Department of Analytical, Environmental and Forensic Sciences, MRC-PHE Centre for Environment and Health, King's College London, London, UK.
Center for Health Protection, National Institute for Public Health and the Environment (RIVM), Bilthoven, The Netherlands.
Environ Mol Mutagen. 2021 Apr;62(4):252-264. doi: 10.1002/em.22429. Epub 2021 Mar 15.
TP53 harbors somatic mutations in more than half of human tumors with some showing characteristic mutation spectra that have been linked to environmental exposures. In bladder cancer, a unique distribution of mutations amongst several codons of TP53 has been hypothesized to be caused by environmental carcinogens including 4-aminobiphenyl (4-ABP). 4-ABP undergoes metabolic activation to N-hydroxy-4-aminobiphenyl (N-OH-4-ABP) and forms pre-mutagenic adducts in DNA, of which N-(deoxyguanosin-8-yl)-4-ABP (dG-C8-4-ABP) is the major one. Human TP53 knock-in mouse embryo fibroblasts (HUFs) are a useful model to study the influence of environmental carcinogens on TP53-mutagenesis. By performing the HUF immortalization assay (HIMA) TP53-mutant HUFs are generated and mutations can be identified by sequencing. Here we studied the induction of mutations in human TP53 after treatment of primary HUFs with N-OH-4-ABP. In addition, mutagenicity in the bacterial lacZ reporter gene and the formation of dG-C8-4-ABP, measured by P-postlabelling analysis, were determined in N-OH-4-ABP-treated primary HUFs. A total of 6% TP53-mutants were identified after treatment with 40 μM N-OH-4-ABP for 24 hr (n = 150) with G>C/C>G transversion being the main mutation type. The mutation spectrum found in the TP53 gene of immortalized N-OH-4-ABP-treated HUFs was unlike the one found in human bladder cancer. DNA adduct formation (~40 adducts/10 nucleotides) was detected after 24 hr treatment with 40 μM N-OH-4-ABP, but lacZ mutagenicity was not observed. Adduct levels decreased substantially (sixfold) after a 24 hr recovery period indicating that primary HUFs can efficiently repair the dG-C8-4-ABP adduct possibly before mutations are fixed. In conclusion, the observed difference in the N-OH-4-ABP-induced TP53 mutation spectrum to that observed in human bladder tumors do not support a role of 4-ABP in human bladder cancer development.
TP53 基因在超过一半的人类肿瘤中存在体细胞突变，其中一些具有与环境暴露相关的特征性突变谱。在膀胱癌中，TP53 中几个密码子的突变分布被假设是由环境致癌物引起的，包括 4-氨基联苯（4-ABP）。4-ABP 经过代谢激活生成 N-羟基-4-氨基联苯（N-OH-4-ABP），并在 DNA 中形成前诱变加合物，其中 N-（脱氧鸟嘌呤-8-基）-4-ABP（dG-C8-4-ABP）是主要的一种。人 TP53 基因敲入小鼠胚胎成纤维细胞（HUFs）是研究环境致癌物对 TP53 突变影响的有用模型。通过进行 HUF 永生化测定（HIMA），生成 TP53 突变的 HUFs，然后通过测序鉴定突变。在这里，我们研究了用 N-OH-4-ABP 处理原代 HUFs 后人类 TP53 中的突变诱导。此外，还通过 P-后标记分析测定了 N-OH-4-ABP 处理的原代 HUFs 中细菌 lacZ 报告基因的诱变活性和 dG-C8-4-ABP 的形成。用 40μM N-OH-4-ABP 处理 24 小时后，共鉴定出 6%的 TP53 突变体（n=150），主要的突变类型为 G>C/C>G 颠换。在 N-OH-4-ABP 处理的永生化 HUFs 的 TP53 基因中发现的突变谱与人类膀胱癌中的不同。在用 40μM N-OH-4-ABP 处理 24 小时后检测到约 40 个加合物/10 个核苷酸的 DNA 加合物形成，但未观察到 lacZ 诱变活性。在 24 小时恢复后，加合物水平显著降低（六倍），表明原代 HUFs 可以有效地修复 dG-C8-4-ABP 加合物，可能在突变固定之前。总之，与在人类膀胱癌中观察到的 N-OH-4-ABP 诱导的 TP53 突变谱不同，这并不支持 4-ABP 在人类膀胱癌发展中的作用。
