Suppressive subtractive hybridisation reveals differential expression of serglycin, sorcin, bone marrow proteoglycan and prostate-tumour-inducing gene I (PTI-1) in drug-resistant and sensitive tumour cell lines of haematopoetic origin
Suppressive subtractive hybridisation reveals differential expression of serglycin, sorcin, bone marrow proteoglycan and prostate-tumour-inducing gene I (PTI-1) in drug-resistant and sensitive tumour cell lines of haematopoetic origin
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DOI：
10.1016/s0959-8049(99)00202-6
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发表时间：
1999-11-01
影响因子：
8.4
通讯作者：
Bertram, J
中科院分区：
文献类型：
--
作者：
Beyer-Sehlmeyer, G;Hiddemann, W;Bertram, J
治疗诱导的耐药性的发展仍然是最重要的治疗限制之一。然而，一个综合的观点，耐药发展的分子机制一般是失踪。为了阐明这种耐药性发展的网络，我们建立了造血来源的六种肿瘤细胞系（Jurkat，U937，HL 60，DoHH-2，K562和ARH 77）的耐药（多柔比星（DX），甲氨蝶呤（MTX），顺铂（cisPt），长春新碱（Vin））衍生物。通过抑制性消减杂交分析药物敏感亲本细胞系及其耐药衍生物的差异基因表达。在对克隆的PCR片段进行差异表达的斑点印迹筛选和测序后，通过北方印迹分析证实差异表达。为了区分仅与一种或另一种研究药物相关的差异表达基因，将各种耐药亚系（多柔比星耐药、甲氨蝶呤耐药、顺铂耐药Jurkat细胞）的cDNA合并，并与敏感的亲本细胞系进行比较。此外，合并不同造血肿瘤细胞系的耐药衍生物的cDNA，并与相应的敏感造血细胞系的合并cDNA进行比较，以消除与耐药性无关的细胞系间变异。作为该筛选的结果，以下基因在耐药变体中显示出更高（至少2倍）或排他性表达：sergycin、sorcin、BMPG（骨髓蛋白聚糖基因）和PTI-1（前列腺肿瘤诱导基因1）。此外，我们的小组先前发现在耐药结肠癌细胞系LoVo H67 P中上调的hsp 90的表达升高，发现在耐药HL 60细胞中过表达。(C)1999 Elsevier Science Ltd.保留所有权利。
The development of therapy-induced drug resistance is still one of the most important therapeutic limitations. Nevertheless, an integrating view of the molecular mechanisms underlying resistance development in general is missing. In order to shed some light on the network of this resistance development, we established drug-resistant (doxorubicin (DX), methotrexate (MTX), cisplatin (cisPt), vincristine (Vin)) derivatives of six tumour cell lines (Jurkat, U937, HL60, DoHH-2, K562 and ARH77) of haematopoetic origin. Differential gene expression of drug-sensitive parental cell lines and the drug-resistant derivatives thereof was analysed by suppressive subtractive hybridisation. After dot blot screening for differential expression and sequencing of the cloned PCR fragments, differential expression was confirmed by Northern blot analysis. In an attempt to discriminate for differentially expressed genes only related to one or the other of the investigated drugs, the cDNAs of various resistant sublines (doxorubicin-, methotrexate-, cisplatin-resistant Jurkat cells) were pooled and compared with the sensitive parental cell line. In addition, cDNAs of the resistant derivatives of the different haematopoetic tumour cell lines were pooled and compared with the pooled cDNAs of the corresponding sensitive haematopoetic cell lines to eliminate cell line to cell line variations that were not related to drug resistance. As a result of this screening, the following genes showed a higher (at least 2-fold) or exclusive expression in the drug-resistant variants: serglycin, sorcin, BMPG (bone marrow proteoglycan gene) and PTI-1 (prostate-tumour-inducing gene 1). In addition, elevated expression of hsp90, previously found by our group to be upregulated in the drug-resistant colon carcinoma cell line LoVo H67P was found to be overexpressed in drug-resistant HL60 cells. (C) 1999 Elsevier Science Ltd. All rights reserved.