Effects of Chemopreventive Natural Compounds on the Accuracy of 8-oxo-7,8-dihydro-2′-deoxyguanosine Translesion Synthesis^{[ # ]}

• Abstract
• Introduction
• Results
• Discussion
• Comparison of the 2 analytical methods
• Modulation of 8-oxodG TLS by phytochemicals
• Modulation of 8-oxodG TLS: possible mechanisms and consequences in cancer chemoprevention and therapy
• Material and Methods
• Chemicals
• Cell culture
• Primer extension assay
• Influence of phytochemicals on 8-oxodG TLS bypass
• Statistical analysis
• Contributorsʼ Statement
• References

Abstract

Translesion synthesis is a DNA damage tolerance mechanism that relies on a series of specialized DNA polymerases able to bypass a lesion on a DNA template strand during replication or post-repair synthesis. Specialized translesion synthesis DNA polymerases pursue replication by inserting a base opposite to this lesion, correctly or incorrectly depending on the lesion nature, involved DNA polymerase(s), sequence context, and still unknown factors. To measure the correct or mutagenic outcome of 8-oxo-7,8-dihydro-2′-deoxyguanosine bypass by translesion synthesis, a primer-extension assay was performed in vitro on a template DNA bearing this lesion in the presence of nuclear proteins extracted from human intestinal epithelial cells (FHs 74 Int cell line); the reaction products were analyzed by both denaturing capillary electrophoresis (to measure the yield of translesion elongation) and pyrosequencing (to determine the identity of the nucleotide inserted in front of the lesion). The influence of 14 natural polyphenols on the correct or mutagenic outcome of translesion synthesis through 8-oxo-7,8-dihydro-2′-deoxyguanosine was then evaluated in 2 experimental conditions by adding the polyphenol either (i) to the reaction mix during the primer extension assay; or (ii) to the culture medium, 24 h before cell harvest and nuclear proteins extraction. Most of the tested polyphenols significantly influenced the outcome of translesion synthesis, either through an error-free (apigenin, baicalein, sakuranetin, and myricetin) or a mutagenic pathway (epicatechin, chalcone, genistein, magnolol, and honokiol).

Introduction

Although some cancers are directly related to heredity, most of them are linked to molecular and cellular modifications accumulated in the course of life, arising through DNA damage, either from the endogenous metabolism or from exposure to environmental agents, workplace hazards, and life habits [1], [2]. Carcinogenesis is classically divided into 3 main steps: (i) the initiation that results from a cell exposure to 1 or several genotoxic agents of chemical, physical, or biological (pathogens) origins; (ii) the promotion, during which an initiated cell granted with a selective growth advantage proliferates and forms a preneoplastic focus; (iii) the progression that consists in the clonal expansion of these cells, with an increase of metastatic potential and angiogenesis [3], [4]. In addition, it is increasingly accepted that cancers can also result from epigenetic modifications (DNA methylation, histone modification, post-transcriptional regulation by microRNA). These modifications induce reversible changes in the expression of genes without DNA sequence alteration [5], [6].

According to a joint report of WHO and IARC [7], [8], 18.1 million new cases and 9.6 million deaths linked to cancer were recorded in 2018. During the 2 following decades, the number of new cases is expected to increase by 50% to reach 27 million per year in 2040. This report also considers that 40% of new cancer cases could be avoided by changes in life habits (a diet rich in fruits and vegetables, regular physical activity, etc.) and by avoiding risk factors (tobacco, pollution, overweight, ionizing and non-ionizing radiations, etc.).

While he was studying vitamin A analogs for carcinogenesis prevention, Dr. Michael Sporn defined in 1976 the term “chemoprevention” as “the strategy that uses natural, synthetic or biological agents to reverse, suppress or prevent either the initial phases of carcinogenesis or the progression of premalignant cells to invasive disease” [9]. The definition has been further refined, now designating the use of natural or pharmacological agents, relatively nontoxic, that inhibit cancer development by either blocking DNA damages that initiate carcinogenesis or stopping/inverting premalignant cell progression. These agents may act by affecting cellular and molecular events implicated in the 3 steps of carcinogenesis or epigenetic alterations [10], [11], [12], [13]. Chemopreventive agents are usually divided into 2 groups: (i) blocking agents that prevent carcinogens from reaching their target, undergoing a metabolic activation, or interacting with macromolecules (DNA, RNA, and proteins) (“barrier” function); and (ii) suppressing agents that inhibit the malignant transformation of initiated cells by interfering with the promotion or progression steps [11], [13], [14]. Blocking agents mainly act on transmembrane transports, elimination of reactive oxygen species (ROS), phase II metabolism (activation of, for example, glutathione S-transferase, glucuronosyltransferase, or sulfotransferase), and maintenance of genome integrity (increase in DNA replication and repair fidelity, by activation of error-free DNA repair and/or inhibition of error-prone pathways) [11], [12], [13], [15]. Suppressing agents can act on cell division to restore a lost equilibrium between proliferation and apoptosis, on angiogenesis, or on events leading to metastasis [15], [16]. According to Tsuda et al., the ultimate goal of cancer chemoprevention would be to live without cancer or to live with cancer without feeling the symptoms until the natural end of life [14].

Numerous studies established a link between the consumption of fruits and vegetables and a significant reduction in cancer risk [12], [17], [18], [19]. A wide variety of phytochemicals, particularly polyphenols, whose primary functions are to protect plants from photosynthetic stress, ROS, microorganisms, and herbivore feeding, have demonstrated chemopreventive effects against some cancers through several mechanisms including anti-inflammatory and antioxidant [20] and specific beneficial effects on the regulation of key proteins implicated in cell cycle control, cell differentiation, inflammation, DNA methylation, apoptosis, angiogenesis, tumor growth, and metastasis (see reviews [16], [21] and Supporting Information Table S1). The phytochemicals tested in the present study are mainly polyphenols and have been selected after a literature survey of their purported cancer chemoprevention properties (Supporting Information Table S1) and for their previously demonstrated effects on DNA polymerases [22], [23]. Although effects related to DNA damage repair are probably not the main chemoprevention-related activities of polyphenols, we want to explore whether these compounds might have a role in genome maintenance integrity through modulation of DNA damage tolerance and repair. To do so, we focused on DNA TLS, a major mechanism of DNA damage tolerance that allows the cells to avoid replication blockade in the presence of a lesion on DNA. We selected 8-oxodG, an oxidative lesion whose presence on a DNA strand may result either from the insertion of an oxidized guanine during DNA synthesis or from the oxidation of a guanine on the DNA strand. This lesion is highly mutagenic due to its ability to undergo an anti-syn conformational change that functionally mimics T during replication [24]. This feature makes 8-oxodG more prone to a mismatch with adenine, which can lead to a C: G to A: T transversion, a mutation common to many cancers [25], [26]. The 8-oxodG lesion itself is very prone to oxidation into products with a higher mutagenic potential, implying an even higher rate of C: G to A: T transversions [27], [28], [29].

In eukaryotes, the occurrence of such a lesion can be avoided/corrected by 3 known mechanisms: (i) hydrolysis of 8-oxodGTP, catalyzed by the NUDIX hydrolase MutT homolog 1 [30], which prevents its incorporation into DNA; (ii) excision base repair (BER) via the 8-oxoguanine DNA glycosylase 1 (OGG1) [31]; and (iii) error correction by MutY homolog [32], a glycosylase specialized in the removal of mismatched adenine in front of an 8-oxodG.

In vitro, the classical replicative DNA polymerases (DNA pol α, δ, and ε [33]) are mainly “error-prone” toward an 8-oxodG because of their conformational properties. Also, in front of the lesion, the specialized DNA pol η, pol λ [34], and pol ζ [34], [35] mainly incorporate a cytosine, while DNA pol κ preferentially inserts an adenine [36]. DNA pol η has been shown to prevent mismatching of 8-oxodG during plasmid replication and to affect mutation frequency [37]. Rodriguez et al. suggested that 8-oxodG blocks the replication fork progression in vivo but that DNA pol η can replicate this lesion with accuracy [38].

