Publications by authors named "Kathleen Dickman"

33 Publications

Additive Effects of Arsenic and Aristolochic Acid in Chemical Carcinogenesis of Upper Urinary Tract Urothelium.

Cancer Epidemiol Biomarkers Prev 2021 Feb 4;30(2):317-325. Epub 2020 Dec 4.

Department of Urology, National Taiwan University Hospital, Taipei, Taiwan.

Background: Aristolochic acids (AA) and arsenic are chemical carcinogens associated with urothelial carcinogenesis. Here we investigate the combined effects of AA and arsenic toward the risk of developing upper tract urothelial carcinoma (UTUC).

Methods: Hospital-based ( = 89) and population-based (2,921 cases and 11,684 controls) Taiwanese UTUC cohorts were used to investigate the association between exposure to AA and/or arsenic and the risk of developing UTUC. In the hospital cohort, AA exposure was evaluated by measuring aristolactam-DNA adducts in the renal cortex and by identifying A>T mutations in tumors. In the population cohort, AA exposure was determined from prescription health insurance records. Arsenic levels were graded from 0 to 3 based on concentrations in well water and the presence of arseniasis-related diseases.

Results: In the hospital cohort, 43, 26, and 20 patients resided in grade 0, 1+2, and 3 arseniasis-endemic areas, respectively. Aristolactam-DNA adducts were present in >90% of these patients, indicating widespread AA exposure. A>T mutations in were detected in 28%, 44%, and 22% of patients residing in grade 0, 1+2, and 3 arseniasis-endemic areas, respectively. Population studies revealed that individuals who consumed more AA-containing herbs had a higher risk of developing UTUC in both arseniasis-endemic and nonendemic areas. Logistic regression showed an additive effect of AA and arsenic exposure on the risk of developing UTUC.

Conclusions: Exposure to both AA and arsenic acts additively to increase the UTUC risk in Taiwan.

Impact: This is the first study to investigate the combined effect of AA and arsenic exposure on UTUC.
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http://dx.doi.org/10.1158/1055-9965.EPI-20-1090DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8083126PMC
February 2021

Proximal Tubular Vacuolization and Hypersensitivity to Drug-Induced Nephrotoxicity in Male Mice With Decreased Expression of the NADPH-Cytochrome P450 Reductase.

Toxicol Sci 2020 02;173(2):362-372

Department of Pharmacology and Toxicology, College of Pharmacy, University of Arizona, Tucson, Arizona.

The effect of variations in the expression of cytochrome P450 reductase (CPR or POR) is determined in mice with decreased POR expression to identify potential vulnerabilities in people with low POR expression. There is an age-dependent appearance of increasing vacuolization in the proximal tubules of the renal cortex in 4- to 9-month-old male (but not female) Cpr-low (CL) mice. These mice have low POR expression in all cells of the body and upregulation of lysosome-associated membrane protein 1 expression in the renal cortex. Vacuolization is also seen in extrahepatic CL and extrarenal CL male mice, but not in mice with tissue-specific Por deletion in liver, intestinal epithelium, or kidney. The occurrence of vacuolization is accompanied by increases in serum blood-urea-nitrogen levels. Male CL mice are hypersensitive to cisplatin- and gentamicin-induced renal toxicity at 3 months of age, before proximal tubular (PT) vacuoles are detectable. At doses that do not cause renal toxicity in wild-type mice, both drugs cause substantial increases in serum blood-urea-nitrogen levels and PT vacuolization in male but not female CL mice. The hypersensitivity to drug-induced renal toxicity is accompanied by increases in circulating drug levels. These novel findings demonstrate deficiency of renal function in mice with globally reduced POR expression and suggest that low POR expression may be a risk factor for drug-induced nephrotoxicity in humans.
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http://dx.doi.org/10.1093/toxsci/kfz225DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8000068PMC
February 2020

Non-invasive detection of urothelial cancer through the analysis of driver gene mutations and aneuploidy.

Elife 2018 03 20;7. Epub 2018 Mar 20.

Masonic Cancer Center, University of Minnesota, Minneapolis, United States.

Current non-invasive approaches for detection of urothelial cancers are suboptimal. We developed a test to detect urothelial neoplasms using DNA recovered from cells shed into urine. UroSEEK incorporates massive parallel sequencing assays for mutations in 11 genes and copy number changes on 39 chromosome arms. In 570 patients at risk for bladder cancer (BC), UroSEEK was positive in 83% of those who developed BC. Combined with cytology, UroSEEK detected 95% of patients who developed BC. Of 56 patients with upper tract urothelial cancer, 75% tested positive by UroSEEK, including 79% of those with non-invasive tumors. UroSEEK detected genetic abnormalities in 68% of urines obtained from BC patients under surveillance who demonstrated clinical evidence of recurrence. The advantages of UroSEEK over cytology were evident in low-grade BCs; UroSEEK detected 67% of cases whereas cytology detected none. These results establish the foundation for a new non-invasive approach for detection of urothelial cancer.
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http://dx.doi.org/10.7554/eLife.32143DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5860864PMC
March 2018

Human liver-kidney model elucidates the mechanisms of aristolochic acid nephrotoxicity.

