Search

In the original version of [this article][1] ([1][2]), authors Thomas Karasic and Jeffrey Meyer are mistakenly omitted from the author list. This error has been corrected in the latest online HTML and PDF versions of the article and the author contributions and disclosures have been updated as

Collectively liver cancer accounts for nearly 9% of all reported cancer deaths and is one of the most common causes of cancer mortality worldwide http://gco.iarc.fr/ (1). The incidence of liver cancer varies enormously globally and unfortunately the burden of this nearly always fatal disease is much greater in the less economically developed countries of Asia and sub-Saharan Africa (2). HCC is also the most rapidly rising solid tumor in the US and Central America and is overrepresented in minority communities, including African-Americans, Hispanic/Latino-Americans and Asian-Americans (1,3,4). Overall, there are more than 850,000 new cases each year and more than 300,000 deaths annually in the People’s Republic of China (P.R.C.) alone (2). In contrast with most common cancers in the economically developed world where over 90% of cases are diagnosed after the age of 45, in high-risk regions for liver cancer onset begins to occur in both men and women by 20 years of age and peaks between 40-49 years of age in men and between 50-59 years of age in women (5-7). Gender differences in liver cancer incidence have also been well described and worldwide the number of cases among men were 554,000 and 228,000 among women in 2012 (8)(9). To date, the significant etiological factors associated with development of HCC have been defined by biomarker studies and they are infection in early life with hepatitis B virus (HBV) and lifetime exposure to high levels of aflatoxin B1 (AFB1) in the diet (10,11). Over the past 25 years, an appreciation for the role of the hepatitis C virus (HCV) has also emerged. HCV is contributing to HCC being the most rapidly rising solid tumor in the US and Japan (12). Detailed knowledge of the etiology of HCC has spurred many mechanistic studies to understand the pathogenesis of this nearly always-fatal disease (10,13,14). Fortunately, the successful development and deployment of some highly effective new drugs that cure HCV infection is a major advance and will hopefully diminish the role of this virus in liver cancer (15,16). Alcohol is a recognized human carcinogen and alcoholic cirrhosis and heavy alcohol use have been repeatedly associated with an increase in HCC risk (23). However, it is unclear if alcohol use in the absence of cirrhosis influences HCC development (24). Several studies have demonstrated an increased risk of HCC up to 5-fold with consumption of more than 80g of alcohol per day or approximately 6-7 drinks per day (23). The risk of HCC ranges from borderline significant to doubled with chronic alcohol consumption of less than 80g/day (23). A synergism between alcohol and HBV and HCV infections has also been described (23,25). In economically developed countries the dramatic rise in overweight and nonalcoholic fatty liver disease has also been related to increased HCC (26-28). Of major concern for the future are the role that obesity, diabetes and general underlying fatty liver disease will play in the development of liver cancer (29-31). Both therapeutic and pre-disease interventions will need to be deployed now to blunt the impact of these risk factors in the decades to come. Collectively, the development and validation of a new generation of biomarkers reflecting complex processes such as inflammation will need to be deployed. References 1. Ferlay, J., et al. (2015) Cancer incidence and mortality worldwide: sources, methods and major patterns in GLOBOCAN 2012. Int J Cancer., 136, E359-86. doi: 10.1002/ijc.29210. Epub 2014 Oct 9. 2. (2014) World Cancer Report 2014. International Agency for Research on Cancer. 3. (2005) American Cancer Society Cancer Facts and Figures 2005. 22-27. 4. El-Serag, H.B., et al. (2014) - Epidemiology of hepatocellular carcinoma in the United States: where are we? - Hepatology. 2014 Nov;60(5):1767-75. doi: 10.1002/hep.27222. Epub 2014 Aug 25., 1767-75. 