Crespi C L, Penman B W, Gelboin H V, Gonzalez F J
GENTEST Corporation, Woburn, MA 01801.
Carcinogenesis. 1991 Jul;12(7):1197-201. doi: 10.1093/carcin/12.7.1197.
We have developed a human B-lymphoblastoid cell line, designated 2D6/Hol, which stably expresses human cytochrome P450 CYP2D6 cDNA. This cell line exhibits bufuralol 1'-hydroxylase activity and immunologically detectable CYP2D6 protein. The specific activity of (+)-bufuralol 1'-hydroxylase in microsomes from 2D6/Hol cells was comparable to that observed in human liver microsomes. This cell line was used to examine the mutagenicity activation of three tobacco smoke-derived nitrosamines, N-nitrosonornicotine (NNN), 1-(N-methyl-N-nitrosamino)-1-(3-pyridinyl)-4-butanal) (NNA) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), by CYP2D6. Exposure of 2D6/Hol cells to NNK concentrations of 30-90 micrograms/ml induced a concentration-dependent decrease in relative survival and increase in mutant fraction at the hypoxanthine guanine phosphoribosyl transferase (hprt) locus. In contrast, NNK was non-mutagenic and non-cytotoxic to control cells at exposure concentrations up to 150 micrograms/ml. NNK mutagenicity in 2D6/Hol cells was compared to the responses observed in isogenic cell lines expressing human CYP1A2 (1A2/Hol), human CYP2A3 (2A3/Hol) and human CYP2E1 (2E1/Hol). These three additional human cytochrome P450-expressing cell lines were also found to be sensitive to NNK-induced mutagenicity and cytotoxicity. We found no evidence for CYP2D6-mediated activation of NNN or NNA. NNN was non-cytotoxic and non-mutagenic to both control and 2D6/Hol cells. NNA was equally cytotoxic and mutagenic to control cells and 2D6/Hol cells. The activation of NNA to a mutagen may have been carried out by P450 native to the AHH-1 TK +/- cell line. The 2D6/Hol cell line, in conjunction with the control cell line and other isogenic cell lines expressing other human cytochrome P450 cDNAs provides a useful system for the examination of the role of the polymorphic CYP2D6 in human procarcinogen activation and drug metabolism.
我们构建了一种人B淋巴母细胞系，命名为2D6/Hol，它能稳定表达人细胞色素P450 CYP2D6 cDNA。该细胞系具有布呋洛尔1'-羟化酶活性，且能通过免疫检测到CYP2D6蛋白。2D6/Hol细胞微粒体中(+)-布呋洛尔1'-羟化酶的比活性与人肝微粒体中的相当。该细胞系用于检测细胞色素P450 CYP2D6对三种烟草烟雾衍生的亚硝胺，即N-亚硝基降烟碱（NNN）、1-(N-甲基-N-亚硝胺基)-1-(3-吡啶基)-4-丁醛（NNA）和4-(甲基亚硝胺基)-1-(3-吡啶基)-1-丁酮（NNK）的致突变激活作用。将2D6/Hol细胞暴露于浓度为30 - 90微克/毫升的NNK中，会导致次黄嘌呤鸟嘌呤磷酸核糖基转移酶（hprt）位点的相对存活率呈浓度依赖性下降，突变率增加。相比之下，在暴露浓度高达150微克/毫升时，NNK对对照细胞无致突变性且无细胞毒性。将2D6/Hol细胞中NNK的致突变性与在表达人CYP1A2（1A2/Hol）、人CYP2A3（2A3/Hol）和人CYP2E1（2E1/Hol）的同基因细胞系中观察到的反应进行比较。还发现这三种额外的表达人细胞色素P450的细胞系对NNK诱导的致突变性和细胞毒性也敏感。我们没有发现CYP2D6介导激活NNN或NNA的证据。NNN对对照细胞和2D6/Hol细胞均无细胞毒性和致突变性。NNA对对照细胞和2D6/Hol细胞的细胞毒性和致突变性相同。NNA向致突变物的激活可能是由AHH-1 TK +/-细胞系自身的P450进行的。2D6/Hol细胞系与对照细胞系以及其他表达其他人细胞色素P450 cDNA的同基因细胞系一起，为研究多态性CYP2D6在人类前致癌物激活和药物代谢中的作用提供了一个有用的系统。