The emergence of the concept of eukaryotic TLS as a major mechanism for DNA damage tolerance has led to the development of several assays to monitor its speed and/or fidelity, to characterize the activities of specialized DNA polymerases, and to determine their role regarding specific lesions. However, most of these assays show limitations in resolution, reproducibility, or quantification; in addition, they are generally not adapted to screening because they are too time- and labor-consuming [34], [39], [40]. To overcome these limitations, our laboratories have developed and analytically validated bioassays for DNA polymerase kinetics and fidelity, using capillary electrophoresis [41] and pyrosequencing [42]. In the present study, these methods were applied to screen the influence of various phytochemicals on the kinetics and fidelity of nuclear protein extracts at performing TLS past 8-oxo-dG.

Results

In a previous work [43], the cytotoxicity of all the polyphenols to be studied here was evaluated on FHs74Int intestinal epithelial cells in culture over 24 h. Cell survival curves (percentage of cell survival as a function of the compound concentration) allowed to determine the IC10. If the IC10 was above 30 µM, this latter concentration was selected as the working dose in the present study. Typical plasma levels of these compounds being generally in the µM range, a concentration of 30 µM was considered as a threshold value under which the occurrence of nonspecific effects is negligible. A preliminary step of compound solubilization in DMSO was carried out to achieve a maximum concentration of 0.5% of DMSO after dilution in the cell culture medium. The same DMSO amount was added to the medium of the “control” cell flasks.

To measure the correct or mutagenic outcome of 8-oxodG bypass by TLS DNA polymerases, we performed in vitro replication of a lesion-containing template DNA by extension of a 39-mer primer annealing just 5′ of the 8-oxodG lesion; DNA polymerases were obtained as total nuclear proteins extracted from normal human intestinal epithelial cells as previously described [41], [42]. The reaction products (both the initial 39-mer primer and the primer extended by 1 or 2 nucleotide[s] in front of the lesion) were analyzed by both capillary electrophoresis ([Fig. 1]) (kinetics measurement by quantification of the 40- and 41-mer extension products) [41] and pyrosequencing ([Fig. 2]) (identification and quantification of nucleotides inserted in front of the lesion) [42].

In a first set of experiments, the “error-prone” TLS was evaluated by capillary electrophoresis in the presence of dATP and individual polyphenols added to the primer extension reaction mix. The proportions of the 40-mer extension product relatively to the 39-mer initial primer and normalized to control experiment are shown in [Fig. 3].

When added to the reaction mixture during this primer extension assay, apigenin, baicalein, sakuranetin, myricetin, and arbutin reduced the incorrect dAMP insertion in front of the 8-oxodG lesion. By contrast, under the same conditions, epicatechin stimulated dAMP insertion.

In a second set of experiments, the “error-free” TLS was evaluated by the addition of dCTP to the primer extension reaction mix, which led to primer elongation by 1 or 2 nucleotides ([Fig. 1]). The proportions of primer extension products (40- and 41-mer) to the 39-mer primer and normalized to control experiment are shown in [Fig. 4].

When added to the reaction mixture during this primer extension assay, apigenin, sakuranetin, myricetin, chalcone, genistein, indole-3-carbinol and arbutin reduced the correct dCMP insertion in front of the 8-oxodG lesion. However, both baicalein and naringenin promoted dCMP insertion in the same conditions.

To visualize the overall effects of these phytochemicals on the outcome of 8-oxodG TLS by the nuclear protein extract, the ratios of the percentages of different primer extension products, obtained in the presence of dATP or dCTP, are shown in [Fig. 5].

Apigenin, baicalein, and myricetin significantly promoted the nonmutagenic dCMP incorporation in front of 8-oxodG when added to the primer extension reaction mix. By contrast, chalcone and epicatechin both significantly promoted the mutagenic outcome (i.e., dAMP incorporation) under the same conditions.

We then studied the effect of each polyphenol added to the cell culture for 24 h before the extraction of nuclear proteins to be used in the primer extension assay for 8-oxodG TLS. In a first set of experiments, the “error-prone” TLS was evaluated. The proportions of primer extension products in the presence of dATP (proportions of elongated 40-mer peak area to the 39-mer peak area), normalized to their respective control, are shown in [Fig. 6].

Nuclear protein extracts of both apigenin- and curcumin-treated FHs74Int cells (10 µM and 5 µM, respectively; 24 h) significantly reduced the mutagenic insertion of dAMP in front of the 8-oxodG lesion. Under the same conditions, epicatechin stimulated the insertion of dAMP.

In a second set of experiments, “error-free” TLS was evaluated by primer extension in the presence of dCTP. The proportions of elongated 40- and 41-mer to 39-mer primer normalized to their respective control are shown in [Fig. 7].

Nuclear protein extracts of both apigenin- and curcumin-treated FHs74Int cells (10 µM and 5 µM, respectively; 24 h) significantly reduced the correct insertion of dCMP in front of the 8-oxodG lesion during TLS. Naringenin and sakuranetin both stimulated the insertion of dCMP opposite 8-oxodG under the same conditions.

To visualize the phytochemical effects on the outcome of TLS in front of 8-oxodG, the ratios of the proportions of the different primer extension products obtained in the presence of dATP and dCTP were calculated ([Fig. 8]).

Nuclear protein extracts of both sakuranetin- and myricetin-treated FHs74Int cells significantly promoted the nonmutagenic outcome of TLS in front of 8-oxodG. However, epicatechin significantly promoted the mutagenic outcome of TLS of 8-oxodG under the same conditions.

In a third set of experiments, the magnolol and honokiol neolignans were tested under the same conditions; the results are shown in [Fig. 9].

When added to the cell culture medium for 24 h, both compounds stimulated the mutagenic insertion of dAMP in front of the 8-oxodG lesion but did not affect the correct insertion of dCMP. In conclusion, magnolol and honokiol significantly promoted the mutagenic outcome of this translation synthesis.

The above experiments indicated that the modulatory effects on the TLS in front of 8-oxodG were higher when the phytochemical was in contact with the cells for 24 h before the preparation of nuclear protein extracts. This configuration was therefore maintained for the subsequent experiments, in which, as previously described [42], 8-oxodG bearing primers were tested in vitro for elongation by “treated” nuclear extracts and corresponding “control”. Pyrosequencing of the primer extension products obtained in the presence of nuclear proteins and all 4 dNTPs allowed us to determine the proportions of dAMP and dCMP incorporation in front of the 8-oxodG lesion.

The results obtained for each phytochemical are shown in [Fig. 10]. Only apigenin and epicatechin appeared to significantly modulate the proportions of nucleotides inserted in front of 8-oxodG under these conditions. These compounds increased the mutagenic potential of the lesion by favoring dAMP versus dCMP incorporation. The neolignans magnolol and honokiol did not present a significant effect in this test configuration.

Discussion

Comparison of the 2 analytical methods

The development and analytical validation of the 2 methods applied in this paper are discussed in the previously published development and validation studies [41], [42].

A point-by-point comparison of both analytical methods ([Table 1]) indicates that they are complementary. The capillary electrophoresis method allows the quantification of the nucleotide incorporated in front of the lesion by measuring the rate of synthesis, which corresponds to the speed at which a polymerase inserts a nucleotide in front of the lesion. This is a measurement of the insertion kinetics. The pyrosequencing method does not measure the rate of synthesis but makes it possible to quantitatively detect the correct versus incorrect insertion ratio in the presence of the 4 dNTPs, which is a condition more similar to what happens in vivo. The 2 configurations used in these tests allowed us to study 2 types of effects: (i) when the phytochemical compound is directly added to the primer extension reaction medium, the observed effect is a direct action (activation or inhibition) of the compound on 1 or more enzymes or auxiliary proteins or cofactors involved in the TLS mechanism; and (ii) when the compound is added to the cells 24 h before nuclear protein extraction, the observed effect is a “cellular” effect resulting from a response of the cell to the compound, possibly involving alterations in the expression of enzymes, auxiliary proteins or cofactors involved in the TLS mechanism.

Modulation of 8-oxodG TLS by phytochemicals

With the notable exception of quercetin, all tested phytochemicals demonstrated an effect in at least 1 of the performed tests.

Apigenin showed an inhibitory effect on the kinetics of TLS, reducing both dAMP and dCMP insertion in front of the 8-oxodG lesion and favored the mutagenic outcome of the TLS. Two previous studies reported that apigenin administration to mice greatly decreased the expression of the PCNA, a processivity factor required for DNA replication and TLS [44], [45]. This observation is consistent with the inhibition of primer extension measured in our different experiments.