5. Chen, J.G., et al. (2006) Trends in the incidence of cancer in Qidong, China, 1978-2002. Int J Cancer, 119, 1447-54. 6. Vatanasapt, V., et al. (1995) Cancer incidence in Thailand, 1988-1991. Cancer Epidemiol.Biomarkers Prev., 4, 475-483. 7. Parkin, D.M., et al. (2005) Global cancer statistics, 2002. CA Cancer J Clin, 55, 74-108. 8. Torre, L.A., et al. (2015) Global cancer statistics, 2012. CA Cancer J Clin., 65, 87-108. doi: 10.3322/caac.21262. Epub 2015 Feb 4. 9. Wogan, G.N., et al. (1967) Dose-response characteristics of aflatoxin B1 carcinogenesis in the rat. Cancer Res, 27, 2370-6. 10. Kensler, T.W., et al. (2003) Translational strategies for cancer prevention in liver. Nature Reviews, 3, 321-329. 11. Block, T.M., et al. (2003) Molecular viral oncology of hepatocellular carcinoma. Oncogene, 22, 5093-5107. 12. Tanaka, V., et al. (2002) A comparison of the molecular clock of hepatitis C virus in the United States and Japan predicts that hepatocellular carcinoma incidence in the United States will increase over the next two decades. Proc.Natl.Acad.Sci.USA, 99, 15584-15589. 13. Kensler, T.W., et al. (2004) Chemoprevention of hepatocellular carcinoma in aflatoxin endemic areas. Gastroenterology, 127, S310-S318. 14. Groopman, J.D., et al. (2008) Protective interventions to prevent aflatoxin-induced carcinogenesis in developing countries. Annu Rev Public Health, 29, 187-203. 15. Stasi, C., et al. (2015) The epidemiological changes of HCV and HBV infections in the era of new antiviral therapies and the anti-HBV vaccine. J Infect Public Health, 004. 16. Barth, H. (2015) Hepatitis C virus: Is it time to say goodbye yet? Perspectives and challenges for the next decade. World J Hepatol., 7, 725-37. doi: 10.4254/wjh.v7.i5.725. 17. Yu, M.C., et al. (2004) Environmental factors and risk for hepatocellular carcinoma. Gastroenterology, 127, S72-8. 18. Falk, H., et al. (1974) Hepatic disease among workers at a vinyl chloride polymerization plant. JAMA, 230, 59-63. 19. Dragani, T.A., et al. (2008) Occupational exposure to vinyl chloride and risk of hepatocellular carcinoma. Cancer Causes Control. 20. Forman, D., et al. (1985) Exposure to vinyl chloride and angiosarcoma of the liver: a report of the register of cases. Br J Ind Med, 42, 750-3. 21. Mastrangelo, G., et al. (2004) Increased risk of hepatocellular carcinoma and liver cirrhosis in vinyl chloride workers: synergistic effect of occupational exposure with alcohol intake. Environ Health Perspect, 112, 1188-92. 22. Wong, R.H., et al. (2003) Interaction of vinyl chloride monomer exposure and hepatitis B viral infection on liver cancer. J.Occ.Med., 45, 379-383. 23. Morgan, T.R., et al. (2004) Alcohol and hepatocellular carcinoma. Gastroenterology, 127, S87-96. 24. Fattovich, G., et al. (2004) Hepatocellular carcinoma in cirrhosis: incidence and risk factors. Gastroenterology, 127, S35-50. 25. Singal, A.K., et al. (2007) Mechanisms of synergy between alcohol and hepatitis C virus. J Clin Gastroenterol, 41, 761-72. 26. Takamatsu, S., et al. (2008) Influence of risk factors for metabolic syndrome and non-alcoholic fatty liver disease on the progression and prognosis of hepatocellular carcinoma. Hepatogastroenterology, 55, 609-14. 27. Ohki, T., et al. (2008) Obesity is an independent risk factor for hepatocellular carcinoma development in chronic hepatitis C patients. Clin Gastroenterol Hepatol, 6, 459-64. 28. El-Serag, H.B. (2007) Epidemiology of hepatocellular carcinoma in USA. Hepatol Res, 37 Suppl 2, S88-94. 29. Khan, F.Z., et al. (2015) Advances in hepatocellular carcinoma: Nonalcoholic steatohepatitis-related hepatocellular carcinoma. World J Hepatol., 7, 2155-61. doi: 10.4254/wjh.v7.i18.2155. 30. Saran, U., et al. (2015) Hepatocellular Carcinoma and Lifestyles. J Hepatol, 028. 31. McGlynn, K.A., et al. (2015) Global epidemiology of hepatocellular carcinoma: an emphasis on demographic and regional variability. Clin Liver Dis., 19, 223-38. doi: 10.1016/j.cld.2015.01.001. Epub 2015 Feb 26. Citation Format: John D. Groopman. The changing etiology of liver cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr SY27-01.