JCI Insight 2017 11 16;2(22). Epub 2017 Nov 16.

Department of Environmental and Occupational Health Sciences and.

Environmental exposures pose a significant threat to human health. However, it is often difficult to study toxicological mechanisms in human subjects due to ethical concerns. Plant-derived aristolochic acids are among the most potent nephrotoxins and carcinogens discovered to date, yet the mechanism of bioactivation in humans remains poorly understood. Microphysiological systems (organs-on-chips) provide an approach to examining the complex, species-specific toxicological effects of pharmaceutical and environmental chemicals using human cells. We microfluidically linked a kidney-on-a-chip with a liver-on-a-chip to determine the mechanisms of bioactivation and transport of aristolochic acid I (AA-I), an established nephrotoxin and human carcinogen. We demonstrate that human hepatocyte-specific metabolism of AA-I substantially increases its cytotoxicity toward human kidney proximal tubular epithelial cells, including formation of aristolactam adducts and release of kidney injury biomarkers. Hepatic biotransformation of AA-I to a nephrotoxic metabolite involves nitroreduction, followed by sulfate conjugation. Here, we identify, in a human tissue-based system, that the sulfate conjugate of the hepatic NQO1-generated aristolactam product of AA-I (AL-I-NOSO3) is the nephrotoxic form of AA-I. This conjugate can be transported out of liver via MRP membrane transporters and then actively transported into kidney tissue via one or more organic anionic membrane transporters. This integrated microphysiological system provides an ex vivo approach for investigating organ-organ interactions, whereby the metabolism of a drug or other xenobiotic by one tissue may influence its toxicity toward another, and represents an experimental approach for studying chemical toxicity related to environmental and other toxic exposures.
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http://dx.doi.org/10.1172/jci.insight.95978DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5752374PMC
November 2017

A Rapid Throughput Method To Extract DNA from Formalin-Fixed Paraffin-Embedded Tissues for Biomonitoring Carcinogenic DNA Adducts.

Chem Res Toxicol 2017 12 27;30(12):2130-2139. Epub 2017 Nov 27.

Masonic Cancer Center, Division of Carcinogenesis and Chemoprevention and Department of Medicinal Chemistry, ‡Department of Laboratory Medicine and Pathology, and §Department of Urology, University of Minnesota , Minneapolis, Minnesota 55455, United States.

Formalin-fixed paraffin-embedded (FFPE) tissues are rarely used for screening DNA adducts of carcinogens because the harsh conditions required to reverse the formaldehyde-mediated DNA cross-links can destroy DNA adducts. We recently adapted a commercial silica-based column kit used in genomics to manually isolate DNA under mild conditions from FFPE tissues of rodents and humans and successfully measured DNA adducts of several carcinogens including aristolochic acid I (AA-I), 4-aminobiphenyl (4-ABP), and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) (Yun et al. (2013) Anal. Chem. 85, 4251-8, and Guo et al. (2016) Anal. Chem. 88, 4780-7). The DNA retrieval methodology is robust; however, the procedure is time-consuming and labor intensive, and not amenable to rapid throughput processing. In this study, we have employed the Promega Maxwell 16 MDx system, which is commonly used in large scale genomics studies, for the rapid throughput extraction of DNA. This system streamlines the DNA isolation procedure and increases the sample processing rate by about 8-fold over the manual method (32 samples versus 4 samples processed per hour). High purity DNA is obtained in satisfactory yield for the measurements of DNA adducts by ultra performance liquid chromatography-electrospray-ionization-ion trap-multistage scan mass spectrometry. The measurements show that the levels of DNA adducts of AA-I, 4-ABP, and PhIP in FFPE rodent and human tissues are comparable to those levels measured in DNA from matching tissues isolated by the commercial silica-based column kits and in DNA from fresh frozen tissues isolated by the conventional phenol-chloroform extraction method. The isolation of DNA from tissues is one major bottleneck in the analysis of DNA adducts. This rapid throughput methodology greatly decreases the time required to process DNA and can be employed in large-scale epidemiology studies designed to assess the role of chemical exposures and DNA adducts in cancer risk.
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http://dx.doi.org/10.1021/acs.chemrestox.7b00218DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5822421PMC
December 2017

Aristolochic Acid in the Etiology of Renal Cell Carcinoma.

Cancer Epidemiol Biomarkers Prev 2016 12 23;25(12):1600-1608. Epub 2016 Aug 23.

Department of Pharmacological Sciences, Stony Brook University, Stony Brook, New York.

