Mutagenicity of 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N–OH-PhIP) in human TP53 knock-in (Hupki) mouse embryo fibroblasts

Lisa Hölzl-Armstrong; Sarah Moody; Jill E. Kucab; Edwin P. Zwart; Medjda Bellamri; Mirjam Luijten; Robert J. Turesky; Michael R. Stratton; Volker M. Arlt; David H. Phillips

doi:10.1016/j.fct.2020.111855

Mutagenicity of 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N–OH-PhIP) in human TP53 knock-in (Hupki) mouse embryo fibroblasts

Lisa Hölzl-Armstrong, Sarah Moody, Jill E. Kucab, Edwin P. Zwart, Medjda Bellamri, Mirjam Luijten, Robert J. Turesky, Michael R. Stratton, Volker M. Arlt^*, David H. Phillips

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

4 Citations (Scopus)

Abstract

2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is a possible human carcinogen formed in cooked fish and meat. PhIP is bioactivated by cytochrome P450 enzymes to form 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N–OH-PhIP), a genotoxic metabolite that reacts with DNA leading to the mutation-prone DNA adduct N-(deoxyguanosin-8-yl)-PhIP (dG-C8-PhIP). Here, we studied N–OH-PhIP-induced whole genome mutagenesis in human TP53 knock-in (Hupki) mouse embryo fibroblasts (HUFs) immortalised and subjected to whole genome sequencing (WGS). In addition, mutagenicity of N–OH-PhIP in TP53 and the lacZ reporter gene were assessed. TP53 mutant frequency in HUF cultures treated with N–OH-PhIP (2.5 μM for 24 h, n = 90) was 10% while no TP53 mutations were found in untreated controls (DMSO for 24 h, n = 6). All N–OH-PhIP-induced TP53 mutations occurred at G:C base pairs with G > T/C > A transversions accounting for 58% of them. TP53 mutations characteristic of those induced by N–OH-PhIP have been found in human tumours including breast and colorectal, which are cancer types that have been associated with PhIP exposure. LacZ mutant frequency increased 25-fold at 5 μM N–OH–PHIP and up to ~350 dG-C8-PhIP adducts/10⁸ nucleosides were detected by ultra-performance liquid chromatography-electrospray ionisation multistage scan mass spectrometry (UPLC-ESI-MS³) at this concentration. In addition, a WGS mutational signature defined by G > T/C > A transversions was present in N–OH-PhIP-treated immortalised clones, which showed similarity to COSMIC SBS4, 18 and 29 signatures found in human tumours.

Original language	English
Article number	111855
Journal	Food and Chemical Toxicology
Volume	147
DOIs	https://doi.org/10.1016/j.fct.2020.111855
Publication status	Published - Jan 2021

Keywords

Dietary carcinogen
DNA adducts
Mutation
PhIP
TP53
Whole genome sequencing

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1016/j.fct.2020.111855

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Hölzl-Armstrong, L., Moody, S., Kucab, J. E., Zwart, E. P., Bellamri, M., Luijten, M., Turesky, R. J., Stratton, M. R., Arlt, V. M., & Phillips, D. H. (2021). Mutagenicity of 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N–OH-PhIP) in human TP53 knock-in (Hupki) mouse embryo fibroblasts. Food and Chemical Toxicology, 147, Article 111855. https://doi.org/10.1016/j.fct.2020.111855

@article{07de7448675f4a11b2fa9d23954db388,

title = "Mutagenicity of 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N–OH-PhIP) in human TP53 knock-in (Hupki) mouse embryo fibroblasts",

