Yadollahi-Farsani M, Gooderham N J, Davies D S, Boobis A R
Department of Clinical Pharmacology, Royal Postgraduate Medical School, London, UK.
Carcinogenesis. 1996 Apr;17(4):617-24. doi: 10.1093/carcin/17.4.617.
The mutagenic 'fingerprint' of the cooked food carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) was determined in a Chinese hamster cell line genetically engineered to express human CYP1A2 (XEMh1A2-MZ). The parental Chinese hamster V79 and XEMh1A2-MZ cells were exposed to PhIP at various concentrations for 24h. There was a dose-dependent increase in frequency of mutations at the hypoxanthine-guanine phosphoribosyltransferase (hprt) locus only in the metabolically competent XEMh1A2-MZ cells. The mutant frequency ranged from 25 to 90 X 10(-6) with final concentrations of 2.5 to 100 microM PhIP compared to 8 X 10(-6) in the solvent controls and the V79MZ cells. The molecular nature of the PhIP-induced mutations in XEMh1A2-MZ cells was determined by examining DNA sequence modifications at the hprt locus in forty five 6-thioguanine resistant (6-TGr) mutant clones. Single base substitutions predominantly GC-->TA transversions, were the major class of PhIP-induced mutation. However, a -1 frameshift 'hotspot' in a 5'-GGGA sequence was also observed. With the exception of a compound modification, all of the PhIP-induced mutations involved G.C base pairs. This is consistent with the previously observed PhIP-induced mutations in cultured mammalian cells and 32P-postlabelling experiments that show PhIP adducts to the guanine base and that major adduct is at the C8 position. Furthermore, nearly all of these mutations involved guanine bases on the non-transcribed strand which is possibly indicative of preferential repair of PhIP adducts from the transcribed strand. Nearest neighbor analysis of induced base substitutions indicates a preference for 5' guanine and 3' adenine. These data effectively define a mutation 'fingerprint' for PhIP, which may provide the basis for definitive studies on the role of PhIP in diet associated cancers such as tumours of colon. It is, therefore, intriguing that in their recent report of mutation in tumours of the colon induced by PhIP in male rate Kakiuchi et al. (Proc. Natl Acad. Sci. USA, 92, 910-914) report that four out of eight tumors had identical mutation of the tumour suppressor gene apc which is comprised of a -1 G frameshift in a 5'-GGGA sequence.
在一种经基因工程改造以表达人CYP1A2的中国仓鼠细胞系（XEMh1A2 - MZ）中，测定了烹饪食物致癌物2 - 氨基 - 1 - 甲基 - 6 - 苯基咪唑并[4,5 - b]吡啶（PhIP）的诱变“指纹”。将亲本中国仓鼠V79细胞和XEMh1A2 - MZ细胞暴露于不同浓度的PhIP中24小时。仅在具有代谢活性的XEMh1A2 - MZ细胞中，次黄嘌呤 - 鸟嘌呤磷酸核糖转移酶（hprt）基因座处的突变频率呈剂量依赖性增加。与溶剂对照和V79MZ细胞中的8×10⁻⁶相比，终浓度为2.5至100μM的PhIP时，突变频率范围为25至90×10⁻⁶。通过检查45个6 - 硫鸟嘌呤抗性（6 - TGr）突变克隆中hprt基因座处的DNA序列修饰，确定了XEMh1A2 - MZ细胞中PhIP诱导突变的分子性质。单碱基取代主要是GC→TA颠换，是PhIP诱导突变的主要类型。然而，在一个5'-GGGA序列中也观察到一个 - 1移码“热点”。除了一种复合修饰外，所有PhIP诱导的突变都涉及G.C碱基对。这与之前在培养的哺乳动物细胞中观察到的PhIP诱导突变以及³²P后标记实验一致，这些实验表明PhIP与鸟嘌呤碱基形成加合物，且主要加合物位于C8位置。此外，几乎所有这些突变都涉及非转录链上的鸟嘌呤碱基，这可能表明优先从转录链修复PhIP加合物。对诱导碱基取代的邻位分析表明对5'鸟嘌呤和3'腺嘌呤有偏好。这些数据有效地定义了PhIP的突变“指纹”，这可能为确定PhIP在与饮食相关的癌症（如结肠癌）中的作用的确定性研究提供基础。因此，有趣的是，在Kakiuchi等人（《美国国家科学院院刊》，92，910 - 914）最近关于雄性大鼠中PhIP诱导的结肠癌肿瘤突变的报告中，他们报告说八分之四的肿瘤具有肿瘤抑制基因apc的相同突变，该突变由5'-GGGA序列中的一个 - 1 G移码组成。
