Reduced folate carrier gene expression in childhood acute lymphoblastic leukemia: relationship to immunophenotype and ploidy.
Reduced folate carrier gene expression in childhood acute lymphoblastic leukemia: relationship to immunophenotype and ploidy.
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发表时间：
1998-09
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通讯作者：
Long Zhang;J. Taub;Michael S. Williamson;So C Wong;B. Hukku;J. Pullen;Y. Ravindranath;L. Matherly
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作者：
Long Zhang;J. Taub;Michael S. Williamson;So C Wong;B. Hukku;J. Pullen;Y. Ravindranath;L. Matherly
用竞争性聚合酶链式反应检测人白血病中的还原型叶酸载体(RFC)转录本。在RFC和β-肌动蛋白竞争性模板存在的情况下，对总RNA进行逆转录和扩增。将RFC转录本归一化为β-肌动蛋白转录本。在一系列K562亚系中，通过PCR检测的RFC转录本的范围约为30倍，与Northern分析的结果非常一致，并且与蛋白质印迹和[~3H]甲氨蝶呤转运的RFC蛋白的比例不同。在48例急性淋巴细胞白血病(ALL)患儿的49个样本中，RFC转录本的变化范围超过88倍。15例T细胞和33例B-前体ALL的RFC转录本的中位数相似(RFC/β-肌动蛋白分别为6.13×10(-3)和7.92×10(-3))，41例诊断(7.20×10(-3))和8例复发(5.58×10(-3))。而RFC转录本的聚合酶链式反应测量结果与B-前体ALL细胞(n=10)的氨甲喋呤转运变化相似，而对于T-ALL细胞(n=12)，这些参数之间没有明显的关系。超二倍体B前体母细胞(n=11)具有大于52条染色体和3-5个拷贝的21号染色体，其RFC转录水平中位数约为二倍体B前体母细胞的3倍。在三个带有获得性21三体的B-前体样本中，有两个的RFC转录水平也升高。我们的结果表明，RFC基因的表达比T-ALL更能预测B-前体细胞摄取甲氨蝶呤的能力，并且B-前体ALL细胞中21号染色体拷贝的增加通常与RFC转录本的增加有关。因此，超二倍体B-前体均接受以抗代谢药物为基础的化疗的儿童预后良好，甲氨蝶呤和甲氨蝶呤聚谷氨酸的高水平累积可能部分反映了RFC基因表达和甲氨蝶呤转运能力的提高。
Reduced folate carrier (RFC) transcripts in human leukemias were measured by a competitive PCR assay. Total RNAs were reverse transcribed and amplified in the presence of competitive templates for RFC and beta-actin. RFC transcripts were normalized to transcripts for beta-actin. In a series of K562 sublines, a approximately 30-fold range of RFC transcripts measured by PCR assay closely agreed with results of Northern analysis and varied in proportion to RFC protein on Western blots and [3H]methotrexate transport. RFC transcripts varied over a 88-fold range in 49 specimens from 48 children with acute lymphoblastic leukemia (ALL). Median RFC transcripts were similar for 15 T-cell and 33 B-precursor ALL samples (RFC/beta-actin = 6.13 x 10(-3) and 7.92 x 10(-3), respectively) and for 41 diagnostic (7.20 x 10(-3)) and 8 relapse (5.58 x 10(-3)) samples. Whereas PCR measurements of RFC transcripts approximated changes in methotrexate transport in B-precursor ALL blasts (n = 10), for T-ALL blasts (n = 12) there was no apparent relationship between these parameters. For hyperdiploid B-precursor blasts (n = 11) with greater than 52 chromosomes and three to five copies of chromosome 21, the median RFC transcript level was approximately 3-fold higher than that for diploid B-precursor blasts. RFC transcripts were also elevated for two of three B-precursor specimens with acquired trisomy 21. Our results suggest that RFC gene expression is far more predictive of methotrexate uptake capacity in B-precursor than T-ALL and that increased copies of chromosome 21 in B-precursor ALL blasts are generally associated with increased RFC transcripts. Hence, the good prognosis for children with hyperdiploid B-precursor ALL treated with antimetabolite-based chemotherapy and the high levels of methotrexate and methotrexate polyglutamates accumulated may, in part, reflect elevated RFC gene expression and capacities for methotrexate transport.