Our data indicate that baicalein has an effect only when directly added to the reaction mixture during in vitro primer extension assay. This flavone has already shown a modest inhibitory effect on DNA pol γ, a polymerase involved in the replication of mitochondrial DNA [46]. Previous in vitro data indicate that baicalein treatment of HEK293T cells, derived from embryonic renal tissue, causes increased PCNA ubiquitination and DNA pol η deubiquitination; this DNA pol η ubiquitination is necessary for its activation and involvement in 8-oxodG TLS [47].

As indicated by capillary electrophoresis, the treatment of cells by naringenin stimulates the correct insertion of dCTP in front of the lesion. However, a few references in the literature indicate downregulation of PCNA expression after oral administration of naringenin nanoparticles [48], which seems not consistent with the observed effects herein.

Although treatment of cells with sakuranetin has demonstrated antimutagenic modulation in our assays in vitro, no literature reference was found so far that could explain this type of effect. Myricetin had an inhibitory effect on the incorporation of nucleotides in front of the 8-oxodG lesion in our in vitro primer extension assays analyzed by capillary electrophoresis. This is consistent with effects described for myricetin, which was found to be a competitive inhibitor of some DNA polymerases (pol α, Ki = 1.6 µM; pol I from E. coli, Ki = 0.37 µM) and HIV reverse transcriptase (Ki = 0.08 µM) [46]. In a more recent study, Shiomi et al. have shown that myricetin could inhibit the activities of DNA pol α, β, and κ by more than 80% [22]; the structural elements responsible for these activities have been ascribed to the hydroxyl groups on the aromatic ring B of the flavonoids [22]. This assertion, however, is inconsistent with our data as 2 compounds hydroxylated on ring B (quercetin and epicatechin) did not show comparable activities. Apart from this reduction in TLS kinetics, our work made it possible to characterize a nonmutagenic effect of myricetin on the outcome of 8-oxodG translation synthesis.

In all our experiments, epicatechin demonstrated stimulatory effects on the insertion of dAMP in front of the 8-oxodG lesion, promoting the mutagenic outcome of this TLS. In the literature, epicatechin has in vitro inhibitory effects on DNA polymerase λ that are known to be involved in the “error-free” replication of 8-oxodG [49], which is in agreement with our data.

Chalcone and genistein both had a single effect, namely an inhibition of dCMP insertion in front of the lesion and an ability to promote the mutagenic outcome of 8-oxodG TLS when these compounds are added to the reaction mixture. Literature data indicate that treatment of HEK293T cells with genistein results in a decrease in the amount of unmodified and ubiquitinated PCNA bound to chromatin [47], however, such an effect will most probably not be induced by the simple addition of the flavonoid to the reaction mixture.

Capillary electrophoresis experiment indicated that curcumin, indole-3-carbinol, and arbutin had mainly inhibitory effects on the kinetics of nucleotides insertion in front of the 8-oxodG lesion. Curcumin is also described as a fairly specific inhibitor of DNA polymerase λ, involved in the “error-free” replication of 8-oxodG [49] with no effect on the activity of DNA polymerases α, γ, δ, ε, and β [50], [51].

Honokiol and magnolol both stimulated the incorrect dAMP incorporation in front of the 8-oxodG lesion and thus promoted the mutagenic outcome of this TLS. Magnolol induced inhibition of DNA synthesis and PCNA expression in vitro in vascular smooth muscle cells [52]. Since the decrease in PCNA expression promotes the mutagenic outcome of the 8-oxodG TLS [34], our results are in agreement with those previous studies.

Modulation of 8-oxodG TLS: possible mechanisms and consequences in cancer chemoprevention and therapy

In TLS, stimulation or inhibition of a specific nucleotide insertion by a xenobiotic could be attributed to (i) a selective capacity of inhibition toward certain specialized DNA polymerases (depending on whether the xenobiotic can inhibit a DNA polymerase exhibiting an “error-prone” or an “error-free” behavior toward the considered lesion, the relay by another DNA polymerase, possibly more or less faithful, can take place, favoring a non-mutagenic or mutagenic outcome of the TLS, respectively); and/or (ii) modulation of the complex mechanisms of interplay between replicative and specialized polymerases at the level of the replication fork [53].

Based on the kinetic effects induced by the addition of tested phytochemicals to the reaction medium, we can classify them into 2 groups; (i) compounds that inhibit the insertion of dATP in front of 8-oxodG: myricetin > apigenin > arbutin > baicalein > sakuranetin and (ii) compounds inhibiting the insertion of dCTP in front of 8-oxodG: arbutin > chalcone > myricetin > sakuranetin > apigenin > genistein > indole-3-carbinol. Regarding the kinetic effects induced by cell incubation with the tested phytochemicals, we can differentiate: (i) compounds inhibiting the insertion of dATP in front of 8-oxodG: apigenin > curcumin and (ii) compounds inhibiting the insertion of dCTP in front of 8-oxodG: apigenin ~ curcumin. Regarding the mutagenic effects induced by cell incubation with the tested phytochemicals, we can distinguish compounds promoting the incorrect insertion of dATP opposite 8-oxodG: apigenin > epicatechin.

The inhibitory activity of myricetin could be explained by studies of molecular docking on DNA polymerase that showed binding to the polymerization catalytic site, competing with the incoming nucleotide, and blocking the DNA elongation process; a similar effect was shown for the stilbene resveratrol [23].

As suggested by Shiomi et al. [22], the number of hydroxyl groups on the tested polyphenols appears related to their inhibitory activity on dATP or dCTP insertion in front of 8-oxodG; however, establishing a structure-activity relationship is certainly premature at this stage and would require the use of pure DNA polymerases.

Furthermore, it appears that, besides their inhibitory activities on DNA polymerases, some of the studied polyphenols modulate the outcome of TLS, suggesting different affinities for the catalytic site depending on the involved polymerases. Since DNA polymerases involved in TLS present structural differences from replicative polymerases, especially at their catalytic site [54], it is reasonable to hypothesize that the polyphenols could exert different inhibitory activities against individual DNA polymerases.

Such modulations of TLS could be of high importance in the course of exposure to mutagens. Pushing an individual cell towards a mutagenic or nonmutagenic TLS could initiate or prevent a given mutational event. From this perspective, reduced to the cell level, modest modulating activity is most probably susceptible to effects. In addition, such TLS effects add to the many biological processes also modulated by these compounds and provide additional clues to the mechanisms involved in their cancer chemopreventive activities. Nevertheless, although chemoprevention by phytochemicals appears as an acceptable, nonexpensive, and rapidly applicable modality for cancer limitation, the in vitro concentrations tested in the literature are often supra-physiological and therefore likely not to be reached in vivo [11]. Moreover, polyphenolic compounds are frequently consumed under their various glycoside forms and converted in vivo into other forms, which diminishes their bioavailability as is [55]. The chemopreventive activities of polyphenols indeed depend on a series of parameters that include the dose, the route and frequency of administration, eventual chemical interactions, pharmacokinetic parameters, and individual intestinal microbiotas. The real impact of these substances on health remains to be evaluated through epidemiological data and pharmacokinetic studies coupled with clinical trials.

In therapeutic strategies against cancer, the selective inhibition of DNA polymerases or the polymerase interplays by phytochemicals could have a dual effect, sensitizing the tumor to therapy and preventing the emergence of resistance to treatment [56]. This corresponds to the principle of “synthetic lethality” by which defects in DNA repair pathways can be tolerated alone in a cancer cell but become lethal when combined [57]. Some inhibitors of specialized DNA polymerases have already been studied for this attractive potential. Eicosapentaenoic acid (C20: 5ω3), a DNA pol β, pol δ, and pol ε inhibitor, sensitizes cancer cells to radiotherapy in vitro [58]. The inhibition of error-prone DNA polymerase expression by interfering RNAs sensitizes cells to anticancer agents and prevents the emergence of resistances; this has been shown for Rev3, the catalytic subunit of the TLS DNA polymerase zeta, and cisplatin, both in vitro and in vivo, [59] and for Rev1, a DNA repair protein involved in the polymerase switching-event during TLS, and cisplatin or cyclophosphamide [56]. This indicates that inhibiting the expression and/or activity of DNA polymerases may increase anticancer therapeutic effects, reduce the development of resistance to DNA-damaging radio-/chemotherapy, and thus improve the clinical outcome of treatments. However, highly selective inhibitors that could be used as an adjuvant in cancer therapies remain to be discovered. The polyphenols investigated here could give a clue as to the structure of such agents.