Many components of outdoor air pollution, such as benzene, are known to be carcinogenic to humans. However, due to spatial and temporal variation in air pollution, ambient measures of single pollutant exposures may be limited for generalizing across diverse settings. Covalent adducts of air pollutants with the free thiol at the Cys34 residue of human serum albumin represent biomarkers of recent air pollution exposure ( Citation Format: Joshua W. Smith, Robert N. O'Meally, Derek Ng, Thomas W. Kensler, Robert N. Cole, John D. Groopman. Signatures of chemoprotection from ambient exposure to outdoor air pollution in the human serum albumin Cys34 adductome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 603.

Hepatocellular carcinoma (HCC) continues to be a major cause of cancer death globally, with aflatoxin B1 (AFB1) as a prevalent risk factor in low- and middle-income countries. MicroRNAs have been shown to be differentially expressed in HCC and may serve as predictive biomarkers. In this study, we analyzed tumor and non-tumor tissue from rats dosed with AFB1 (200 µg/kg BW) for 28 consecutive days that received vehicle only or AFB1 plus the chemopreventive agent CDDO-Im (30 µmol), and control animals (Johnson et al., CaPR, 2014). Total RNA was isolated from tumor or non-tumor tissue at the time of sacrifice and sequenced. MicroRNA transcriptomic analysis revealed 17 miRNAs significantly upregulated (> 5 fold) in tumors compared to non-tumor tissue. The top ten dysregulated miRNAs determined by fold change and biological significance were selected for further investigation: rno-miR- 205, 200b-3p, 182, 429, 31a-5p, 10b-5p, 141-3p, 132-3p, 802-5p. Validation of sequencing results by quantitative PCR (qPCR) confirmed the upregulation of the majority of candidate miRNAs in tumors and rno-miR-224-5p as the most dysregulated miRNA (over 400 fold). We also examined the levels of these candidates in terminal sera of the same animals by qPCR. Circulating miRs-224-5p, 182, and 122-5p were increased (> 1.5 fold) in animals diagnosed with HCC compared to untreated animals and those previously dosed with AFB1 plus CDDO-Im. Analysis of tracking of serum miR-182 by generalized estimating equations (GEE) revealed significantly increased levels (5 fold; CI: 2.17- 2.50) in animals that developed HCC. This sustained increase in serum miR-182 is seen months before any tumors or other symptoms are present, and highlights this miRNA as a potential predictive biomarker in AFB1-induced HCC. Supported by T32ES007141-31A1 and CA197222. Citation Format: Merricka C. Livingstone, Natalie M. Johnson, Bill D. Roebuck, Thomas W. Kensler, John D. Groopman. Dysregulated microRNAs in aflatoxin-induced hepatocellular carcinoma: Serum miR-182 as a potential predictive biomarker [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5401.