Background: Aristolochia species used in the practice of traditional herbal medicine contains aristolochic acid (AA), an established human carcinogen contributing to urothelial carcinomas of the upper urinary tract. AA binds covalently to genomic DNA, forming aristolactam (AL)-DNA adducts. Here we investigated whether AA is also an etiologic factor in clear cell renal cell carcinoma (ccRCC).

Methods: We conducted a population-based case-control study to investigate the linkage between Aristolochia prescription history, cumulative AA consumption, and ccRCC incidence in Taiwan (5,709 cases and 22,836 matched controls). The presence and level of mutagenic dA-AL-I adducts were determined in the kidney DNA of 51 Taiwanese ccRCC patients. The whole-exome sequences of ccRCC tumors from 10 Taiwanese ccRCC patients with prior exposure to AA were determined.

Results: Cumulative ingestion of more than 250 mg of AA increased risk of ccRCC (OR, 1.25), and we detected dA-AL-I adducts in 76% of Taiwanese ccRCC patients. Furthermore, the distinctive AA mutational signature was evident in six of 10 sequenced ccRCC exomes from Taiwanese patients.

Conclusions: This study strongly suggests that AA contributes to the etiology of certain RCCs.

Impact: The current study offers compelling evidence implicating AA in a significant fraction of the RCC arising in Taiwan and illustrates the power of integrating epidemiologic, molecular, and genetic data in the investigation of cancer etiology. Cancer Epidemiol Biomarkers Prev; 25(12); 1600-8. ©2016 AACR.
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http://dx.doi.org/10.1158/1055-9965.EPI-16-0219DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5533284PMC
December 2016

Recurrence pattern and TP53 mutation in upper urinary tract urothelial carcinoma.

Oncotarget 2016 Jul;7(29):45225-45236

Department of Urology, National Taiwan University Hospital, Taipei, Taiwan.

TP53 mutation patterns are associated with prognosis of various cancers. This study was designed to investigate the association between TP53 mutation patterns and recurrence patterns in upper urinary tract urothelial carcinoma (UTUC) patients. A total of 165 consecutive UTUC patients who underwent nephroureterectomies were enrolled for measuring mutation patterns of TP53 gene from exome 2 to 11. Bladder recurrence, contralateral UTUC recurrence, and metastases were compared among groups by using log-rank test and Cox proportional hazard model. Single base substitution as an A:T to T:A transversion was noted in 55 (33.3%) patients (AT group). Forty-two (25.5%) patients had TP53 mutations with only other than A:T to T:A transversion (NAT group), and 68 patients (41.2%) had wide-type TP53 (WT group). AT group was predominately female (64%, 52%, 29%, respectively), had a higher incidence of end-stage renal disease (24%, 14%, 10%, respectively), and had more high-grade tumors (82%, 74%, 62%, respectively) compared to NAT and WT groups. With adjustment of tumor grade/stages, bladder and contralateral UTUC recurrence-free survival duration was shortest in NAT (p < 0.001) and AT group (p < 0.001), respectively. NAT group had a shorter metastasis-free survival duration than the other two groups combined (p = 0.018). As a result, A:T to T:A transversion increased contralateral UTUC recurrence risk, but other mutations in TP53 raised the hazard of bladder recurrence and metastases. Therefore, TP53 mutation pattern may be a useful biomarker to predict recurrence patterns of UTUC patients.
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http://dx.doi.org/10.18632/oncotarget.9904DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5216718PMC
July 2016

Low-Coverage Exome Sequencing Screen in Formalin-Fixed Paraffin-Embedded Tumors Reveals Evidence of Exposure to Carcinogenic Aristolochic Acid.

Cancer Epidemiol Biomarkers Prev 2015 Dec 17;24(12):1873-81. Epub 2015 Sep 17.

Molecular Mechanisms and Biomarkers Group, International Agency for Research on Cancer, Lyon, France.

Background: Dietary exposure to cytotoxic and carcinogenic aristolochic acid (AA) causes severe nephropathy typically associated with urologic cancers. Monitoring of AA exposure uses biomarkers such as aristolactam-DNA adducts, detected by mass spectrometry in the kidney cortex, or the somatic A>T transversion pattern characteristic of exposure to AA, as revealed by previous DNA-sequencing studies using fresh-frozen tumors.

Methods: Here, we report a low-coverage whole-exome sequencing method (LC-WES) optimized for multisample detection of the AA mutational signature, and demonstrate its utility in 17 formalin-fixed paraffin-embedded urothelial tumors obtained from 15 patients with endemic nephropathy, an environmental form of AA nephropathy.

Results: LC-WES identified the AA signature, alongside signatures of age and APOBEC enzyme activity, in 15 samples sequenced at the average per-base coverage of approximately 10×. Analysis at 3 to 9× coverage revealed the signature in 91% of the positive samples. The exome-wide distribution of the predominant A>T transversions exhibited a stochastic pattern, whereas 83 cancer driver genes were enriched for recurrent nonsynonymous A>T mutations. In two patients, pairs of tumors from different parts of the urinary tract, including the bladder, harbored overlapping mutation patterns, suggesting tumor dissemination via cell seeding.