Material and Methods

Chemicals

Oligonucleotides and primers were purchased from Sigma-Aldrich. Pyrosequencing and capillary electrophoresis reagents were obtained from Qiagen and Analis, respectively. Apigenin (purity ≥ 97%), arbutin (purity ≥ 98%), baicalein (purity ≥ 98%), curcumin (purity ≥ 65%), epicatechin (purity ≥ 90%), genistein (purity ≥ 97%), indole-3-carbinol, myricetin (purity ≥ 96.0%), naringenin (purity ≥ 95%), quercetin (purity ≥ 98%), sakuranetin (purity ≥ 95%), and trans-chalcone (purity ≥ 97%) were purchased from Sigma-Aldrich. Honokiol (purity ≥ 99.6%) and magnolol (purity ≥ 99.5%) were from EDQM, Strasbourg, France.

Cell culture

The FHs74Int human intestinal epithelial cell line (ATCC # CCL-241) was obtained from the American Type Culture Collection. Cells were cultured in DMEM high-glucose medium supplemented with 10% FBS Gold, 10 mM HEPES, 2 mM L-glutamine, 1% non-essential amino acid, 100 000 U/L penicillin, 100 mg/L streptomycin (obtained from Lonza), 30 ng/µL EGF, and 10 µg/µL insulin (obtained from Sigma) and maintained at 37 °C in a humidified atmosphere of 5% CO₂ in the air. Nuclear protein extraction and dosage were performed as previously described [41], [42].

Primer extension assay

The capillary electrophoresis ([Fig. 1]) and pyrosequencing ([Fig. 2]) techniques applied for the primer extension assays have been fully described and analytically validated [41], [42], indicating their suitability for the determination of 8-oxodG TLS kinetics and fidelity.

Briefly, the reaction is carried out in TLS buffer pH 7.0 (50 mM Tris-Base, 0.25 µg/µL BSA, 1 mM DTT, 5 mM MgCl₂) containing 20 nM of hybridized oligonucleotides, 100 µM of dATP, dCTP, or dNTP mixture, and 0.4 µg/µL of nuclear protein extract (final reaction volume, 20 µL). The reaction mixture is then incubated at 25 °C for 10 min, and the reaction is stopped by adding 1 µL of 0.5 M EDTA. The mixture is then diluted in TE buffer pH 7.9 (10 mM Tris-Base, 1 mM EDTA) to obtain a final DNA concentration of 5 nM and stored at − 20 °C protected from light.

Influence of phytochemicals on 8-oxodG TLS bypass

Two configurations of phytochemical addition were selected, by adding the polyphenol either (i) to the reaction mixture during the primer extension assay (to study an eventual direct influence of tested compounds on translesion elongation); or (ii) to the culture medium, 24 h before cell harvest and nuclear proteins extraction (to study an eventual cellular effect leading to modulation of translesion elongation). Control experiments were performed without compound and with 0.5% DMSO (w/v) concentration in the reaction mix.

Statistical analysis

All the experiments were conducted in triplicate. Statistical analyses were performed with GraphPad Prism 8 software; 2-way or 1-way ANOVA tests were used to compare experimental data; p-values < 0.05 were considered significant.

Contributorsʼ Statement

Data collection: A. Nachtergael, M. Spanoghe, D. Lanterbecq; design of the study: A. Nachtergael, P. Duez, A. Belayew; statistical analysis: A. Nachtergael, P. Duez; analysis and interpretation of the data: A. Nachtergael, P. Duez, A. Belayew; drafting the manuscript: A. Nachtergael; revision of the manuscript: M. Spanoghe, D. Lanterbecq, P. Duez, A. Belayew.

Conflict of Interest

The authors declare that they have no conflict of interest.

Acknowledgements

This research was supported by the Foundation “Plants for Health” (Award 2020) and the University of Mons (UMONS). The help of Hakan Baykus and Pascal Vannuffel for the pyrosequencing experiments is kindly acknowledged.

^# Dedicated to Professor Arnold Vlietinck on the occasion of his 80th birthday.

