In a large population-based case–control study from Northern Italy, intakes of combined fruits and vegetables, only fruits and only vegetables were associated with a 30, 21 and 24% lower risk of lung cancer, respectively. A diet rich in quercetin foods was associated with 53% lower risk of lung cancer. The inverse associations for quercetin-rich foods were seen in both women and men, ever smokers, and were strongest in the heaviest smokers. The beneficial effect of a quercetin-rich diet did not differ by histological subtypes. Analyses to examine gene–diet interaction between dietary quercetin and polymorphisms of P450 and GST genes showed no evidence that variants of these metabolic genes modulate the inverse associations between a quercetin-rich diet and lung cancer risk. Notably, in a small subset of cases with dietary information and gene expression data, we observed a downregulation of P450s genes and upregulation of GST genes in subjects with high frequency of intake of quercetin-rich foods. This finding is consistent with an influence of dietary quercetin on mRNA expression of key metabolic genes in human lung tissues and suggests a possible mechanism for the protective effect of quercetin-rich food consumption against lung cancer risk. Importantly, the metabolic genes affected by quercetin intake are key regulators of the metabolism of tobacco carcinogens, suggesting an interplay between quercetin intake, tobacco smoking and risk of lung cancer.
The vast epidemiological evidence showing that fruit intake lower the risk of lung cancer is convincing, whereas the evidence is not consistent for vegetables (2
). Recently, the National Institutes of Health–American Association of Retired Persons prospective cohort study in the USA reported no relationship between combined fruits and vegetables or either fruits or vegetables alone and lung cancer risk (37
). However, high intake of foods belonging to the Rosacea botanical family, which included some quercetin-rich foods, but not all, reduced the risk of lung cancer (37
The current literature on the relationship between quercetin-rich foods and lung cancer risk is limited and equivocal. Our data corroborate the results from prospective cohort studies (14
) and some case–control studies (16
), but not others (18
). The two prospective cohort studies were conducted in Finland. The Alpha-Tocopherol, Beta-Carotene Cancer Prevention Study (14
), including only smokers, and the Finnish Mobile Clinic Health Examination (15
) showed statistically significant 44 and 58% lower risks of lung cancer comparing highest versus lowest quartile of intake, respectively. Our finding of an inverse association among ever smokers corroborates recent results from a population-based case–control study in the USA that found a 37% lower risk of lung cancer among tobacco smokers but no relationship among never smokers (17
). The two studies showing discrepant results were either very small (103 cases/206 controls) (18
) or relied on proxy interview in 30% of the cases (19
). The latter study, however, although did not find an association for total quercetin intake, did observe a protective association with quercetin-rich onions.
Quercetin has been observed to inhibit carcinogen-induced tumors in rats (11
) and mice (10
) and cell proliferation in human lung cancer cells (38
). The mechanisms by which quercetin exerts anticarcinogenic properties are multi-fold (8
) and have been shown in both animal and experimental studies. Of interest, quercetin may prevent carcinogenesis by inhibiting expression of P450 enzymes (26
) and has been shown to inhibit hepatic CYP1A1
in rats (39
). Furthermore, experimental studies showed that quercetin also inhibited B(a
)P-induced DNA damage in human Hep G2 cells by altering CYP1A1
gene expression (20
). There are some data showing that quercetin may influence gene expression of GST enzymes, although it is unclear whether quercetin induces (26
) or inhibits GSTP1
The precise mechanism by which quercetin influences metabolic gene expression is speculative. It has been suggested that quercetin competes with PAHs-like B(a
)P for binding to the aryl hydrocarbon receptor, a transcription factor that regulates expression of the CYP1
family, including CYP1A1
). These genes are involved in activating tobacco-related procarcinogens into carcinogenic metabolites (41
). For Phase II genes, quercetin may interact with the antioxidant-responsive element, a promoter factor, and mediate the induction of Phase II genes, like GST
). In a previous study by our group, we observed an upregulation of gene expression with polymorphisms of CYP1A1
in current smokers (30
). In the present study, we found that frequent dietary intake of quercetin resulted in an upregulation of several GST
genes, including GSTM1
as well as a downregulation of several P450
genes in human non-tumor lung tissues. If confirmed, this finding may illustrate a mechanism of quercetin-related protection against tobacco-induced lung carcinogenesis.
There is evidence that variants of metabolic genes may modulate the association between specific dietary constituents, particularly crucifer-derived isothiocyanates, and lung cancer risk (43
). With respect to quercetin, Le Marchand et al.
previously reported on the modifying effect of CYP1A1 MspI
) polymorphism on the association between onions and lung cancer in 72 cases and 453 controls (19
). We extended the quercetin-gene interaction investigation beyond the single polymorphism of CYP1A1
to include additional variants of CYP450
genes in a much larger population. In our study, gene variants were not associated with lung cancer risk after accounting for multiple comparisons using Bonferroni correction. We note that this correction is conservative and may lead to false negative results (44
). Without this adjustment, we observed a suggestive gene–quercetin interaction for CYP1B1_18
= 0.01). The gene–diet analyses as well as the smoking- and histology-stratified results did not replicate Le Marchand's results. The effect of high dietary intake of quercetin-rich foods on P450s and GSTs activities, lowering their ability to biotransform procarcinogens to carcinogenic electrophiles and increasing xenobiotic elimination, respectively, may overcome the effect of individual variants of these metabolic genes.
