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Perspective on this Article from Cruciferous Vegetable Feeding Alters UGT1A1 Activity: Diet- and Genotype-Dependent Changes in Serum Bilirubin in a Controlled Feeding Trial

Increased adiposity and diets high in glycemic load (GL) are associated with increased risk of many chronic diseases including cancer. Using plasma from 80 healthy individuals [40 men/40 women, 29 with DXA-derived low fat mass (FM) and 51 with high FM] in a randomized cross-over–controlled feeding trial and arrays populated with 3,504 antibodies, we measured plasma proteins collected at baseline and end of each of two 28-day controlled diets: a low GL diet high in whole grains, legumes, fruits, and vegetables (WG) and a high GL diet high in refined grains and added sugars (RG). Following univariate testing for proteins differing by diet, we evaluated pathway-level involvement. Among all 80 participants, 172 proteins were identified as differing between diets. Stratifying participants by high and low FM identified 221 and 266 proteins, respectively, as differing between diets (unadjusted P < 0.05). These candidate proteins were tested for overrepresentation in Reactome pathways, corresponding to 142 (of 291) pathways in the high-FM group and 72 (of 274) pathways in the low-FM group. We observed that the cancer-related pathways, DNA Repair, DNA Replication, and Cell Cycle, were overrepresented in the high-FM participants while pathways involved in post-translational protein modification were overrepresented in participants with either FM. Although high-GL diets are associated with increased risk of some cancers, our study further suggests that biology associated with consumption of GL diets is variable depending on an individual's adiposity and dietary recommendations related to cancer prevention be made with the additional consideration of an individual's FM.

Increased adiposity and diets high in glycemic load (GL) are associated with increased risk of many chronic diseases including cancer. Using plasma from 80 healthy individuals [40 men/40 women, 29 with DXA-derived low fat mass (FM) and 51 with high FM] in a randomized cross-over–controlled feeding trial and arrays populated with 3,504 antibodies, we measured plasma proteins collected at baseline and end of each of two 28-day controlled diets: a low GL diet high in whole grains, legumes, fruits, and vegetables (WG) and a high GL diet high in refined grains and added sugars (RG). Following univariate testing for proteins differing by diet, we evaluated pathway-level involvement. Among all 80 participants, 172 proteins were identified as differing between diets. Stratifying participants by high and low FM identified 221 and 266 proteins, respectively, as differing between diets (unadjusted P < 0.05). These candidate proteins were tested for overrepresentation in Reactome pathways, corresponding to 142 (of 291) pathways in the high-FM group and 72 (of 274) pathways in the low-FM group. We observed that the cancer-related pathways, DNA Repair, DNA Replication, and Cell Cycle, were overrepresented in the high-FM participants while pathways involved in post-translational protein modification were overrepresented in participants with either FM. Although high-GL diets are associated with increased risk of some cancers, our study further suggests that biology associated with consumption of GL diets is variable depending on an individual's adiposity and dietary recommendations related to cancer prevention be made with the additional consideration of an individual's FM.

We and others have reported associations between B vitamins principally involved in one-carbon metabolism and increased lung cancer risk; however results for women have been inconsistent. Here we report on the association of supplemental vitamins B6, folic acid, and B12 intake and lung cancer risk using data from the Women's Health Initiative (WHI) study of postmenopausal women. Between 1993 and 1998, 161,808 women were recruited to participate in the WHI at 40 US clinical centers. After exclusions, 159,232 women were available for analysis and followed prospectively for 18.3 years. Among them, 3,836 incident lung cancer cases were diagnosed. At baseline, supplemental B vitamins from multivitamins, vitamin mixtures, and individual supplements were assessed. Adjusted Cox regression models were used to estimate hazard ratios (HR) and 95% confidence intervals (CI) for associations between supplemental B vitamin intake and lung cancer risk. Relative to non-users, women who took ≥50 mg/d of vitamin B6 had 18% (HR 0.82, 95% CI: 0.70-0.97) reduced lung cancer risk. Associations did not differ significantly by smoking status or lung cancer histology. Intakes of folic acid and vitamin B12 were not associated with risk. There is a need for replication of our findings from large, prospective studies with high-quality measurement of supplement intakes. As such, no recommendation can be made at present on the use of B6 supplements for lung cancer prevention in women. Citation Format: Theodore M. Brasky, Roberta M. Ray, Sandi L. Navarro, Jeannette M. Schenk, Alison M. Newton, Marian L. Neuhouser. Supplemental one-carbon metabolism related B vitamins and lung cancer risk in the Women's Health Initiative [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4654.