• References

• 1 Vineis P, Schatzkin A, Potter JD. Models of carcinogenesis: an overview. Carcinogenesis 2010; 31: 1703-1709
  CrossrefPubMedSearch in Google Scholar
• 2 Poivre M, Nachtergael A, Bunel V, Philippe ON, Duez P. Genotoxicity and Carcinogenicity of herbal Products. In: Pelkonen O, Duez P, Vuorela PM, Vuorela H. eds. Toxicology of herbal Products. Cham: Springer International Publishing; 2017: 179-215
  CrossrefSearch in Google Scholar
• 3 Balmain A. Cancer genetics: from Boveri and Mendel to microarrays. Nat Rev Cancer 2001; 1: 77-82
  CrossrefPubMedSearch in Google Scholar
• 4 Loeb LA, Springgate CF, Battula N. Errors in DNA replication as a basis of malignant changes. Cancer Res 1974; 34: 2311-2321
  PubMedSearch in Google Scholar
• 5 Migheli F, Migliore L. Epigenetic Perturbations in the Context of the multi-hit Hypothesis of Carcinogenesis. In: Maulik N, Karagiannis T. eds. Molecular Mechanisms and Physiology of Disease. New York: Springer; 2014: 383-399
  CrossrefSearch in Google Scholar
• 6 Link A, Balaguer F, Goel A. Cancer chemoprevention by dietary polyphenols: Promising role for epigenetics. Biochem Pharmacol 2010; 80: 1771-1792
  CrossrefPubMedSearch in Google Scholar
• 7 Schüz J, Espina C, Villain P, Herrero R, Leon ME, Minozzi S, Romieu I, Segnan N, Wardle J, Wiseman M, Belardelli F, Bettcher D, Cavalli F, Galea G, Lenoir G, Martin-Moreno JM, Nicula FA, Olsen JH, Patnick J, Primic-Zakelj M, Puska P, van Leeuwen FE, Wiestler O, Zatonski W, Guha N, Kralikova E, McNeill A, Peruga A, Anderson A, Berrino F, Boutron-Ruault MC, Cecchini M, Key TJ, Leitzmann M, Norat T, Powers HJ, Scoccianti C, Auvinen A, de Vries E, Erdmann F, Greinert R, Harrison J, Kesminiene A, McColl N, Friis S, Kogevinas M, Saracci R, Straif K, Vainio H, Almonte M, Anttila A, De Vuyst H, Dillner J, Franceschi S, Gonzalez P, Hall A, Park JY, Armaroli P, Atkin W, Dean PB, de Koning H, Dillner L, Kuipers E, Lansdorp-Vogelaar I, Paci E, Regula J, Suonio E, Törnberg S, Wood LF, Gaudin N, Frie KG, Terrasse V, Winstanley K, Bellisario C, Biagioli E, Cinquini M, Gianola S, Gonzalez Lorenzo M, von Karsa L, Lignini T. European code against cancer 4th edition: 12 ways to reduce your cancer risk. Cancer Epidemiol 2015; 39: S1-S10
  CrossrefPubMedSearch in Google Scholar
• 8 Wild C, Weiderpass E, Stewart BW. World Cancer Report: Cancer Research for Cancer Prevention. In: Wild C, Weiderpass E, Stewart BW. eds. World Cancer Report. Lyon, France: International Agency for Research on Cancer; 2020: 15-45
  Search in Google Scholar
• 9 Sporn MB, Dunlop N, Newton D, Smith J. Prevention of chemical carcinogenesis by vitamin A and its synthetic analogs (retinoids). Fed Proc 1976; 35: 1332-1338
  PubMedSearch in Google Scholar
• 10 Theisen C. Chemoprevention: whatʼs in a name?. J Natl Cancer Inst 2001; 93: 743
  CrossrefPubMedSearch in Google Scholar
• 11 Surh YJ. Cancer chemoprevention with dietary phytochemicals. Nat Rev Cancer 2003; 3: 768-780
  CrossrefPubMedSearch in Google Scholar
• 12 Shukla Y, Pal SK. Dietary cancer chemoprevention: an overview. Int J Hum Genet 2004; 4: 265-276
  CrossrefPubMedSearch in Google Scholar
• 13 De Flora S, Ferguson LR. Overview of mechanisms of cancer chemopreventive agents. Mutat Res Fundam Mol Mech Mutagen 2005; 591: 8-15
  CrossrefPubMedSearch in Google Scholar
• 14 Tsuda H, Ohshima Y, Nomoto H, Fujita K, Matsuda E, Iigo M, Takasuka N, Moore MA. Cancer prevention by natural compounds. Drug Metab Pharmacokinet 2004; 19: 245-263
  CrossrefPubMedSearch in Google Scholar
• 15 Ramos S. Cancer chemoprevention and chemotherapy: dietary polyphenols and signalling pathways. Mol Nutr Food Res 2008; 52: 507-526
  CrossrefPubMedSearch in Google Scholar
• 16 Aggarwal BB, Shishodia S. Molecular targets of dietary agents for prevention and therapy of cancer. Biochem Pharmacol 2006; 71: 1397-1421
  CrossrefPubMedSearch in Google Scholar
• 17 Riboli E, Norat T. Epidemiologic evidence of the protective effect of fruit and vegetables on cancer risk. Am J Clin Nutr 2003; 78: 559S-569S
  CrossrefPubMedSearch in Google Scholar
• 18 La Vecchia C. Mediterranean diet and cancer. Public Health Nutr 2004; 7: 965-968
  CrossrefPubMedSearch in Google Scholar
• 19 Heath AK, Muller DC, van den Brandt PA, Papadimitriou N, Critselis E, Gunter M, Vineis P, Weiderpass E, Fagherazzi G, Boeing H, Ferrari P, Olsen A, Tjønneland A, Arveux P, Boutron-Ruault MC, Mancini FR, Kühn T, Turzanski-Fortner R, Schulze MB, Karakatsani A, Thriskos P, Trichopoulou A, Masala G, Contiero P, Ricceri F, Panico S, Bueno-de-Mesquita B, Bakker MF, van Gils CH, Olsen KS, Skeie G, Lasheras C, Agudo A, Rodríguez-Barranco M, Sánchez MJ, Amiano P, Chirlaque MD, Barricarte A, Drake I, Ericson U, Johansson I, Winkvist A, Key T, Freisling H, His M, Huybrechts I, Christakoudi S, Ellingjord-Dale M, Riboli E, Tsilidis KK, Tzoulaki I. Nutrient-wide association study of 92 foods and nutrients and breast cancer risk. Breast Cancer Res 2020; 22: 5
  CrossrefPubMedSearch in Google Scholar
• 20 Nichols J, Katiyar S. Skin photoprotection by natural polyphenols: anti-inflammatory, antioxidant and DNA repair mechanisms. Arch Dermatol Res 2010; 302: 71-83
  CrossrefPubMedSearch in Google Scholar
• 21 Gullett NP, Ruhul Amin ARM, Bayraktar S, Pezzuto JM, Shin DM, Khuri FR, Aggarwal BB, Surh YJ, Kucuk O. Cancer prevention with natural compounds. Semin Oncol 2010; 37: 258-281
  CrossrefPubMedSearch in Google Scholar
• 22 Shiomi K, Kuriyama I, Yoshida H, Mizushina Y. Inhibitory effects of myricetin on mammalian DNA polymerase, topoisomerase and human cancer cell proliferation. Food Chem 2013; 139: 910-918
  CrossrefPubMedSearch in Google Scholar
• 23 Wu Y, Hsieh TC, Wu JM, Wang X, Christopher JS, Pham AH, Swaby JD, Lou L, Xie ZR. Elucidating the inhibitory effect of resveratrol and its structural analogs on selected nucleotide-related enzymes. Biomolecules 2020; 10: 1223
  CrossrefPubMedSearch in Google Scholar
• 24 McAuley-Hecht KE, Leonard GA, Gibson NJ, Thomson JB, Watson WP, Hunter WN, Brown T. Crystal structure of a DNA duplex containing 8-hydroxydeoxyguanine-adenine base pairs. Biochemistry 1994; 33: 10266-10270
  CrossrefPubMedSearch in Google Scholar
• 25 Arana ME, Kunkel TA. Mutator phenotypes due to DNA replication infidelity. Semin Cancer Biol 2010; 20: 304-311
  CrossrefPubMedSearch in Google Scholar
• 26 Wang D, Kreutzer DA, Essigmann JM. Mutagenicity and repair of oxidative DNA damage: insights from studies using defined lesions. Mutat Res Fundam Mol Mech Mutagen 1998; 400: 99-115
  CrossrefPubMedSearch in Google Scholar
• 27 Henderson PT, Delaney JC, Gu F, Tannenbaum SR, Essigmann JM. Oxidation of 7,8-dihydro-8-oxoguanine affords lesions that are potent sources of replication errors in vivo . Biochemistry 2001; 41: 914-921
  CrossrefPubMedSearch in Google Scholar
• 28 Henderson PT, Delaney JC, Muller JG, Neeley WL, Tannenbaum SR, Burrows CJ, Essigmann JM. The hydantoin lesions formed from oxidation of 7,8-dihydro-8-oxoguanine are potent sources of replication errors in vivo . Biochemistry 2003; 42: 9257-9262
  CrossrefPubMedSearch in Google Scholar