Although this present study hypothesizes on one possible mechanism by which quercetin may exert its anticarcinogenic properties against lung cancer risk by influencing expression of P450
genes, additional mechanisms have been proposed to account for the putative anticarcinogenic effect of quercetin, including scavenging free radicals (45
), inhibiting proliferation by via cell cycle arrest (47
) and apoptosis (38
The findings of beneficial effects with a high quercetin-rich diet could also be attributed to other dietary components found in fruits and vegetables, such as isothiocyanates (found in cruciferous vegetables). In a recent meta-analysis, consumption of cruciferous vegetables was associated with lower risk of lung cancer (6
). In our study, intake of cruciferous vegetables was not statistically associated with lung cancer risk (supplementary Table 5
is available at Carcinogenesis
Online) possibly because there was a limited consumption of these dietary components in this population. Several factors suggest that quercetin may be an independent protective factor in lung cancer etiology. In the present study, the analyses for quercetin-rich foods were adjusted for other fruits and vegetables; moreover, the effective size observed for quercetin-rich foods compared with the findings for combined fruits and vegetables, as well as fruits and vegetables separately, was stronger and persisted for both men and women as well as across histological subtypes.
Study limitations include the possibility of recall bias due to the case–control study design, although the rapid recruitment protocol that allowed study enrollment and interview at the time of the diagnosis and not when the patients were in terminal conditions was designed to minimize such issues. Dietary data derived from FFQs are subject to measurement errors that may be random or systematic (49
). Moreover, because the FFQ in the EAGLE study was targeted to obtain information on specific foods, categories of interest were limited in scope and did not include portion size. The lack of information on portion size limited our assessment of quercetin intake to frequency of quercetin-rich foods consumption and not quercetin intake. Additionally, we were unable to calculate and adjust for total energy intake. Energy adjustment, although not perfect for addressing measurement error in FFQs, has been shown to be a reasonable method by some (50
), whereas others have proposed adjustment for body weight and physical activity as more appropriate methods (51
). Although we adjusted for body mass index, used sex-specific quintile of quercetin-rich food intake and conducted analyses stratified by sex, we cannot totally exclude residual measurement error, as in all dietary studies.
Cigarette smoking has been correlated with a less healthy lifestyle, including higher alcohol consumption, poor diet and lower socioeconomic status (52
). The extensive data available in our study enabled rigorous control for cigarette smoking, alcohol and other factors in the analyses, although residual confounding can never be completely ruled out. Finally, red wine is a rich source for quercetin, whereas white wine is not (29
). The EAGLE's FFQ did not collect information on red and white wine consumption separately; thus, we could not include red wine consumption as part of the summary measure for quercetin-rich foods. However, we verified that consumption of wine overall did not modify the association between quercetin-rich food and lung cancer risk by adjusting for total wine consumption. This suggested that, if any, the effect of quercetin contained in wine was modest.
To our knowledge, our study examined the largest combination of SNPs in P450 genes and the first to examine the role of GST polymorphisms in the relationship between dietary quercetin and lung cancer risk. However, there are other plausible candidate genes that could be explored, e.g. genes involved in glucuronidation and sulfation, which may lead to novel findings in future studies. Lastly, the microarray expression results were based on a small sample of adenocarcinoma cases only. Moreover, mRNA expression may not predict protein expression levels due to posttranscriptional and posttranslational modifications as well as other factors. Therefore, this finding requires confirmation in a larger population with protein expression data.
Our study has several strengths. It is a large population-based case–control study with high participation rates and detailed information on smoking history as well as many other risk factors. The large sample size permitted investigation by histological subtypes and smoking status with adequate power. The comprehensive genotype data on metabolic genes permitted selection of specific candidate genes for an investigation of gene–diet interaction that extends beyond previous studies on one or a couple of genes. Additional data on mRNA expression from human lung tissues enabled an investigation into the influence of dietary quercetin on expression of metabolic genes. And lastly, cases were rapidly ascertained and surrogate participants were not needed.
In conclusion, higher frequencies of intake of quercetin-rich foods were associated with lower risk of lung cancer in this Italian population. The inverse association did not differ by histological subtypes, were stronger among heavy smokers, and was not affected by variants of P450 and GST genes. Downregulation of several P450 genes and upregulation of GST genes involved in the metabolism of tobacco carcinogens were observed in human lung tissues of subjects consuming high quercetin-rich foods. This finding provides potential provocative mechanistic insights into the role of dietary quercetin in tobacco-induced lung carcinogenesis. Further studies exploring this relationship are warranted.