Conclusions: LC-WES analysis of archived tumor tissues is a reliable method applicable to investigations of both the exposure to AA and its biologic effects in human carcinomas.

Impact: By detecting cancers associated with AA exposure in high-risk populations, LC-WES can support future molecular epidemiology studies and provide evidence-base for relevant preventive measures.
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http://dx.doi.org/10.1158/1055-9965.EPI-15-0553DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4806408PMC
December 2015

New Approaches for Biomonitoring Exposure to the Human Carcinogen Aristolochic Acid.

Toxicol Res (Camb) 2015 Jul;4(4):763-776

Masonic Cancer Center and Department of Medicinal Chemistry, University of Minnesota, Minneapolis, MN 55455, USA.

Aristolochic acids (AA) are found in all herbaceous plants, many of which have been used worldwide for medicinal purposes for centuries. AA are causal agents of the chronic kidney disease entity termed aristolochic acid nephropathy (AAN) and potent upper urinary tract carcinogens in humans. AAN and upper urinary tract cancers are endemic in rural areas of Croatia and other Balkan countries where exposure to AA occurs through the ingestion of home-baked bread contaminated with seeds. In Asia, exposure to AA occurs through usage of traditional Chinese medicinal herbs containing . Despite warnings from regulatory agencies, traditional Chinese herbs containing AA continue to be used world-wide. In this review, we highlight novel approaches to quantify exposure to AA, by analysis of aristolactam (AL) DNA adducts, employing ultraperformance liquid chromatography-electrospray ionization/multistage mass spectrometry (UPLC-ESI/MS). DNA adducts are a measure of internal exposure to AA and serve as an important end point for cross-species extrapolation of toxicity data and human risk assessment. The level of sensitivity of UPLC-ESI/MS surpasses the limits of detection of AL-DNA adducts obtained by P-postlabeling techniques, the most widely employed methods for detecting putative DNA adducts in humans. AL-DNA adducts can be measured by UPLC-ESI/MS, not only in fresh frozen renal tissue, but also in formalin-fixed, paraffin-embedded (FFPE) samples, an underutilized biospecimen for assessing chemical exposures, and in exfoliated urinary cells, a non-invasive approach. The frequent detection of AL DNA adducts in renal tissues, combined with the characteristic mutational spectrum induced by AA in and other genes provides compelling data for a role of AA in upper urothelial tract cancer.
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http://dx.doi.org/10.1039/C5TX00052ADOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4564010PMC
July 2015

Krüppel-like factor 6 regulates mitochondrial function in the kidney.

J Clin Invest 2015 Mar 17;125(3):1347-61. Epub 2015 Feb 17.

Maintenance of mitochondrial structure and function is critical for preventing podocyte apoptosis and eventual glomerulosclerosis in the kidney; however, the transcription factors that regulate mitochondrial function in podocyte injury remain to be identified. Here, we identified Krüppel-like factor 6 (KLF6), a zinc finger domain transcription factor, as an essential regulator of mitochondrial function in podocyte apoptosis. We observed that podocyte-specific deletion of Klf6 increased the susceptibility of a resistant mouse strain to adriamycin-induced (ADR-induced) focal segmental glomerulosclerosis (FSGS). KLF6 expression was induced early in response to ADR in mice and cultured human podocytes, and prevented mitochondrial dysfunction and activation of intrinsic apoptotic pathways in these podocytes. Promoter analysis and chromatin immunoprecipitation studies revealed that putative KLF6 transcriptional binding sites are present in the promoter of the mitochondrial cytochrome c oxidase assembly gene (SCO2), which is critical for preventing cytochrome c release and activation of the intrinsic apoptotic pathway. Additionally, KLF6 expression was reduced in podocytes from HIV-1 transgenic mice as well as in renal biopsies from patients with HIV-associated nephropathy (HIVAN) and FSGS. Together, these findings indicate that KLF6-dependent regulation of the cytochrome c oxidase assembly gene is critical for maintaining mitochondrial function and preventing podocyte apoptosis.
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http://dx.doi.org/10.1172/JCI77084DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4362257PMC
March 2015

Chronic dietary exposure to aristolochic acid and kidney function in native farmers from a Croatian endemic area and Bosnian immigrants.

Clin J Am Soc Nephrol 2015 Feb 13;10(2):215-23. Epub 2015 Jan 13.

Department of Pharmacological Sciences, State University of New York at Stony Brook, Stony Brook, New York.

Background And Objectives: Improvements in agricultural practices in Croatia have reduced exposure to consumption of aristolochic acid-contaminated flour and development of endemic (Balkan) nephropathy. Therefore, it was hypothesized that Bosnian immigrants who settled in an endemic area in Croatia 15-30 years ago would be at lower risk of developing endemic nephropathy because of reduced exposure to aristolochic acid. To test this hypothesis, past and present exposure to aristolochic acid, proximal tubule damage as a hallmark of endemic nephropathy, and prevalence of CKD in Bosnian immigrants were analyzed.