• 29 Henderson PT, Neeley WL, Delaney JC, Gu F, Niles JC, Hah SS, Tannenbaum SR, Essigmann JM. Urea lesion formation in DNA as a consequence of 7, 8-dihydro-8-oxoguanine oxidation and hydrolysis provides a potent source of point mutations. Chem Res Toxicol 2004; 18: 12-18
  CrossrefPubMedSearch in Google Scholar
• 30 Maki H, Sekiguchi M. MutT protein specifically hydrolyses a potent mutagenic substrate for DNA synthesis. Nature 1992; 355: 273-275
  CrossrefPubMedSearch in Google Scholar
• 31 Radicella JP, Dherin C, Desmaze C, Fox MS, Boiteux S. Cloning and characterization of hOGG1, a human homolog of the OGG1 gene of Saccharomyces cerevisiae. Proc Natl Acad Sci U S A 1997; 94: 8010-8015
  CrossrefPubMedSearch in Google Scholar
• 32 McCann JAB, Berti PJ. Adenine release is fast in MutY-catalyzed hydrolysis of G : A and 8-oxo-G : A DNA mismatches. J Biol Chem 2003; 278: 29587-29592
  CrossrefPubMedSearch in Google Scholar
• 33 Hübscher U, Maga G. DNA replication and repair bypass machines. Curr Opin Chem Biol 2011; 15: 627-635
  CrossrefPubMedSearch in Google Scholar
• 34 Maga G, Villani G, Crespan E, Wimmer U, Ferrari E, Bertocci B, Hubscher U. 8-oxo-guanine bypass by human DNA polymerases in the presence of auxiliary proteins. Nature 2007; 447: 606-608
  CrossrefPubMedSearch in Google Scholar
• 35 Kamiya H, Yamaguchi A, Suzuki T, Harashima H. Roles of specialized DNA polymerases in mutagenesis by 8-hydroxyguanine in human cells. Mutat Res Fundam Mol Mech Mutagen 2010; 686: 90-95
  CrossrefPubMedSearch in Google Scholar
• 36 Irimia A, Eoff RL, Guengerich FP, Egli M. Structural and functional elucidation of the mechanism promoting error-prone synthesis by human DNA polymerase kappa opposite the 7,8-dihydro-8-oxo-2′-deoxyguanosine adduct. J Biol Chem 2009; 284: 22467-22480
  CrossrefPubMedSearch in Google Scholar
• 37 Lee DH, Pfeifer GP. Translesion synthesis of 7,8-dihydro-8-oxo-2′-deoxyguanosine by DNA polymerase eta in vivo . Mutat Res Fundam Mol Mech Mutagen 2008; 641: 19-26
  CrossrefPubMedSearch in Google Scholar
• 38 Rodriguez GP, Song JB, Crouse GF. In vivo bypass of 8-oxodG. PLoS Genet 2013; 9: e1003682
  CrossrefPubMedSearch in Google Scholar
• 39 Tietz D. Nucleic Acid Electrophoresis. Heidelberg: Springer; 1998
  CrossrefSearch in Google Scholar
• 40 Ding L, Williams K, Ausserer W, Bousse L, Dubrow R. Analysis of plasmid samples on a microchip. Anal Biochem 2003; 316: 92-102
  CrossrefPubMedSearch in Google Scholar
• 41 Nachtergael A, Charles C, Spanoghe M, Gadenne M, Belayew A, Duez P. Measurement of translesion synthesis by fluorescent capillary electrophoresis: 7,8-Dihydro-8-oxodeoxyguanosine bypass modulation by natural products. Anal Biochem 2013; 440: 23-31
  CrossrefPubMedSearch in Google Scholar
• 42 Nachtergael A, Belayew A, Duez P. Pyrosequencing for the quantitative assessment of 8-oxodG bypass DNA synthesis. DNA Repair (Amst) 2014; 22: 147-152
  CrossrefPubMedSearch in Google Scholar
• 43 Charles C, Nachtergael A, Ouedraogo M, Belayew A, Duez P. Effects of chemopreventive natural products on non-homologous end-joining DNA double-strand break repair. Mutat Res Genet Toxicol Environ Mutagen 2014; 768: 33-41
  CrossrefPubMedSearch in Google Scholar
• 44 Shukla S, MacLennan GT, Flask CA, Fu P, Mishra A, Resnick MI, Gupta S. Blockade of β-catenin signaling by plant flavonoid apigenin suppresses prostate carcinogenesis in TRAMP mice. Cancer Res 2007; 67: 6925-6935
  CrossrefPubMedSearch in Google Scholar
• 45 Liu LZ, Fang J, Zhou Q, Hu X, Shi X, Jiang BH. Apigenin inhibits expression of vascular endothelial growth factor and angiogenesis in human lung cancer cells: implication of chemoprevention of lung cancer. Mol Pharmacol 2005; 68: 635-643
  CrossrefPubMedSearch in Google Scholar
• 46 Ono K, Nakane H. Mechanisms of inhibition of various cellular DNA and RNA polymerases by several flavonoids. J Biochem 1990; 108: 609-613
  CrossrefPubMedSearch in Google Scholar
• 47 Fox JT, Sakamuru S, Huang R, Teneva N, Simmons SO, Xia M, Tice RR, Austin CP, Myung K. High-throughput genotoxicity assay identifies antioxidants as inducers of DNA damage response and cell death. Proc Natl Acad Sci U S A 2012; 109: 5423-5428
  CrossrefPubMedSearch in Google Scholar
• 48 Sulfikkarali N, Krishnakumar N, Manoharan S, Nirmal R. Chemopreventive efficacy of naringenin-loaded nanoparticles in 7,12-dimethylbenz(a)anthracene induced experimental oral carcinogenesis. Pathol Oncol Res 2013; 19: 287-296
  CrossrefPubMedSearch in Google Scholar
• 49 Matsubara K, Saito A, Tanaka A, Nakajima N, Akagi R, Mori M, Mizushina Y. Epicatechin conjugated with fatty acid is a potent inhibitor of DNA polymerase and angiogenesis. Life Sci 2007; 80: 1578-1585
  CrossrefPubMedSearch in Google Scholar
• 50 Mizushina Y, Hirota M, Murakami C, Ishidoh T, Kamisuki S, Shimazaki N, Takemura M, Perpelescu M, Suzuki M, Yoshida H, Sugawara F, Koiwai O, Sakaguchi K. Some anti-chronic inflammatory compounds are DNA polymerase λ-specific inhibitors. Biochem Pharmacol 2003; 66: 1935-1944
  CrossrefPubMedSearch in Google Scholar
• 51 Takeuchi T, Ishidoh T, Iijima H, Kuriyama I, Shimazaki N, Koiwai O, Kuramochi K, Kobayashi S, Sugawara F, Sakaguchi K, Yoshida H, Mizushina Y. Structural relationship of curcumin derivatives binding to the BRCT domain of human DNA polymerase λ . Genes Cells 2006; 11: 223-235
  CrossrefPubMedSearch in Google Scholar
• 52 Wu CH, Chen CW, Chen HC, Chang WC, Shu MJ, Hung JS. Elucidating the inhibitory mechanisms of magnolol on rat smooth muscle cell proliferation. J Pharmacol Sci 2005; 99: 392-399
  CrossrefPubMedSearch in Google Scholar
• 53 Vaisman A, Woodgate R. Translesion DNA polymerases in eukaryotes: What makes them tick?. Crit Rev Biochem Mol Biol 2017; 52: 274-303
  CrossrefPubMedSearch in Google Scholar
• 54 Hubscher U, Spadani S, Villani G, Maga G. Structural and functional Aspects of the eukaryotic DNA polymerase families. In: Hubscher U, Spadani S, Villani G, Maga G. eds. DNA Polymerase Discovery, Characterization and Functions in cellular DNA Transactions. Hackensack, NJ: World Scientific; 2010: 111-160
  Search in Google Scholar
• 55 Murakami A, Ashida H, Terao J. Multitargeted cancer prevention by quercetin. Cancer Lett 2008; 269: 315-325
  CrossrefPubMedSearch in Google Scholar
• 56 Xie K, Doles J, Hemann MT, Walker GC. Error-prone translesion synthesis mediates acquired chemoresistance. Proc Natl Acad Sci U S A 2010; 107: 20792-20797
  CrossrefPubMedSearch in Google Scholar
• 57 Shaheen M, Allen C, Nickoloff JA, Hromas R. Synthetic lethality: exploiting the addiction of cancer to DNA repair. Blood 2011; 117: 6074-6082
  CrossrefPubMedSearch in Google Scholar
• 58 Lange SS, Takata K, Wood RD. DNA polymerases and cancer. Nat Rev Cancer 2011; 11: 96-110
  CrossrefPubMedSearch in Google Scholar
• 59 Doles J, Oliver TG, Cameron ER, Hsu G, Jacks T, Walker GC, Hemann MT. Suppression of Rev3, the catalytic subunit of Pol{zeta}, sensitizes drug-resistant lung tumors to chemotherapy. Proc Natl Acad Sci U S A 2010; 107: 20786-20791
  CrossrefPubMedSearch in Google Scholar