abstract = "2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is a possible human carcinogen formed in cooked fish and meat. PhIP is bioactivated by cytochrome P450 enzymes to form 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N–OH-PhIP), a genotoxic metabolite that reacts with DNA leading to the mutation-prone DNA adduct N-(deoxyguanosin-8-yl)-PhIP (dG-C8-PhIP). Here, we studied N–OH-PhIP-induced whole genome mutagenesis in human TP53 knock-in (Hupki) mouse embryo fibroblasts (HUFs) immortalised and subjected to whole genome sequencing (WGS). In addition, mutagenicity of N–OH-PhIP in TP53 and the lacZ reporter gene were assessed. TP53 mutant frequency in HUF cultures treated with N–OH-PhIP (2.5 μM for 24 h, n = 90) was 10% while no TP53 mutations were found in untreated controls (DMSO for 24 h, n = 6). All N–OH-PhIP-induced TP53 mutations occurred at G:C base pairs with G > T/C > A transversions accounting for 58% of them. TP53 mutations characteristic of those induced by N–OH-PhIP have been found in human tumours including breast and colorectal, which are cancer types that have been associated with PhIP exposure. LacZ mutant frequency increased 25-fold at 5 μM N–OH–PHIP and up to ~350 dG-C8-PhIP adducts/108 nucleosides were detected by ultra-performance liquid chromatography-electrospray ionisation multistage scan mass spectrometry (UPLC-ESI-MS3) at this concentration. In addition, a WGS mutational signature defined by G > T/C > A transversions was present in N–OH-PhIP-treated immortalised clones, which showed similarity to COSMIC SBS4, 18 and 29 signatures found in human tumours.",

keywords = "Dietary carcinogen, DNA adducts, Mutation, PhIP, TP53, Whole genome sequencing",

author = "Lisa H{\"o}lzl-Armstrong and Sarah Moody and Kucab, {Jill E.} and Zwart, {Edwin P.} and Medjda Bellamri and Mirjam Luijten and Turesky, {Robert J.} and Stratton, {Michael R.} and Arlt, {Volker M.} and Phillips, {David H.}",

note = "Funding Information: Lisa H{\"o}lzl-Armstrong was supported by a PhD studentship from the MRC Centre for Environment and Health. Work at King's College London and the Wellcome Trust Sanger Institute was supported by the Cancer Research UK Grand Challenge Award “Mutographs of Cancer” (grant C98/A24032 ). Other funding was from the National Cancer Institute of the National Institutes of Health ( R01CA122320 to R.J.T.); Mass spectrometry was carried out in the Analytical Biochemistry Shared Resource of the Masonic Cancer Center, University of Minnesota , funded in part by Cancer Center Support ( CA-077598 ). Publisher Copyright: {\textcopyright} 2020 Elsevier Ltd Copyright: Copyright 2021 Elsevier B.V., All rights reserved.",

year = "2021",

month = jan,

doi = "10.1016/j.fct.2020.111855",

language = "English",

volume = "147",

journal = "Food and Chemical Toxicology",

issn = "0278-6915",

publisher = "Elsevier",

}

TY - JOUR

T1 - Mutagenicity of 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N–OH-PhIP) in human TP53 knock-in (Hupki) mouse embryo fibroblasts

AU - Hölzl-Armstrong, Lisa

AU - Moody, Sarah

AU - Kucab, Jill E.

AU - Zwart, Edwin P.

AU - Bellamri, Medjda

AU - Luijten, Mirjam

AU - Turesky, Robert J.

AU - Stratton, Michael R.

AU - Arlt, Volker M.

AU - Phillips, David H.

N1 - Funding Information: Lisa Hölzl-Armstrong was supported by a PhD studentship from the MRC Centre for Environment and Health. Work at King's College London and the Wellcome Trust Sanger Institute was supported by the Cancer Research UK Grand Challenge Award “Mutographs of Cancer” (grant C98/A24032 ). Other funding was from the National Cancer Institute of the National Institutes of Health ( R01CA122320 to R.J.T.); Mass spectrometry was carried out in the Analytical Biochemistry Shared Resource of the Masonic Cancer Center, University of Minnesota , funded in part by Cancer Center Support ( CA-077598 ). Publisher Copyright: © 2020 Elsevier Ltd Copyright: Copyright 2021 Elsevier B.V., All rights reserved.