A novel acetylenic tricyclic bis-(cyano enone), TBE-31, is a lead compound in a series of tricyclic compounds with enone functionalities in rings A and C. Nanomolar concentrations of this potent multifunctional molecule suppress the induction of the inflammatory protein, inducible nitric oxide synthase, activate phase 2 cytoprotective enzymes in vitro and in vivo, block cell proliferation, and induce differentiation and apoptosis of leukemia cells. Oral administration of TBE-31 also significantly reduces formation of aflatoxin-DNA adducts and decreases size and number of aflatoxin-induced preneoplastic hepatic lesions in rats by >90%. Because of the two cyano enones in rings A and C, TBE-31 may directly interact with DTT and protein targets such as Keap1 that contain reactive cysteine residues. The above findings suggest that TBE-31 should also be tested for chemoprevention and chemotherapy in relevant models of cancer and against other chronic, degenerative diseases in which inflammation and oxidative stress contribute to disease pathogenesis. [Cancer Res 2008;68(16):6727–33]

Background: Aflatoxin, which is a risk factor for developing hepatocellular carcinoma, may also cause gallbladder cancer (GBC). We hypothesized that GBC patients have higher aflatoxin exposure than gallstone patients. Methods: We used samples and data from the Shanghai Biliary Tract Cancer Case-control Study conducted from 1997-2001. Of the 267 GBC and 730 gallstone participants with plasma available, 250 GBC patients and 250 controls with gallstones were randomly selected for measurement of aflatoxin B1 (AFB1)-lysine plasma adducts using isotope dilution mass spectrometry. In 54 GBC patients, tumor tissue was examined for the aflatoxin-related R249S TP53 mutation through targeted sequencing. Results: We calculated age- and sex-adjusted odds ratios (ORs) and 95% confidence intervals (CIs), as well as the population attributable fraction (PAF). AFB1-lysine adduct detection was nearly 3-fold higher in GBC patients (OR, 2.96; 95% CI, 1.90 to 4.61). Among the 85 GBC patients (34%) with detectable AFB1-lysine adduct levels, the median was 4.6 pg/mg, compared to 1.2 pg/mg among the 37 gallstone-patient controls (15%) with detectable levels, and the OR was 9.97(95% CI, 2.73 to 36.44) for quartile 4 versus 1 in GBC compared to gallstone patients. The PAF for cancer related to aflatoxin was 22% (95% CI: 17%-27%). In 54 GBC patients, tumor tissue was examined for the aflatoxin-related R249S TP53 mutation through targeted sequencing, however, all patients were negative for the mutation. Conclusions: Aflatoxin is significantly associated with GBC, although it does not appear to be correlated with the R249S TP53 mutation in the gallbladder. If aflatoxin is shown to cause GBC, it may have accounted for up to 22% of the gallbladder cancers in Shanghai during the study period and may account for an even higher proportion in high-risk areas. If verified, reducing aflatoxin exposure may reduce GBC incidence. Citation Format: Jill E. Koshiol, Yu-Tang Gao, Michael Dean, Patricia Egner, Chirag Nepal, Kristine Jones, Bingsheng Wang , Asif Rashid, Wen Lou, Alison Van Dyke, Catterina Ferreccio, Michael Malasky, Ming-Chang Shen, Bin zhu, Jesper Andersen, Allan Hildesheim, Ann W. Hsing, John Groopman. Association of aflatoxin and gallbladder cancer in Shanghai [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-157. doi:10.1158/1538-7445.AM2017-LB-157

Low-income countries across the world are increasingly burdened with growing rates of cancer incidence and mortality. Opportunities for prevention in these populations may be realized most readily for cancer sites with known etiologies that are amenable to intervention. Liver cancer is the sixth-most commonly occurring cancer worldwide and is the second-most common contributor to global cancer mortality. Aflatoxin B1 (AFB1) is a known and potent human hepatocarcinogen, with exposure occurring via ingestion of grains - especially maize - contaminated with Aspergillus flavus. Previous work in regions with endemic levels of liver cancer has shown that reducing AFB1 exposure attenuated liver cancer mortality - even in the presence of other known etiological exposures, such as hepatitis B virus. Guatemala experiences high rates of liver cancer - which is uniquely distributed equally between men and women - and consumes maize as a dietary staple, making the assessment of AFB1 exposure a high priority for cancer prevention efforts in this country. Therefore, we measured circulating levels of the AFB1-albumin adduct, an established biomarker of AFB1 exposure, in adults residing in five geographically and ecologically dispersed departments of Guatemala (Guatemala, Suchitepéquez, Sololá, Escuintla, and Quiché). Healthy men and women aged 40 and older were recruited for the study. Pregnant women or individuals with implanted cardiac devices were excluded from participation. Subjects (n=436) gave informed consent and provided demographic information and a venipuncture blood sample during a clinic visit. Serum was separated from whole blood, digested with protease, subjected to solid-phase extraction, and analyzed with isotope dilution ultrahigh-performance liquid chromatography-triple quadrupole mass spectrometry. Serum albumin was measured by ELISA. AFB1-albumin adducts (pg AFB1-lysine adduct/mg albumin) were detectable in 100% of samples, with a mean ± SEM of 21.6 ± 2.5 and range of 0.2 - 814.8. These levels are similar to those observed in regions of China in the 1980s, prior to reductions in AFB1 exposure and liver cancer mortality. Stratification by demographic factors revealed significant differences in aflatoxin levels in urban (9.6 ± 1.3) versus rural (28.8 ± 3.9) subjects (p ≤ 0.001). Accordingly, AFB1-albumin adduct levels were lowest in the urban municipality of Escuintla (5.8 ± 1.1) and highest in rural Suchitepéquez (36.4 ± 4.8). Exposure was significantly different by sex, with men having higher levels compared to women (30.0 ± 5.4 vs. 15.3 ± 1.3, p = 0.01). These data show that AFB1 exposure in Guatemalan adults is similar to levels previously shown to increase liver cancer risk. Larger cohort and prospective studies are warranted to assess the relationship between liver cancer incidence and AFB1 exposure in this population. Citation Format: Joshua W. Smith, Patricia A. Egner, Mariana Lazo, Alvaro Rivera, Fernanda Kroker, Neal D. Freedman, Katherine A. McGlynn, Manuel Ramirez-Zea, Eliseo Guallar, John D. Groopman. Quantitative measurement of aflatoxin-serum albumin adducts reveals substantial exposure in Guatemala, a country with high rates of liver cancer mortality in men and women [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4247. doi:10.1158/1538-7445.AM2017-4247

Liver cancer is the second leading cause of cancer death worldwide: risk factors include the viruses, HBV and HCV, and the environmental carcinogen aflatoxin B1 (AFB1). Chemoprevention strategies that activate genes controlled by the KEAP1-NRF2 pathway, such as CDDO-Im, afford complete protection against AFB1-induced hepatocarcinogenesis in rats. The opportunity to deploy these types of agents in high-risk populations would be advanced by the validation of biomarkers reflecting their efficacy. MicroRNAs (miRNAs) affecting gene expression are important in normal physiology and have been shown to be frequently dysregulated in cancer. It is our hypothesis that specific rat liver miRNAs are biomarkers that temporally track with AFB1-induced liver cancer, and are modulated in animals that receive complete protection by CDDO-Im. MiRNAs were isolated from archived liver tissue of rats (Johnson et al., CaPR, 2014) that were dosed with AFB1 (200 μg) for 28 consecutive days. Two additional groups received either vehicle only or AFB1 + CDDO-Im (30 μmol). After RNA isolation (miRCURY tissue, Exiqon), samples were profiled by RNA sequencing (Illumina). Fourteen miRNAs were selected based on dynamic range and level of detection for validation by RT-qPCR (TaqMan microRNA assay). MiRNA profiling from animals at the end of the carcinogenic 28-day dosing regimen revealed an increased total number of miRNAs due to AFB1 exposure alone (n≈500) compared to control. Those miRNAs displaying a greater than 10 fold increase in expression between control and AFB1 treatment groups were examined in detail: 541-5p, 34a-5p, 127-3p, 205, 434-3p, 429, 411-5p, 181c-3p, 200b-3p, 221-3p. RNA-seq data showed that these were all upregulated in the AFB1-treated samples. Co-treatment with CDDO-Im shifted expression levels back to control. The miRNAs 192-3p, 92a-3p, 26b-3p, and 375-3p were expressed consistently in all treatment groups and were selected for normalization. Expression levels of the 14 candidate miRNAs were then examined in liver samples obtained from rats after 7, 14 and 21 doses AFB1 and compared to levels determined at week 4 (28 doses AFB1). Quantitative analysis demonstrated an increase in expression of the panel of miRNAs due to AFB1 exposure over the dosing period, potentially indicating a role in early carcinogenesis. MicroRNA expression showed a varying trend over time in animals treated with AFB1 plus CDDO-Im. Three invariant miRNAs (miR-375-3p, 92a-3p, 192-3p) showed little change during the 28-day exposure period. In conclusion, we have identified a panel of miRNAs that are upregulated and track with AFB1 exposure. These changes are abrogated by treatment with CDDO-Im and thus reflect the protective efficacy of the intervention. Supported by T32ES007141-31A1 and CA197222. Citation Format: Merricka C. Livingstone, Bill D. Roebuck, Natalie M. Johnson, Thomas W. Kensler, John D. Groopman. Temporal trends in microRNAs during subchronic aflatoxin dosing and modulation by the chemopreventive oleane triterpenoid, CDDO-Im. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1083.

