Spasokoukotskaja T, Arnér E S, Brosjö O, Gunvén P, Juliusson G, Liliemark J, Eriksson S
Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.
Eur J Cancer. 1995;31A(2):202-8. doi: 10.1016/0959-8049(94)00435-8.
Deoxycytidine kinase (dCK) activates several clinically important drugs, including the recently developed antileukaemic compound 2-chlorodeoxyadenosine (CdA). The distribution of dCK in cells and tissues has previously been determined by activity measurements, which may be unreliable because of the presence of other enzymes with overlapping substrate specificities. Therefore we have measured dCK polypeptide levels in extracts of normal and malignant human peripheral blood mononuclear cells, gastrointestinal tissues and sarcomas, using a specific immunoblotting technique, as well as the phosphorylation of CdA in the same extracts. High levels of dCK were found in all major subpopulations of normal mononuclear leucocytes (120 +/- 19 ng dCK/mg protein) and in B-cell chronic lymphocytic leukaemia (81 +/- 30 ng/mg, n = 23). Hairy-cell leukaemia contained lower levels (28 +/- 23 ng/mg, n = 7), as did three samples of T-cell chronic lymphocytic leukaemia (18 +/- 14 ng/mg). Phytohaemagglutinin stimulation of normal lymphocytes did not lead to any substantial increase in either dCK activity or protein expression (less than 2.5-fold). The human CEM wt T-lymphoblastoid cell line contained 56 +/- 1 ng/dCK/mg protein, while in the CEM ddC50 and AraC8D mutants that lack dCK activity, no dCK polypeptide could be detected. In colon adenocarcinomas, the dCK content was significantly higher (20 +/- 9 ng/mg, n = 20) than in normal colon mucosa (8 +/- 3.5 ng/mg, n = 19, P < 0.05). A similar pattern of dCK expression was found in gastric adenocarcinomas (21 +/- 13 ng/mg, n = 5) and normal stomach mucosa (6 +/- 5 ng/mg, n = 5, P < 0.15). One leiomyosarcoma and one extra-skeletal osteosarcoma showed dCK levels comparable with those found in normal lymphocytes (84 +/- 6 and 109 +/- 4 ng/mg, respectively), while other sarcoma samples contained lower levels, comparable to the gastrointestinal adenocarcinomas (20 +/- 7 ng/mg, n = 12). Thus, dCK is expressed constitutively and predominantly in lymphoid cells, but it is also found in solid non-lymphoid tissues, with increased levels in malignant cells. The phosphorylation of CdA in crude extracts showed a close correlation to the dCK polypeptide level.
脱氧胞苷激酶（dCK）可激活多种具有临床重要性的药物，包括最近研发的抗白血病化合物2-氯脱氧腺苷（CdA）。此前，细胞和组织中dCK的分布是通过活性测定来确定的，由于存在其他具有重叠底物特异性的酶，这种方法可能并不可靠。因此，我们使用一种特异性免疫印迹技术，测定了正常和恶性人外周血单个核细胞、胃肠道组织及肉瘤提取物中的dCK多肽水平，以及相同提取物中CdA的磷酸化情况。在正常单核白细胞的所有主要亚群（120±19 ng dCK/ mg蛋白质）和B细胞慢性淋巴细胞白血病（81±30 ng/mg，n = 23）中发现了高水平的dCK。毛细胞白血病中的水平较低（28±23 ng/mg，n = 7），三个T细胞慢性淋巴细胞白血病样本也是如此（18±14 ng/mg）。用植物血凝素刺激正常淋巴细胞，dCK活性或蛋白质表达均未出现任何显著增加（小于2.5倍）。人CEM wt T淋巴母细胞系含有56±1 ng/dCK/mg蛋白质，而在缺乏dCK活性的CEM ddC50和AraC8D突变体中，未检测到dCK多肽。在结肠腺癌中，dCK含量（20±9 ng/mg，n = 20）显著高于正常结肠黏膜（8±3.5 ng/mg，n = 19，P < 0.05）。在胃腺癌（21±13 ng/mg，n = 5）和正常胃黏膜（6±5 ng/mg，n = 5，P < 0.15）中也发现了类似的dCK表达模式。一个平滑肌肉瘤和一个骨外骨肉瘤的dCK水平与正常淋巴细胞中的相当（分别为84±6和109±4 ng/mg），而其他肉瘤样本的水平较低，与胃肠道腺癌相当（20±7 ng/mg，n = 12）。因此，dCK在淋巴样细胞中组成性且主要表达，但在实体非淋巴组织中也有发现，在恶性细胞中水平升高。粗提物中CdA的磷酸化与dCK多肽水平密切相关。