Design, Setting, Participants, & Measurements: In this cross-sectional observational study from 2005 to 2010, 2161 farmers were divided into groups: indigenous inhabitants from endemic nephropathy and nonendemic nephropathy villages and Bosnian immigrants; α-1 microglobulin-to-creatinine ratio >31.5 mg/g and eGFR<60 ml/min per 1.73 m(2) were considered to be abnormal.

Results: CKD and proximal tubule damage prevalence was significantly lower in Bosnian immigrants than inhabitants of endemic nephropathy villages (6.9% versus 16.6%; P<0.001; 1.3% versus 7.3%; P=0.003, respectively); 20 years ago, Bosnian immigrants observed fewer Aristolochia clematitis in cultivated fields (41.9% versus 67.8%) and fewer seeds among wheat seeds (6.1% versus 35.6%) and ate more purchased than homemade bread compared with Croatian farmers from endemic nephropathy villages (38.5% versus 14.8%, P<0.001). Both Croatian farmers and Bosnian immigrants observe significantly fewer Aristolochia plants growing in their fields compared with 15-30 years ago. Prior aristolochic acid exposure was associated with proximal tubule damage (odds ratio, 1.64; 95% confidence interval, 1.04 to 2.58; P=0.02), whereas present exposure was not (odds ratio, 1.31; 95% confidence interval, 0.75 to 2.30; P=0.33). Furthermore, immigrant status was an independent negative predictor of proximal tubule damage (odds ratio, 0.40; 95% confidence interval, 0.19 to 0.86; P=0.02).

Conclusions: Bosnian immigrants and autochthonous Croats residing in endemic areas are exposed significantly less to ingestion of aristolochic acid than in the past. The prevalence of endemic nephropathy and its associated urothelial cancers is predicted to decrease over time.
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http://dx.doi.org/10.2215/CJN.03190314DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4317733PMC
February 2015

Formalin-fixed paraffin-embedded tissue as a source for quantitation of carcinogen DNA adducts: aristolochic acid as a prototype carcinogen.

Carcinogenesis 2014 Sep 28;35(9):2055-61. Epub 2014 Apr 28.

Division of Environmental Health Sciences, Wadsworth Center, New York State Department of Health, Albany, NY 12201, USA; Masonic Cancer Center and Department of Medicinal Chemistry, University of Minnesota, Minneapolis, MN 55455, USA,

DNA adducts are a measure of internal exposure to genotoxicants. However, the measurement of DNA adducts in molecular epidemiology studies often is precluded by the lack of fresh tissue. In contrast, formalin-fixed paraffin-embedded (FFPE) tissues frequently are accessible, although technical challenges remain in retrieval of high quality DNA suitable for biomonitoring of adducts. Aristolochic acids (AA) are human carcinogens found in Aristolochia plants, some of which have been used in the preparation of traditional Chinese herbal medicines. We previously established a method to measure DNA adducts of AA in FFPE tissue. In this study, we examine additional features of formalin fixation that could impact the quantity and quality of DNA and report on the recovery of AA-DNA adducts in mice exposed to AA. The yield of DNA isolated from tissues fixed with formalin decreased over 1 week; however, the levels of AA-DNA adducts were similar to those in fresh frozen tissue. Moreover, DNA from FFPE tissue served as a template for PCR amplification, yielding sequence data of comparable quality to DNA obtained from fresh frozen tissue. The estimates of AA-DNA adducts measured in freshly frozen tissue and matching FFPE tissue blocks of human kidney stored for 9 years showed good concordance. Thus, DNA isolated from FFPE tissues may be used to biomonitor DNA adducts and to amplify genes used for mutational analysis, providing clues regarding the origin of human cancers for which an environmental cause is suspected.
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http://dx.doi.org/10.1093/carcin/bgu101DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4146416PMC
September 2014

Bioactivation of the human carcinogen aristolochic acid.

Carcinogenesis 2014 Aug 17;35(8):1814-22. Epub 2014 Apr 17.

Department of Pharmacological Sciences, Department of Medicine and.

Unlabelled: Aristolochic acids are potent human carcinogens; the role of phase II metabolism in their bioactivation is unclear. Accordingly, we tested the ability of the partially reduced metabolites, N-hydroxyaristolactams (AL-NOHs), and their N-O-sulfonated and N-O-acetylated derivatives to react with DNA to form aristolactam-DNA adducts. AL-NOHs displayed little or no activity in this regard while the sulfo- and acetyl compounds readily form DNA adducts, as detected by (32)P-post-labeling analysis. Mouse hepatic and renal cytosols stimulated binding of AL-NOHs to DNA in the presence of adenosine 3'-phosphate 5'-phosphosulfate (PAPS) but not of acetyl-CoA. Using Time of Flight liquid chromatography/mass spectrometry, N-hydroxyaristolactam I formed the sulfated compound in the presence of PAPS and certain human sulfotransferases, SULT1B1 > SULT1A2 > SULT1A1 > SULT1A3. The same pattern of SULT reactivity was observed when N-hydroxyaristolactam I was incubated with these enzymes and PAPS and the reaction was monitored by formation of aristolactam-DNA adducts. In the presence of human

