Laurenti Luca, Petlickovski Aleksandar, Rumi Carlo, Gobessi Stefania, Piccioni Paola, Tarnani Michela, Puggioni Pierluigi, Marietti Sara, Sica Simona, Leone Giuseppe, Efremov Dimitar G
Department of Hematology, Catholic University Hospial A. Gemelli, Rome, Italy.
Haematologica. 2005 Nov;90(11):1533-40.
The protein tyrosine kinase ZAP-70 has recently emerged as a major prognostic indicator in chronic lymphocytic leukemia (CLL). ZAP-70 is structurally and functionally homologous to Syk, a key mediator of B-cell receptor signaling. We therefore evaluated ZAP-70 expression in CLL B cells using Syk as an intracellular standard.
The relative amounts of ZAP-70 and Syk were determined in purified B cells from 92 CLL patients using a novel reverse transcriptase/polymerase chain reaction (RT-PCR) procedure that co-amplifies both transcripts with equal efficiency. The ZAP-70/Syk mRNA ratio was correlated with VH gene mutation status, median treatment-free survival and FACS analysis of ZAP-70 expression.
ZAP-70 was expressed in the majority of cases with unmutated VH genes (88%), but also at lower levels in a substantial fraction of cases with mutated VH genes (44%). High levels of ZAP-70, defined as ZAP-70/Syk mRNA ratios above 0.25, were observed mainly in cases with unmutated VH genes and correlated with short treatment-free survival. In contrast, no difference was observed in the median treatment-free survival between patients with low ZAP-70/Syk ratios (0.05-0.25) and patients with no or negligible ZAP-70 expression (ZAP-70/Syk<0.05). In 73 cases ZAP-70 expression was investigated by RT/PCR and FACS analysis; concordance with VH gene mutation status was 86% and 71%, respectively.
ZAP-70 is frequently expressed in CLL B cells, but only high levels correlate with unmutated VH gene status and progressive disease. Expression of ZAP-70 can be accurately assessed by analysis of the ZAP-70/Syk mRNA ratio, thus providing an alternative to FACS analysis.
蛋白酪氨酸激酶ZAP-70最近已成为慢性淋巴细胞白血病（CLL）的一项主要预后指标。ZAP-70在结构和功能上与Syk同源，Syk是B细胞受体信号传导的关键介质。因此，我们以Syk作为细胞内标准，评估了CLL B细胞中ZAP-70的表达。
采用一种新型逆转录酶/聚合酶链反应（RT-PCR）程序，以同等效率共扩增ZAP-70和Syk这两种转录本，测定了92例CLL患者纯化B细胞中ZAP-70和Syk的相对含量。ZAP-70/Syk mRNA比值与VH基因突变状态、中位无治疗生存期以及ZAP-70表达的流式细胞术分析相关。
大多数VH基因未突变的病例（88%）中表达ZAP-70，但相当一部分VH基因发生突变的病例（44%）中ZAP-70表达水平也较低。ZAP-70/Syk mRNA比值高于0.25被定义为ZAP-70高水平，主要在VH基因未突变的病例中观察到，且与较短的无治疗生存期相关。相比之下，ZAP-70/Syk比值低（0.05 - 0.25）的患者与无ZAP-70表达或ZAP-70表达可忽略不计（ZAP-70/Syk < 0.05）的患者，中位无治疗生存期无差异。在73例病例中，通过RT/PCR和流式细胞术分析研究了ZAP-70表达；与VH基因突变状态的一致性分别为86%和71%。
ZAP-70在CLL B细胞中经常表达，但只有高水平与VH基因未突变状态和疾病进展相关。通过分析ZAP-70/Syk mRNA比值可准确评估ZAP-70的表达，从而为流式细胞术分析提供了一种替代方法。