Correspondence

Publication History

Received: 26 December 2020

Accepted after revision: 07 June 2021

Article published online:
08 July 2021

© 2021. Thieme. All rights reserved.

Georg Thieme Verlag KG
Rüdigerstraße 14, 70469 Stuttgart, Germany

• References

• 1 Vineis P, Schatzkin A, Potter JD. Models of carcinogenesis: an overview. Carcinogenesis 2010; 31: 1703-1709
  CrossrefPubMedSearch in Google Scholar
• 2 Poivre M, Nachtergael A, Bunel V, Philippe ON, Duez P. Genotoxicity and Carcinogenicity of herbal Products. In: Pelkonen O, Duez P, Vuorela PM, Vuorela H. eds. Toxicology of herbal Products. Cham: Springer International Publishing; 2017: 179-215
  CrossrefSearch in Google Scholar
• 3 Balmain A. Cancer genetics: from Boveri and Mendel to microarrays. Nat Rev Cancer 2001; 1: 77-82
  CrossrefPubMedSearch in Google Scholar
• 4 Loeb LA, Springgate CF, Battula N. Errors in DNA replication as a basis of malignant changes. Cancer Res 1974; 34: 2311-2321
  PubMedSearch in Google Scholar
• 5 Migheli F, Migliore L. Epigenetic Perturbations in the Context of the multi-hit Hypothesis of Carcinogenesis. In: Maulik N, Karagiannis T. eds. Molecular Mechanisms and Physiology of Disease. New York: Springer; 2014: 383-399
  CrossrefSearch in Google Scholar
• 6 Link A, Balaguer F, Goel A. Cancer chemoprevention by dietary polyphenols: Promising role for epigenetics. Biochem Pharmacol 2010; 80: 1771-1792
  CrossrefPubMedSearch in Google Scholar
• 7 Schüz J, Espina C, Villain P, Herrero R, Leon ME, Minozzi S, Romieu I, Segnan N, Wardle J, Wiseman M, Belardelli F, Bettcher D, Cavalli F, Galea G, Lenoir G, Martin-Moreno JM, Nicula FA, Olsen JH, Patnick J, Primic-Zakelj M, Puska P, van Leeuwen FE, Wiestler O, Zatonski W, Guha N, Kralikova E, McNeill A, Peruga A, Anderson A, Berrino F, Boutron-Ruault MC, Cecchini M, Key TJ, Leitzmann M, Norat T, Powers HJ, Scoccianti C, Auvinen A, de Vries E, Erdmann F, Greinert R, Harrison J, Kesminiene A, McColl N, Friis S, Kogevinas M, Saracci R, Straif K, Vainio H, Almonte M, Anttila A, De Vuyst H, Dillner J, Franceschi S, Gonzalez P, Hall A, Park JY, Armaroli P, Atkin W, Dean PB, de Koning H, Dillner L, Kuipers E, Lansdorp-Vogelaar I, Paci E, Regula J, Suonio E, Törnberg S, Wood LF, Gaudin N, Frie KG, Terrasse V, Winstanley K, Bellisario C, Biagioli E, Cinquini M, Gianola S, Gonzalez Lorenzo M, von Karsa L, Lignini T. European code against cancer 4th edition: 12 ways to reduce your cancer risk. Cancer Epidemiol 2015; 39: S1-S10
  CrossrefPubMedSearch in Google Scholar
• 8 Wild C, Weiderpass E, Stewart BW. World Cancer Report: Cancer Research for Cancer Prevention. In: Wild C, Weiderpass E, Stewart BW. eds. World Cancer Report. Lyon, France: International Agency for Research on Cancer; 2020: 15-45
  Search in Google Scholar
• 9 Sporn MB, Dunlop N, Newton D, Smith J. Prevention of chemical carcinogenesis by vitamin A and its synthetic analogs (retinoids). Fed Proc 1976; 35: 1332-1338
  PubMedSearch in Google Scholar
• 10 Theisen C. Chemoprevention: whatʼs in a name?. J Natl Cancer Inst 2001; 93: 743
  CrossrefPubMedSearch in Google Scholar
• 11 Surh YJ. Cancer chemoprevention with dietary phytochemicals. Nat Rev Cancer 2003; 3: 768-780
  CrossrefPubMedSearch in Google Scholar
• 12 Shukla Y, Pal SK. Dietary cancer chemoprevention: an overview. Int J Hum Genet 2004; 4: 265-276
  CrossrefPubMedSearch in Google Scholar
• 13 De Flora S, Ferguson LR. Overview of mechanisms of cancer chemopreventive agents. Mutat Res Fundam Mol Mech Mutagen 2005; 591: 8-15
  CrossrefPubMedSearch in Google Scholar
• 14 Tsuda H, Ohshima Y, Nomoto H, Fujita K, Matsuda E, Iigo M, Takasuka N, Moore MA. Cancer prevention by natural compounds. Drug Metab Pharmacokinet 2004; 19: 245-263
  CrossrefPubMedSearch in Google Scholar
• 15 Ramos S. Cancer chemoprevention and chemotherapy: dietary polyphenols and signalling pathways. Mol Nutr Food Res 2008; 52: 507-526
  CrossrefPubMedSearch in Google Scholar
• 16 Aggarwal BB, Shishodia S. Molecular targets of dietary agents for prevention and therapy of cancer. Biochem Pharmacol 2006; 71: 1397-1421
  CrossrefPubMedSearch in Google Scholar
• 17 Riboli E, Norat T. Epidemiologic evidence of the protective effect of fruit and vegetables on cancer risk. Am J Clin Nutr 2003; 78: 559S-569S
  CrossrefPubMedSearch in Google Scholar
• 18 La Vecchia C. Mediterranean diet and cancer. Public Health Nutr 2004; 7: 965-968
  CrossrefPubMedSearch in Google Scholar
• 19 Heath AK, Muller DC, van den Brandt PA, Papadimitriou N, Critselis E, Gunter M, Vineis P, Weiderpass E, Fagherazzi G, Boeing H, Ferrari P, Olsen A, Tjønneland A, Arveux P, Boutron-Ruault MC, Mancini FR, Kühn T, Turzanski-Fortner R, Schulze MB, Karakatsani A, Thriskos P, Trichopoulou A, Masala G, Contiero P, Ricceri F, Panico S, Bueno-de-Mesquita B, Bakker MF, van Gils CH, Olsen KS, Skeie G, Lasheras C, Agudo A, Rodríguez-Barranco M, Sánchez MJ, Amiano P, Chirlaque MD, Barricarte A, Drake I, Ericson U, Johansson I, Winkvist A, Key T, Freisling H, His M, Huybrechts I, Christakoudi S, Ellingjord-Dale M, Riboli E, Tsilidis KK, Tzoulaki I. Nutrient-wide association study of 92 foods and nutrients and breast cancer risk. Breast Cancer Res 2020; 22: 5
  CrossrefPubMedSearch in Google Scholar
• 20 Nichols J, Katiyar S. Skin photoprotection by natural polyphenols: anti-inflammatory, antioxidant and DNA repair mechanisms. Arch Dermatol Res 2010; 302: 71-83
  CrossrefPubMedSearch in Google Scholar
• 21 Gullett NP, Ruhul Amin ARM, Bayraktar S, Pezzuto JM, Shin DM, Khuri FR, Aggarwal BB, Surh YJ, Kucuk O. Cancer prevention with natural compounds. Semin Oncol 2010; 37: 258-281
  CrossrefPubMedSearch in Google Scholar
• 22 Shiomi K, Kuriyama I, Yoshida H, Mizushina Y. Inhibitory effects of myricetin on mammalian DNA polymerase, topoisomerase and human cancer cell proliferation. Food Chem 2013; 139: 910-918
  CrossrefPubMedSearch in Google Scholar
• 23 Wu Y, Hsieh TC, Wu JM, Wang X, Christopher JS, Pham AH, Swaby JD, Lou L, Xie ZR. Elucidating the inhibitory effect of resveratrol and its structural analogs on selected nucleotide-related enzymes. Biomolecules 2020; 10: 1223
  CrossrefPubMedSearch in Google Scholar
• 24 McAuley-Hecht KE, Leonard GA, Gibson NJ, Thomson JB, Watson WP, Hunter WN, Brown T. Crystal structure of a DNA duplex containing 8-hydroxydeoxyguanine-adenine base pairs. Biochemistry 1994; 33: 10266-10270
  CrossrefPubMedSearch in Google Scholar
• 25 Arana ME, Kunkel TA. Mutator phenotypes due to DNA replication infidelity. Semin Cancer Biol 2010; 20: 304-311
  CrossrefPubMedSearch in Google Scholar
• 26 Wang D, Kreutzer DA, Essigmann JM. Mutagenicity and repair of oxidative DNA damage: insights from studies using defined lesions. Mutat Res Fundam Mol Mech Mutagen 1998; 400: 99-115
  CrossrefPubMedSearch in Google Scholar
• 27 Henderson PT, Delaney JC, Gu F, Tannenbaum SR, Essigmann JM. Oxidation of 7,8-dihydro-8-oxoguanine affords lesions that are potent sources of replication errors in vivo . Biochemistry 2001; 41: 914-921
  CrossrefPubMedSearch in Google Scholar
• 28 Henderson PT, Delaney JC, Muller JG, Neeley WL, Tannenbaum SR, Burrows CJ, Essigmann JM. The hydantoin lesions formed from oxidation of 7,8-dihydro-8-oxoguanine are potent sources of replication errors in vivo . Biochemistry 2003; 42: 9257-9262