Background: Biomarkers of low-grade systemic inflammation are used to study the associations of inflammation with chronic diseases, including cancer. However, relatively little is known about the intraindividual variability of most of these measures. Methods: Fasting serum samples, collected at baseline and the end of ≥3-week washout periods in a four-diet crossover feeding trial, were used to measure the inflammatory markers high sensitivity C-reactive protein (hsCRP), interleukin (IL)-6, tumor necrosis factor (TNF)-α, IL-8, and soluble TNF receptor (sTNFR) I and II. Participants included 62 men and women for analyses of IL-6 and CRP and 56 for analyses of IL-8, TNF-α, and sTNFRs, aged 20 to 40, who were free of factors known to influence inflammation, for example, chronic disease, medication use, heavy alcohol use, smoking, and obesity (body mass index >30 kg/m2). Intraclass correlations (ICC) were estimated using random effects ANOVA, across all four time points (∼6 weeks apart). Results: ICCs for TNF-α and sTNFR I and II were very high: ICC = 0.92 [95% confidence interval (CI), 0.89–0.96], 0.92 (95% CI, 0.88–0.95), and 0.90 (95% CI, 0.85–0.94), respectively. ICCs for IL-8 and hsCRP were 0.73 (95% CI, 0.63–0.83) and 0.62 (95% CI, 0.49–0.75), respectively. The ICC for IL-6 was considerably lower, ICC = 0.48 (95% CI, 0.36–0.62). Three measures of IL-6 would be needed to achieve a reliability coefficient (Cronbach α) of 0.75. Conclusions: With the exception of IL-6, reliability of all inflammatory markers in our panel was high. Impact: This suggests that a single measure accurately captures the short-term (e.g., 4–6 months) variability within an individual. Cancer Epidemiol Biomarkers Prev; 21(7); 1167–70. ©2012 AACR.

Glutathione S-transferases (GST) detoxify a wide range of carcinogens. Isothiocyanates (ITC), from cruciferous vegetables, are substrates for and inducers of GST. GST variants may alter ITC clearance such that response to crucifers varies by genotype. In a randomized cross-over trial, we tested the hypothesis that changes in serum GSTA1/2 concentration in response to cruciferous vegetable feeding depends on GSTM1/GSTT1 genotype. Thirty-three men and 34 women (age 20-40 years) ate four 14-day controlled diets—basal (vegetable-free), basal supplemented with two different doses of crucifers (“single dose” and “double dose”), and single-dose cruciferous-plus-apiaceous vegetables—fed per kilogram of body weight. Fasting bloods from days 0, 7, 11, and 14 of each diet period were analyzed for serum GSTA1/2 by ELISA. GSTA1/2 increased with single- and double-dose cruciferous compared with basal diet (10% and 13%, respectively; P = 0.02 and 0.004), but cruciferous-plus-apiaceous did not differ from basal (P = 0.59). Overall, GSTA1/2 was higher in GSTM1-null/GSTT1-null than GSTM1+/GSTT1+ individuals (4,198 ± 338 and 3,372 ± 183 pg/mL; P = 0.03). The formal interaction of genotype-by-diet was not statistically significant, but the GSTA1/2 increase during the single-dose cruciferous diet was among GSTM1-null/GSTT1-null individuals (by 28%; P = 0.008), largely explained by GSTM1-null/GSTT1-null men (by 41%; P = 0.01). GSTA1/2 increased during the double-dose cruciferous diet in both GSTM1-null/GSTT1-null men (by 35%; P = 0.04) and GSTM1+/GSTT1+ men (by 26%; P = 0.01) but not in women. In summary, cruciferous vegetable supplementation increased GSTA1/2, but the effect was most marked in GSTM1-null/GSTT1-null men. (Cancer Epidemiol Biomarkers Prev 2009;18(11):2974–8)