Globally, driven by increased life expectancy, the overall incidence of cancer is expected to nearly double over the next 20 years. Nonetheless, the target organs where these diagnoses will occur will still vary widely from country to country because of divergent exposures and other etiologic factors (1). One cancer that currently illustrates this geographic disparity is primary liver cancer (PLC). Currently, PLC is the third leading cause of cancer mortality worldwide with an estimated 696,000 deaths in 2008. Half of these PLC deaths of those deaths occur in the People's Republic of China. The major etiological factors associated with PLC in China, as established from prospective cohort studies, are chronic infection with hepatitis B virus (HBV) and extended exposure to high levels of aflatoxin in the diet, especially from corn and groundnuts (2). The largest study, comprising more than 18,000 men residing in Shanghai in the 1980s, examined HBV infection and aflatoxin exposure as independent and interactive risk factors for PLC (3). This nested case control study revealed a statistically significant increase in the relative risk (RR) of 7.3 for men who were chronically infected with HBV (HBsAg) but unexposed to aflatoxin. In men on whom urinary aflatoxin but not HBV biomarkers were detected, the RR was 3 (4). Furthermore, in men exhibiting both urinary aflatoxin biomarkers and positive HBsAg status, the RR was 59.4.3 A subsequent cohort study in Taiwan confirmed these results.4 In light of this synergistic interaction, programs to eliminate either factor, or both, should have substantial impact on the burden of PLC. Chronic infection with HBV has been long regarded as the major cause of PLC and initiatives have been undertaken to implement universal immunization programs. It is estimated that more than 90% of countries now routinely vaccinate newborns against HBV, and approximately 70% are now delivering 3 immunization doses. Newborns are targeted because HBV transmission is often vertical from mother to child at birth or during the perinatal period. In 1986, Taiwan became the first region to vaccinate all newborns against HBV. Since then, the number of HBV carriers in the juvenile population has declined dramatically. As of 2009, incidence of hepatocellular carcinoma was significantly lower among children aged 6-19 years in vaccinated compared with antecedent unvaccinated birth cohorts (5). For many years, we have been working with colleagues in Qidong, China, located at the mouth of the Yangtze River, an endemic area for liver cancer, where early age of onset and high mortality rates from PLC are reported (6). HBV vaccination of newborns in the rural townships was initiated during the 1980s but did not become universal until 2002. Crude rates of PLC per 100,000 residents of Qidong rose slightly from 1972 through 1990 at levels around 50/100,000 before increasing in the 1990s to a stable rate of about 75/100,000. Such data have led to the perception that Qidong remains an irrevocably high-risk region (7). However, the latter PLC mortality rate increase largely reflects a dramatically altered age distribution in the population rising from 17.0% over 50 years of age in 1973 to 22.7% in 1988 and 34.2% in 2003. Prognosis for survival from liver cancer following diagnosis remains poor: currently In order to accurately describe aflatoxin exposures over the past quarter century, frozen serum samples collected since 1989 were measured for aflatoxin-albumin adduct content (9–11). There has been a dramatic decline in levels of exposure from the 1980s to the present. Median levels of aflatoxin-albumin adducts from residents of the villages of Daxin and HeZuo declined from 19.3 pg/mg albumin in 1989, to 3.6 in 1995, to 2.3 in 1999, to 1.4 in 2003 and undetectable (i.e., Acknowledgments This work has been funded by the National Institutes of Health USA (P01 ES 006052) and the National Science and Technology Mega-Projects of China (No. 2008ZX10002-015, No. 2012ZX10002-008. Citation Format: John D. Groopman, Thomas W. Kensler. Global burden of liver cancer: Emerging needs in Asia. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr SY14-04. doi:10.1158/1538-7445.AM2013-SY14-04