PY - 2021/1

Y1 - 2021/1

N2 - 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is a possible human carcinogen formed in cooked fish and meat. PhIP is bioactivated by cytochrome P450 enzymes to form 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N–OH-PhIP), a genotoxic metabolite that reacts with DNA leading to the mutation-prone DNA adduct N-(deoxyguanosin-8-yl)-PhIP (dG-C8-PhIP). Here, we studied N–OH-PhIP-induced whole genome mutagenesis in human TP53 knock-in (Hupki) mouse embryo fibroblasts (HUFs) immortalised and subjected to whole genome sequencing (WGS). In addition, mutagenicity of N–OH-PhIP in TP53 and the lacZ reporter gene were assessed. TP53 mutant frequency in HUF cultures treated with N–OH-PhIP (2.5 μM for 24 h, n = 90) was 10% while no TP53 mutations were found in untreated controls (DMSO for 24 h, n = 6). All N–OH-PhIP-induced TP53 mutations occurred at G:C base pairs with G > T/C > A transversions accounting for 58% of them. TP53 mutations characteristic of those induced by N–OH-PhIP have been found in human tumours including breast and colorectal, which are cancer types that have been associated with PhIP exposure. LacZ mutant frequency increased 25-fold at 5 μM N–OH–PHIP and up to ~350 dG-C8-PhIP adducts/108 nucleosides were detected by ultra-performance liquid chromatography-electrospray ionisation multistage scan mass spectrometry (UPLC-ESI-MS3) at this concentration. In addition, a WGS mutational signature defined by G > T/C > A transversions was present in N–OH-PhIP-treated immortalised clones, which showed similarity to COSMIC SBS4, 18 and 29 signatures found in human tumours.

AB - 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is a possible human carcinogen formed in cooked fish and meat. PhIP is bioactivated by cytochrome P450 enzymes to form 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N–OH-PhIP), a genotoxic metabolite that reacts with DNA leading to the mutation-prone DNA adduct N-(deoxyguanosin-8-yl)-PhIP (dG-C8-PhIP). Here, we studied N–OH-PhIP-induced whole genome mutagenesis in human TP53 knock-in (Hupki) mouse embryo fibroblasts (HUFs) immortalised and subjected to whole genome sequencing (WGS). In addition, mutagenicity of N–OH-PhIP in TP53 and the lacZ reporter gene were assessed. TP53 mutant frequency in HUF cultures treated with N–OH-PhIP (2.5 μM for 24 h, n = 90) was 10% while no TP53 mutations were found in untreated controls (DMSO for 24 h, n = 6). All N–OH-PhIP-induced TP53 mutations occurred at G:C base pairs with G > T/C > A transversions accounting for 58% of them. TP53 mutations characteristic of those induced by N–OH-PhIP have been found in human tumours including breast and colorectal, which are cancer types that have been associated with PhIP exposure. LacZ mutant frequency increased 25-fold at 5 μM N–OH–PHIP and up to ~350 dG-C8-PhIP adducts/108 nucleosides were detected by ultra-performance liquid chromatography-electrospray ionisation multistage scan mass spectrometry (UPLC-ESI-MS3) at this concentration. In addition, a WGS mutational signature defined by G > T/C > A transversions was present in N–OH-PhIP-treated immortalised clones, which showed similarity to COSMIC SBS4, 18 and 29 signatures found in human tumours.

KW - Dietary carcinogen

KW - DNA adducts

KW - Mutation

KW - PhIP

KW - TP53

KW - Whole genome sequencing

UR - http://www.scopus.com/inward/record.url?scp=85098523565&partnerID=8YFLogxK

U2 - 10.1016/j.fct.2020.111855

DO - 10.1016/j.fct.2020.111855

M3 - Article

C2 - 33189884

AN - SCOPUS:85098523565

SN - 0278-6915

VL - 147

JO - Food and Chemical Toxicology

JF - Food and Chemical Toxicology

M1 - 111855

ER -

Mutagenicity of 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N–OH-PhIP) in human TP53 knock-in (Hupki) mouse embryo fibroblasts

Abstract

Keywords

UN SDGs

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