Higher consumption of cruciferous vegetables is associated with lower risk of several cancers. Isothiocyanates (ITC), the bioactive compounds derived from glucosinolates in cruciferous vegetables, are thought to exert chemoprotective effects through modulation of several pathways critical to carcinogenesis. Urinary ITC excretion is used in epidemiologic studies as a measure of cruciferous vegetable intake; however, considerable inter-individual variation in ITC excretion is observed. Several factors may influence glucosinolate metabolism and excretion, including gut microbial hydrolysis and genetic variation in human enzymes. Thus, intake is not necessarily equivalent to ITC exposure. In a randomized cross-over trial, we evaluated total urinary ITC excretion in healthy participants (n=12 men; n=13 women) in response to 100 μmol glucosinolate intake from three acute feedings of cooked cruciferous vegetables– watercress (67 g), broccoli (125 g), and cabbage (250 g). The vegetables were fed alongside a standardized frozen pasta meal (227 g). GSTM1 genotyping (present or null) was conducted on buccal cell DNA. Total urinary ITC excretion (dithiocarbamates plus glutathione-derived conjugates) was assayed by HPLC from 24 h urine collections adjusted for completeness by creatinine. Linear mixed models were used to assess the effects of dietary exposure and urinary ITC excretion. Mean urinary ITC excretion was 7.21 (95% CI: 5.81, 8.94); 8.35 (95% CI: 6.73, 10.35); and 10.08 (95% CI: 8.20, 12.41) μmol/24 h for watercress, cabbage and broccoli, respectively. As a group, participants excreted significantly more ITC after consumption of broccoli compared to watercress (P=0.01). Intra- and inter-individual variances across all 3 vegetables were 0.29 and 0.07, respectively, and the intra-class correlation (ICC) was 0.20 when evaluating the three dietary measures as replicates of 100 μmol glucosinolate intakes. There were no main effects of GSTM1 genotype (n=9 positive; n=16 null) or ethnicity (n=14 Caucasians; n=9 Asians). Urinary ITC excretion profiles significantly differed between broccoli and watercress, as well as within and between individuals (ICCs below 0.45 indicate poor correlation between repeated measures). This suggests that ITC bioavailability within individuals differs by type of crucifer consumed. ITC are involved in the modulation of several pathways involved in carcinogenesis, therefore an individual's actual exposure to ITC may affect their ultimate risk of cancer. Citation Format: Sandi L. Navarro, Hannah Frenkel, Wendy K. Thomas, Karen Makar, C.Y. Wang, Johanna W. Lampe. Urinary isothiocyanate excretion profiles in response to acute feeding of various cruciferous vegetables in humans. [abstract]. In: Proceedings of the Eleventh Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2012 Oct 16-19; Anaheim, CA. Philadelphia (PA): AACR; Cancer Prev Res 2012;5(11 Suppl):Abstract nr A101.

Biomarkers of low-grade systemic inflammation are used to study the associations of inflammation with chronic diseases, including cancers. However, little is known about the reliability (i.e., intraindividual variability) of most of these measures. Fasting (12-hour) serum samples, which were collected at the end of ≥3 week washout periods in a 4-diet crossover feeding trial, were used to measure the inflammatory markers hsCRP, IL-6, TNF-α, IL-8 and sTNFRI and II. Participants included 32 men and 31 women (5 men and 2 women were missing from analyses of IL-8, TNF-α, and sTNFR I and II), aged 20-40, who were free of factors known to influence inflammation, e.g., chronic disease, medication use, heavy alcohol use, smoking and obesity (body mass index >30 kg/m2). Intraclass correlations (ICC) were estimated using random effects analysis of variance, across all 4 time points (∼5 weeks apart). For TNF-α and sTNFRI & II, the ICCs were very high (ICC= 0.92, 95% CI: 0.89 - 0.96; 0.92, 95% CI: 0.88 - 0.95; and 0.90, 95% CI: 0.85 - 0.94, respectively). The ICCs for IL-8 and hsCRP were 0.73 (95% CI: 0.63-0.83 and 0.68 (95% CI: 0.58-0.78), respectively. The ICC for IL-6 was considerably lower, (ICC = 0.48, 95% CI: 0.36-0.62). With the exception of IL-6, reliability of all inflammatory markers in our panel over 4 time points was high, suggesting that a single measure accurately captures the variability within an individual and is a good measure for use in epidemiologic studies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5486. doi:1538-7445.AM2012-5486

Although there is evidence from studies of prostate cancer cell lines and rodent models that several supplements may have anti-inflammatory, anti-oxidant, or other anti-cancer properties, few epidemiologic studies have examined the association between non-vitamin, non-mineral, specialty supplement use and prostate cancer risk. Participants, 50-76 years, were 35,239 male members of the VITamins And Lifestyle (VITAL) cohort who were residents of western Washington State, and who completed an extensive baseline questionnaire in 2000-2002. Participants responded about their frequency (days/week) and duration (years) of specialty supplement uses. 1,602 incident invasive prostate cancers were obtained from the Surveillance, Epidemiology, and End Results registry. Multivariate-adjusted hazards ratios (HR) and 95% confidence intervals (95% CI) were estimated by Cox proportional hazards models. Any use of grapeseed supplements was associated with a 41% (HR 0.59, 95% CI: 0.40-0.86) reduced risk of total prostate cancer. Although not statistically different, the association was somewhat stronger for low-grade (HR 0.58, 95% CI: 0.33-1.03) than high-grade tumors (HR 0.82, 95% CI: 0.34-2.00). There were no associations for use of chondroitin, co-enzyme Q10, fish oil, garlic, ginkgo biloba, ginseng, glucosamine, or saw palmetto. Grapeseed may be a potential chemopreventive agent, however as current evidence is limited, it should not yet be promoted for prevention of prostate cancer. This work is supported by NCI grants R25-CA94880, R01-CA142545, and K05-CA154337 Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1915. doi:10.1158/1538-7445.AM2011-1915