Nad(p)h: quinone oxidoreductase, the ability of aristolochic acid I to bind DNA covalently was increased significantly by addition of PAPS and SULT1B1. We conclude from these studies that AL-NOHs, formed following partial nitroreduction of aristolochic acids, serve as substrates for SULT1B1, producing N-sulfated esters, which, in turn, are converted to highly active species that react with DNA and, potentially, cellular proteins, resulting in the genotoxicity and nephrotoxicity associated with ingestion of aristolochic acids by humans.
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http://dx.doi.org/10.1093/carcin/bgu095DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4123648PMC
August 2014

Adipose tissue insulin resistance due to loss of PI3K p110α leads to decreased energy expenditure and obesity.

Am J Physiol Endocrinol Metab 2014 May 1;306(10):E1205-16. Epub 2014 Apr 1.

Department of Physiology and Biophysics, Stony Brook University, Stony Brook, New York; Department of Veterans Affairs Medical Center, Northport, New York

Adipose tissue is a highly insulin-responsive organ that contributes to metabolic regulation. Insulin resistance in the adipose tissue affects systemic lipid and glucose homeostasis. Phosphoinositide 3-kinase (PI3K) mediates downstream insulin signaling in adipose tissue, but its physiological role in vivo remains unclear. Using Cre recombinase driven by the aP2 promoter, we created mice that lack the class 1A PI3K catalytic subunit p110α or p110β specifically in the white and brown adipose tissue. The loss of p110α, not p110β, resulted in increased adiposity, glucose intolerance and liver steatosis. Mice lacking p110α in adipose tissue exhibited a decrease in energy expenditure but no change in food intake or activity compared with control animals. This low energy expenditure is a consequence of low cellular respiration in the brown adipocytes caused by a decrease in expression of key mitochondrial genes including uncoupling protein-1. These results illustrate a critical role of p110α in the regulation of energy expenditure through modulation of cellular respiration in the brown adipose tissue and suggest that compromised insulin signaling in adipose tissue might be involved in the onset of obesity.
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http://dx.doi.org/10.1152/ajpendo.00625.2013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4025064PMC
May 2014

Mutational signature of aristolochic acid exposure as revealed by whole-exome sequencing.

Sci Transl Med 2013 Aug;5(197):197ra102

Ludwig Center for Cancer Genetics and Therapeutics and the Howard Hughes Medical Institute at Johns Hopkins Kimmel Cancer Center, Baltimore, MD 21231, USA.

In humans, exposure to aristolochic acid (AA) is associated with urothelial carcinoma of the upper urinary tract (UTUC). Exome sequencing of UTUCs from 19 individuals with documented exposure to AA revealed a remarkably large number of somatic mutations and an unusual mutational signature attributable to AA. Most of the mutations (72%) in these tumors were A:T-to-T:A transversions, located predominantly on the nontranscribed strand, with a strong preference for deoxyadenosine in a consensus sequence (T/CAG). This trinucleotide motif overlaps the canonical splice acceptor site, possibly accounting for the excess of splice site mutations observed in these tumors. The AA mutational fingerprint was found frequently in oncogenes and tumor suppressor genes in AA-associated UTUC. The AA mutational signature was observed in one patient's tumor from a UTUC cohort without previous indication of AA exposure. Together, these results directly link an established environmental mutagen to cancer through genome-wide sequencing and highlight its power to reveal individual exposure to carcinogens.
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http://dx.doi.org/10.1126/scitranslmed.3006200DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3973132PMC
August 2013

Human formalin-fixed paraffin-embedded tissues: an untapped specimen for biomonitoring of carcinogen DNA adducts by mass spectrometry.

Anal Chem 2013 May 10;85(9):4251-8. Epub 2013 Apr 10.

Division of Environmental Health Sciences, Wadsworth Center, New York State Department of Health, Albany, New York 12201, United States.