  CrossrefPubMedSearch in Google Scholar
• 29 Henderson PT, Neeley WL, Delaney JC, Gu F, Niles JC, Hah SS, Tannenbaum SR, Essigmann JM. Urea lesion formation in DNA as a consequence of 7, 8-dihydro-8-oxoguanine oxidation and hydrolysis provides a potent source of point mutations. Chem Res Toxicol 2004; 18: 12-18
  CrossrefPubMedSearch in Google Scholar
• 30 Maki H, Sekiguchi M. MutT protein specifically hydrolyses a potent mutagenic substrate for DNA synthesis. Nature 1992; 355: 273-275
  CrossrefPubMedSearch in Google Scholar
• 31 Radicella JP, Dherin C, Desmaze C, Fox MS, Boiteux S. Cloning and characterization of hOGG1, a human homolog of the OGG1 gene of Saccharomyces cerevisiae. Proc Natl Acad Sci U S A 1997; 94: 8010-8015
  CrossrefPubMedSearch in Google Scholar
• 32 McCann JAB, Berti PJ. Adenine release is fast in MutY-catalyzed hydrolysis of G : A and 8-oxo-G : A DNA mismatches. J Biol Chem 2003; 278: 29587-29592
  CrossrefPubMedSearch in Google Scholar
• 33 Hübscher U, Maga G. DNA replication and repair bypass machines. Curr Opin Chem Biol 2011; 15: 627-635
  CrossrefPubMedSearch in Google Scholar
• 34 Maga G, Villani G, Crespan E, Wimmer U, Ferrari E, Bertocci B, Hubscher U. 8-oxo-guanine bypass by human DNA polymerases in the presence of auxiliary proteins. Nature 2007; 447: 606-608
  CrossrefPubMedSearch in Google Scholar
• 35 Kamiya H, Yamaguchi A, Suzuki T, Harashima H. Roles of specialized DNA polymerases in mutagenesis by 8-hydroxyguanine in human cells. Mutat Res Fundam Mol Mech Mutagen 2010; 686: 90-95
  CrossrefPubMedSearch in Google Scholar
• 36 Irimia A, Eoff RL, Guengerich FP, Egli M. Structural and functional elucidation of the mechanism promoting error-prone synthesis by human DNA polymerase kappa opposite the 7,8-dihydro-8-oxo-2′-deoxyguanosine adduct. J Biol Chem 2009; 284: 22467-22480
  CrossrefPubMedSearch in Google Scholar
• 37 Lee DH, Pfeifer GP. Translesion synthesis of 7,8-dihydro-8-oxo-2′-deoxyguanosine by DNA polymerase eta in vivo . Mutat Res Fundam Mol Mech Mutagen 2008; 641: 19-26
  CrossrefPubMedSearch in Google Scholar
• 38 Rodriguez GP, Song JB, Crouse GF. In vivo bypass of 8-oxodG. PLoS Genet 2013; 9: e1003682
  CrossrefPubMedSearch in Google Scholar
• 39 Tietz D. Nucleic Acid Electrophoresis. Heidelberg: Springer; 1998
  CrossrefSearch in Google Scholar
• 40 Ding L, Williams K, Ausserer W, Bousse L, Dubrow R. Analysis of plasmid samples on a microchip. Anal Biochem 2003; 316: 92-102
  CrossrefPubMedSearch in Google Scholar
• 41 Nachtergael A, Charles C, Spanoghe M, Gadenne M, Belayew A, Duez P. Measurement of translesion synthesis by fluorescent capillary electrophoresis: 7,8-Dihydro-8-oxodeoxyguanosine bypass modulation by natural products. Anal Biochem 2013; 440: 23-31
  CrossrefPubMedSearch in Google Scholar
• 42 Nachtergael A, Belayew A, Duez P. Pyrosequencing for the quantitative assessment of 8-oxodG bypass DNA synthesis. DNA Repair (Amst) 2014; 22: 147-152
  CrossrefPubMedSearch in Google Scholar
• 43 Charles C, Nachtergael A, Ouedraogo M, Belayew A, Duez P. Effects of chemopreventive natural products on non-homologous end-joining DNA double-strand break repair. Mutat Res Genet Toxicol Environ Mutagen 2014; 768: 33-41
  CrossrefPubMedSearch in Google Scholar
• 44 Shukla S, MacLennan GT, Flask CA, Fu P, Mishra A, Resnick MI, Gupta S. Blockade of β-catenin signaling by plant flavonoid apigenin suppresses prostate carcinogenesis in TRAMP mice. Cancer Res 2007; 67: 6925-6935
  CrossrefPubMedSearch in Google Scholar
• 45 Liu LZ, Fang J, Zhou Q, Hu X, Shi X, Jiang BH. Apigenin inhibits expression of vascular endothelial growth factor and angiogenesis in human lung cancer cells: implication of chemoprevention of lung cancer. Mol Pharmacol 2005; 68: 635-643
  CrossrefPubMedSearch in Google Scholar
• 46 Ono K, Nakane H. Mechanisms of inhibition of various cellular DNA and RNA polymerases by several flavonoids. J Biochem 1990; 108: 609-613
  CrossrefPubMedSearch in Google Scholar
• 47 Fox JT, Sakamuru S, Huang R, Teneva N, Simmons SO, Xia M, Tice RR, Austin CP, Myung K. High-throughput genotoxicity assay identifies antioxidants as inducers of DNA damage response and cell death. Proc Natl Acad Sci U S A 2012; 109: 5423-5428
  CrossrefPubMedSearch in Google Scholar
• 48 Sulfikkarali N, Krishnakumar N, Manoharan S, Nirmal R. Chemopreventive efficacy of naringenin-loaded nanoparticles in 7,12-dimethylbenz(a)anthracene induced experimental oral carcinogenesis. Pathol Oncol Res 2013; 19: 287-296
  CrossrefPubMedSearch in Google Scholar
• 49 Matsubara K, Saito A, Tanaka A, Nakajima N, Akagi R, Mori M, Mizushina Y. Epicatechin conjugated with fatty acid is a potent inhibitor of DNA polymerase and angiogenesis. Life Sci 2007; 80: 1578-1585
  CrossrefPubMedSearch in Google Scholar
• 50 Mizushina Y, Hirota M, Murakami C, Ishidoh T, Kamisuki S, Shimazaki N, Takemura M, Perpelescu M, Suzuki M, Yoshida H, Sugawara F, Koiwai O, Sakaguchi K. Some anti-chronic inflammatory compounds are DNA polymerase λ-specific inhibitors. Biochem Pharmacol 2003; 66: 1935-1944
  CrossrefPubMedSearch in Google Scholar
• 51 Takeuchi T, Ishidoh T, Iijima H, Kuriyama I, Shimazaki N, Koiwai O, Kuramochi K, Kobayashi S, Sugawara F, Sakaguchi K, Yoshida H, Mizushina Y. Structural relationship of curcumin derivatives binding to the BRCT domain of human DNA polymerase λ . Genes Cells 2006; 11: 223-235
  CrossrefPubMedSearch in Google Scholar
• 52 Wu CH, Chen CW, Chen HC, Chang WC, Shu MJ, Hung JS. Elucidating the inhibitory mechanisms of magnolol on rat smooth muscle cell proliferation. J Pharmacol Sci 2005; 99: 392-399
  CrossrefPubMedSearch in Google Scholar
• 53 Vaisman A, Woodgate R. Translesion DNA polymerases in eukaryotes: What makes them tick?. Crit Rev Biochem Mol Biol 2017; 52: 274-303
  CrossrefPubMedSearch in Google Scholar
• 54 Hubscher U, Spadani S, Villani G, Maga G. Structural and functional Aspects of the eukaryotic DNA polymerase families. In: Hubscher U, Spadani S, Villani G, Maga G. eds. DNA Polymerase Discovery, Characterization and Functions in cellular DNA Transactions. Hackensack, NJ: World Scientific; 2010: 111-160
  Search in Google Scholar
• 55 Murakami A, Ashida H, Terao J. Multitargeted cancer prevention by quercetin. Cancer Lett 2008; 269: 315-325
  CrossrefPubMedSearch in Google Scholar
• 56 Xie K, Doles J, Hemann MT, Walker GC. Error-prone translesion synthesis mediates acquired chemoresistance. Proc Natl Acad Sci U S A 2010; 107: 20792-20797
  CrossrefPubMedSearch in Google Scholar
• 57 Shaheen M, Allen C, Nickoloff JA, Hromas R. Synthetic lethality: exploiting the addiction of cancer to DNA repair. Blood 2011; 117: 6074-6082
  CrossrefPubMedSearch in Google Scholar
• 58 Lange SS, Takata K, Wood RD. DNA polymerases and cancer. Nat Rev Cancer 2011; 11: 96-110
  CrossrefPubMedSearch in Google Scholar
• 59 Doles J, Oliver TG, Cameron ER, Hsu G, Jacks T, Walker GC, Hemann MT. Suppression of Rev3, the catalytic subunit of Pol{zeta}, sensitizes drug-resistant lung tumors to chemotherapy. Proc Natl Acad Sci U S A 2010; 107: 20786-20791
  CrossrefPubMedSearch in Google Scholar

• Supplementary Material