Isothiocyanates (ITC) in cruciferous vegetables modulate several signaling pathways involved in carcinogenesis, including nuclear factor kappa-B (NF-κB), a transcription factor and central mediator in the generation of inflammation. Glutathione S-transferase (GST)M1 & GSTT1 metabolize ITC; genotypic differences in biologic response to cruciferous vegetable intake, and variation in ITC pharmacokinetics have been reported. Previously, we found reduced serum IL-6 and CRP concentrations in response to cruciferous (broccoli family) and apiaceous (carrot family) vegetable feeding compared to a basal diet in a randomized, crossover trial. Reductions were most marked among individuals with a GSTM1-null genotype. Here, we extend our panel of biomarkers in this intervention to include serum interleukin (IL)-8, tumor necrosis factor-alpha (TNF-α), and soluble TNF receptors I & II (sTNFRI & II). We tested whether supplementation of cruciferous or apiaceous vegetables alters serum concentrations of these markers, and whether this response is GSTM1/GSTT1 genotype-dependent. Men (n=26) and women (n=30), 20-40 y, ate four 14-day controlled diets – basal (fruit &vegetable-free), basal + two doses of crucifers (single & double “dose”), and single-dose cruciferous + apiaceous vegetables – fed per kg body weight. Fasting bloods from days 0 & 14 of each period were analyzed for serum inflammatory markers (IL-8 & TNF-a using the High Sensitivity Human Cytokine Panel, sTNFRs using the Human Soluble Cytokine Receptor Panel, Millipore). In this healthy population, we observed no overall reduction in any of the current biomarkers in response to vegetable supplementation. However, similarly to what we observed with IL-6 and CRP, both sTNFRI & II were lower at day 14 on the double-dose cruciferous vegetable diet among GSTM1-null individuals (P=0.006 & 0.007, respectively). NF-kB is one regulator of these inflammatory markers; but overlap with other pro-inflammatory pathways that may be differently regulated by diet, e.g., STAT3, may preclude observation of an effect. Such diet interventions may be more effective in populations with higher circulating inflammation biomarkers. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3682. doi:10.1158/1538-7445.AM2011-3682

Regular use of aspirin is associated with lower colon cancer risk. High inter-individual variation in aspirin metabolism has been attributed to several factors, including sex, concomitant use of other drugs or alcohol, urine pH, ethnicity, and variants in metabolizing enzymes involved in glucuronidation of aspirin metabolites [i.e., UDP glucuronosyltransferases (UGT)]. However, little is known about the impact of other exposures, including dietary factors. Dietary constituents of citrus fruits, cruciferous vegetables and soy have been shown to induce UGT. Higher intakes may alter UGT activity and affect the ratio of aspirin metabolites excreted. We evaluated, cross-sectionally, whether urinary excretion of aspirin [ASA] and its metabolites (salicylic acid [SA], salicyluric acid [SUA], salicyluric phonolic glucuronide [SUPG], salicyl acyl and phenolic acid glucuronides [SAG/SPG]) differed by UGT1A6 genotype and dietary factors. Following an oral dose of 650 mg aspirin, healthy men (N=264) and women (N=264), 20–40 years, collected urine over an 8-h period. Participant exclusion criteria included medical history of gastrointestinal, hepatic or renal disorders, and exposure to non-dietary factors known to influence biotransformation enzymes, e.g., medications, excessive alcohol, and smoking. FFQ data were available for 481 participants. Multiple linear regression was used to determine whether there were differences in aspirin metabolism by UGT1A6 genotype, or daily intake of protein, total fruit or vegetable intake, and soy, citrus and cruciferous vegetable intake, adjusted for body mass index (BMI), age, sex, ethnicity, urine volume and pH, and energy intake. There were statistically significant differences in the ratios of metabolites excreted, (specific metabolite/sum of metabolites), between sexes and between ethnic groups. Men excreted more SUA (P Of the factors evaluated, ethnicity and sex were the greatest, albeit modest, contributors to variability in aspirin metabolism (R2=2−13%), whereas UGT1A6 had no effect. Our results also suggest that diet may influence aspirin metabolism, but the effects do not appear to be via glucuronidation. Citation Information: Cancer Prev Res 2010;3(1 Suppl):B99.