DNA adducts represent internal dosimeters to measure exposure to environmental and endogenous genotoxicants. Unfortunately, in molecular epidemiologic studies, measurements of DNA adducts often are precluded by the unavailability of fresh tissue. In contrast, formalin-fixed paraffin embedded (FFPE) tissues frequently are accessible for biomarker discovery. We report here that DNA adducts of aristolochic acids (AAs) can be measured in FFPE tissues at a level of sensitivity comparable to freshly frozen tissue. AAs are nephrotoxic and carcinogenic compounds found in Aristolochia herbaceous plants, many of which have been used worldwide for medicinal purposes. AAs are implicated in the etiology of aristolochic acid nephropathy and upper urinary tract carcinoma. 8-Methoxy-6-nitrophenanthro-[3,4-d]-1,3-dioxole-5-carboxylic acid (AA-I) is a component of Aristolochia herbs and a potent human urothelial carcinogen. AA-I reacts with DNA to form the aristolactam (AL-I)-DNA adduct 7-(deoxyadenosin-N(6)-yl) aristolactam I (dA-AL-I). We established a method to quantitatively retrieve dA-AL-I from FFPE tissue. Adducts were measured, using ultraperformance liquid chromatography/mass spectrometry, in liver and kidney tissues of mice exposed to AA-I, at doses ranging from 0.001 to 1 mg/kg body weight. dA-AL-I was then measured in 10-μm thick tissue-sections of FFPE kidney from patients with upper urinary tract cancers; the values were comparable to those observed in fresh frozen samples. The limit of quantification of dA-AL-I was 3 adducts per 10(9) DNA bases per 2.5 μg of DNA. The ability to retrospectively analyze FFPE tissues for DNA adducts may provide clues to the origin of human cancers for which an environmental cause is suspected.
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http://dx.doi.org/10.1021/ac400612xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3904361PMC
May 2013

Variation in presentation and presence of DNA adducts and p53 mutations in patients with endemic nephropathy--an environmental form of the aristolochic acid nephropathy.

Kidney Blood Press Res 2013 26;37(1):1-8. Epub 2013 Feb 26.

School of Medicine, University of Zagreb, Department of Nephrology, Hypertension, Dialysis and Transplantation, University Hospital Center Zagreb, Zagreb, Croatia.

Background: Endemic nephropathy (EN) and associated urothelial cell cancers (UUC) are an environmental form of aristolochic acid nephropathy where the most probable rout of ingestion of aristolochic acid (AA) was made by bread contaminated with AA, leading to chronic dietary intoxication. Clinical courses of three members of the same family, similarly exposed to toxin, who exhibited different clinical courses of the disease are presented.

Methods: Questionnaires on AA exposure were taken. Tissue samples were obtained during therapeutic nephrouretectomies. Histopathology, immunohistochemical detection of p53, p53 mutation screening in tumor DNA and analysis on the presence of aristolactam (AL)-DNA adducts were performed.

Results: Case 1 had UUC with typical EN histopathological signs, whereas Case 2 had bilateral UUCs with typical EN histopathological signs. In contrast, the patient in Case 3 initially showed renal insufficiency, complicated afterwards by right UUC, and later on by left UUC with histopathological end-stage chronic changes but without typical EN changes. AA-DNA adducts and specific p53 mutational spectra (A:T→ T:A transversion) were found in tissues of cases 1 and 2.

Conclusion: Diverse clinical courses seem to be related not to differences in exposure but to differences in metabolic activation or detoxification of AA and/or DNA repair resulting from different genetic polymorphisms.
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http://dx.doi.org/10.1159/000343394DOI Listing
January 2014

Aristolochic acid-induced upper tract urothelial carcinoma in Taiwan: clinical characteristics and outcomes.

Int J Cancer 2013 Jul 12;133(1):14-20. Epub 2013 Feb 12.

Department of Urology, National Taiwan University Hospital, Taipei, Taiwan.

Aristolochic acid (AA), a component of all Aristolochia-based herbal medicines, is a potent nephrotoxin and human carcinogen associated with upper urinary tract urothelial carcinoma (UUC). To investigate the clinical and pathological characteristics of AA-induced UUC, this study included 152 UUC patients, 93 of whom had been exposed to AA based on the presence of aristolactam-DNA adducts in the renal cortex. Gene sequencing was used to identify tumors with A:T-to-T:A transversions in TP53, a mutational signature associated with AA. Cases with both aristolactam-DNA adducts and A:T-to-T:A transversions in TP53 were defined as AA-UUC, whereas patients lacking both of these biomarkers were classified as non-AA-UUC. Cases with either biomarker were classified as possible-AA-UUC. Forty (26%), 60 (40%), and 52 (34%) patients were classified as AA-UUC, possible-AA-UUC and non-AA-UUC, respectively. AA-UUC patients were younger (median ages: 64, 68, 68 years, respectively; p=0.189), predominately female (65%, 42%, 35%, respectively; p=0.011), had more end-stage renal disease (28%, 10%, 12%, respectively; p=0.055), and were infrequent smokers (5%, 22%, 33%, respectively; p=0.07) compared to possible-AA-UUC and non-AA-UUC patients. All 14 patients who developed contralateral UUC had aristolactam-DNA adducts; ten of these also had signature mutations. The contralateral UUC-free survival period was shorter in AA-UUC compared to possible- or non-AA-UUC (p=0.019 and 0.002, respectively), whereas no differences among groups were observed for bladder cancer recurrence. In conclusion, AA-UUC patients tend to be younger and female, and have more advanced renal disease. Notably, AA exposure was associated with an increased risk for developing synchronous bilateral and metachronous contralateral UUC.
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http://dx.doi.org/10.1002/ijc.28013DOI Listing
July 2013

Biomonitoring of aristolactam-DNA adducts in human tissues using ultra-performance liquid chromatography/ion-trap mass spectrometry.