Primary liver cancer (PLC) causes nearly 700,000 annual deaths worldwide predominately in Asia and sub-Saharan Africa; >370,000 deaths occur in the People's Republic of China, alone. Two factors synergistically associated with PLC are chronic infection with hepatitis B (HBV) and repetitive ingestion of aflatoxin (AFB)-contaminated foods. Utilizing information collected over several decades by the Qidong Cancer Registry, mortality rates from PLC were compared to timelines for regional implementation of HBV vaccination and levels of AFB exposure. The concentration of the AFB-lysine adduct was quantified by isotope dilution mass spectrometry in randomized, age-matched subsets of archived serum samples collected from 1989-2012. Economic, social and public health changes occurring during this time period were examined including implementation of a universal vaccination program and the adaptation of an open policy of food provisionment which led to rice replacing maize as the major dietary staple in the Qidong region. Although the crude rates of PLC increased from 1972-1990s (50/100,000 to a steady-state rate of 75/100,000), much of the increase can be attributed to a proportionate aging population. Upon stratifying by birth cohort, no significant change in mortality rate from PLC was found for individuals over 40 years of age. However, 60-75% mortality declines in 20-24, 25-29, 30-34 year olds born 1973-177 compared to similar birth cohorts of 1958-1962 were observed. The prevalence of HBV surface antigen positivity was unchanged; moreover, all these birth cohorts were born before the universal HBV vaccination of newborns. Median levels of the aflatoxin biomarker dropped from 19.3 pg/mg albumin in 1989 to non-detectable by 2009. The estimated population attributable risk of AFB-related PLC decreased from 45% in 1989 to 11% in 2012. Significant reductions in PLC mortality occurred prior to the introduction of universal hepatitis vaccination in Qidongese newborns. Agricultural and economic reforms instituted in the 1980s leading to the establishment of rice as a dietary staple was associated with a 40-fold reduction in AFB exposure. Thus, by extrapolation, changes in food-based policies altering exposure to carcinogens in the diet can have a dramatic impact on PLC mortality worldwide. Supported by NIEHS P01 ES 006052, NST China No. 2008ZX10002-015, No. 2012ZX10002-008. Citation Format: Patricia A. Egner, Jian-Guo Chen, Derek Ng, Lisa P. Jacobson, Alvaro Munoz, Jian-Hua Lu, Yuan-Rong Zhu, Geng-Sun Qian, Tao-Yang Chen, Felicia Wu, Jian-Min Wang, John D. Groopman, Thomas W. Kensler. Reduction in aflatoxin exposure drives decrease in primary liver cancer in Qidong, China. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1381. doi:10.1158/1538-7445.AM2013-1381

Modulation of estrogen depurinating DNA adduct by sulforaphane or KEAP1 knockdown in human breast epithelial cells Sulforaphane (SFN) is a potent inducer of detoxication enzymes such as NAD(P)H:quinone oxidoreductase (NQO1) and glutathione-S-transferases (GST) via the Kelch-like ECH-associated protein 1 (Keap1) - Nuclear Factor- E2-related factor (Nrf2) signaling pathway. NQO1 reduces the carcinogenic estrogen metabolite, catechol estrogen-3,4-quinone (CE-3,4-Q), while GSTs detoxify it through nucleophilic addition. CE-3,4-Q can bind with DNA to form depurinating DNA adducts. Thus, SFN, a isothiocyanate found in broccoli, may alter estrogen metabolism to protect against estrogen-mediated carcinogenesis. MCF10A cells were treated with either vehicle or SFN (10 μM) and either estradiol (E2) or 4-OHE2 (10 μM). NQO1 activity increased 3.2-fold by SFN treatment compared to vehicle. Estrogen metabolites and depurinating DNA adducts in the cell culture medium were partially purified by solid phase extraction and then analyzed by UHPLC-MS/MS. Following E2 treatment, 4-OHE1/2-1-N3Ade and 4-OHE1/2-1-N7Gua adducts were reduced by 60% in SFN treatment; 4-OHE1/2- glutathione conjugates increased 1.9 while 4-OCH3E1/2 increased 3.0 fold with SFN. Following treatment with the proximate metabolite 4-OHE2, 4-OHE1/2-1-N3Ade and 4-OHE 1/2-1-N7Gua adducts also decreased 60% in SFN treated cells compared to vehicle; while 4-OHE1/2-glutathione-conjugates increased 5.0-fold and 4-OCH3E1/2 levels were 3.4-fold higher. To constitutively enhance the expression of Nrf2-regulated genes including NQO1, cells were treated