;11)、t(12;21)、t(1;19)；慢性髓性白血病的t(9;22)p210；以及急性髓性白血病典型的t(8;21)、t(15;17)、inv16。
为了证明该方法的潜在临床应用价值，对247例儿童白血病病例进行了筛查，在30%的病例中发现了上述基因重排。该检测方法的灵敏度和特异性与RT-PCR技术相当，因此可用于监测微小残留病。通过在慢性髓性白血病中鉴定出常见的易位t(9;22)，成功证明了该方法进一步改进即芯片上多重PCR的可行性。
我们的数据表明，基于微阵列的检测方法可以成为临床筛查白血病患者是否存在具有重要诊断和预后意义的特定基因重排的有效且可靠的工具。该方法适合自动化和高通量分析。