with either scrambled or siKEAP1 RNA and E2 or 4-OHE2. NQO1 activity increased 2.2-fold with siKEAP1 treatment. Following E2 treatment, 4-OHE1/2-1-N3Ade and 4-OHE1/2-1-N7Gua adducts dropped 70% in siKEAP1 treated cells compared to scrambled; 4-OHE1/2-glutathione conjugates increased 1.3-fold; however, 4-OCH3E1/2 decreased 50% with siKEAP1 treatment. Following 4-OHE2 treatment, the 4-OHE1/2-1-N3Ade and 4-OHE 1/2-1-N7Gua adducts declined 60% in siKEAP1 treated cells compared to scrambled; 4-OHE1/2-glutathione conjugates increased 1.4-fold while 4-OCH3E1/2 declined 60% with siKEAP1 treatment. SFN or siKEAP1 have similar effects on up-regulating NQO1 transcripts, protein expression and activity levels and on diminution of depurinating estrogen DNA adducts following E2 or 4-OHE2 challenge. However, these pharmacologic and genetic approaches have different effects on COMT. siKEAP1 down-regulates COMT expression, which inhibits 4-OCH3E1 but not 4-CH3E2 formation whilst SFN elevates COMT expression and subsequently levels of 4-OCH3E1/2. Thus, activation of the Nrf2 pathway may account for some but not all of the protective effects of SFN against estrogen-mediated DNA damage. Supported by DOD BCRP Postdoctoral Fellowship103928. Citation Format: Li Yang, Muhammad Zahid, Eleanor G. Rogan, Ercole L. Cavalieri, James D, Yager, Kala Visvanathan, John Groopman, Nancy E. Davidson, Thomas W. Kensler. Modulation of estrogen depurinating DNA adduct by sulforaphane or KEAP1 knockdown in human breast epithelial cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3679. doi:10.1158/1538-7445.AM2013-3679

Epidemiological evidence has suggested that consumption of a diet rich in cruciferous vegetables reduces the risk of several types of cancers and chronic degenerative diseases. In particular, broccoli sprouts are a convenient and rich source of the glucosinolate, glucoraphanin (GR) which can release the chemopreventive agent, sulforaphane (SF), an inducer of glutathione transferases. Two broccoli sprout-derived beverages, one glucoraphanin-rich (GRR) and the other sulforaphane-rich (SFR), were evaluated for SF bioavailability, tolerability and pharmacodynamic action. The GRR beverage relied on the conversion of GR to SF in the gut microflora whereas the SFR beverage was prepared by conversion of GR to SF with myrosinase from daikon sprouts prior to consumption. Study participants were recruited from the farming community of He Zuo Township, Qidong, People's Republic of China, previously documented to have a high incidence of hepatocellular carcinoma and more recently, exposures to substantive levels of air-borne pollutants including phenanthrene, acrolein, crotonaldehyde, benzene and ethylene oxide. Two hundred five individuals were screened at village clinics and 50 willing, healthy participants were randomized into two treatment arms. The study protocol was as follows: a 5-day run-in period, a 7-day administration of beverages, a 5-day washout period, and a 7-day administration of the opposite beverage. Bioavailability of SF, as indicated by excretion of GR, SF and SF-thiol conjugates in urine collected daily 12 hours after dosing, was measured by isotope dilution mass spectrometry. The urinary excretions of SF and its thiols were substantially greater in individuals receiving a SFR beverage compared to a GRR beverage. Determination of possible pharmacodynamic action of the SFR and GRR beverages has been assessed through measures of the urinary excretion of the mercapturic acids of acrolein, crotonaldehyde, ethylene oxide, benzene and 1,3-butadiene using liquid and gas chromatography coupled to mass spectrometry. Statistically significant increases in the levels of excretion of glutathione-derived conjugates of acrolein, crotonaldehyde, and benzene were seen in individuals receiving SFR, GRR or both compared to their pre-intervention baseline values. Increases provoked by SFR were greater than for GRR, but the differences were not significantly different. Thus, chemopreventive intervention may attenuate risks to health associated with exposures to air-borne pollutants in addition to possible effects on aflatoxin exposures in this region. Supported by NIEHS grant P01 ES006052. 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 LB-405. doi:10.1158/1538-7445.AM2011-LB-405