Chem Res Toxicol 2012 May 4;25(5):1119-31. Epub 2012 May 4.

Division of Environmental Health Sciences, Wadsworth Center, New York State Department of Health, Empire State Plaza, Albany, NY 12201, United States.

Aristolochic acids (AAs) are a structurally related family of nephrotoxic and carcinogenic nitrophenanthrene compounds found in Aristolochia herbaceous plants, many of which have been used worldwide for medicinal purposes. AAs have been implicated in the etiology of so-called Chinese herbs nephropathy and of Balkan endemic nephropathy. Both of these disease syndromes are associated with carcinomas of the upper urinary tract (UUC). 8-Methoxy-6-nitrophenanthro-[3,4-d]-1,3-dioxolo-5-carboxylic acid (AA-I) is a principal component of Aristolochia herbs. Following metabolic activation, AA-I reacts with DNA to form aristolactam (AL-I)-DNA adducts. We have developed a sensitive analytical method, using ultraperformance liquid chromatography-electrospray ionization/multistage mass spectrometry (UPLC-ESI/MS(n)) with a linear quadrupole ion-trap mass spectrometer, to measure 7-(deoxyadenosin-N(6)-yl) aristolactam I (dA-AL-I) and 7-(deoxyguanosin-N(2)-yl) aristolactam I (dG-AL-I) adducts. Using 10 μg of DNA for measurements, the lower limits of quantitation of dA-AL-I and dG-AL-I are, respectively, 0.3 and 1.0 adducts per 10(8) DNA bases. We have used UPLC-ESI/MS(n) to quantify AL-DNA adducts in tissues of rodents exposed to AA and in the renal cortex of patients with UUC who reside in Taiwan, where the incidence of this uncommon cancer is the highest reported for any country in the world. In human tissues, dA-AL-I was detected at levels ranging from 9 to 338 adducts per 10(8) DNA bases, whereas dG-AL-I was not found. We conclude that UPLC-ESI/MS(n) is a highly sensitive, specific and robust analytical method, positioned to supplant (32)P-postlabeling techniques currently used for biomonitoring of DNA adducts in human tissues. Importantly, UPLC-ESI/MS(n) could be used to document exposure to AA, the toxicant responsible for AA nephropathy and its associated UUC.
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http://dx.doi.org/10.1021/tx3000889DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3536064PMC
May 2012

Aristolochic acid-associated urothelial cancer in Taiwan.

Proc Natl Acad Sci U S A 2012 May 9;109(21):8241-6. Epub 2012 Apr 9.

Department of Urology, National Taiwan University Hospital, Taipei, Taiwan 10002.

Aristolochic acid, a potent human carcinogen produced by Aristolochia plants, is associated with urothelial carcinoma of the upper urinary tract (UUC). Following metabolic activation, aristolochic acid reacts with DNA to form aristolactam (AL)-DNA adducts. These lesions concentrate in the renal cortex, where they serve as a sensitive and specific biomarker of exposure, and are found also in the urothelium, where they give rise to a unique mutational signature in the TP53 tumor-suppressor gene. Using AL-DNA adducts and TP53 mutation spectra as biomarkers, we conducted a molecular epidemiologic study of UUC in Taiwan, where the incidence of UUC is the highest reported anywhere in the world and where Aristolochia herbal remedies have been used extensively for many years. Our study involves 151 UUC patients, with 25 patients with renal cell carcinomas serving as a control group. The TP53 mutational signature in patients with UUC, dominated by otherwise rare A:T to T:A transversions, is identical to that observed in UUC associated with Balkan endemic nephropathy, an environmental disease. Prominent TP53 mutational hotspots include the adenine bases of (5')AG (acceptor) splice sites located almost exclusively on the nontranscribed strand. A:T to T:A mutations also were detected at activating positions in the FGFR3 and HRAS oncogenes. AL-DNA adducts were present in the renal cortex of 83% of patients with A:T to T:A mutations in TP53, FGFR3, or HRAS. We conclude that exposure to aristolochic acid contributes significantly to the incidence of UUC in Taiwan, a finding with significant implications for global public health.
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http://dx.doi.org/10.1073/pnas.1119920109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3361449PMC
May 2012

Behavior changes in patients with diabetes and hypertension after experiencing shared medical appointments.

J Am Acad Nurse Pract 2012 Jan 5;24(1):43-51. Epub 2011 Aug 5.

School of Nursing, George Mason University, Manassas, Virginia, USA.

Purpose: This project examined recently implemented shared medical appointments (SMAs) at a free clinic for patients with diabetes and/or hypertension. Changes in patients' self-managing behaviors, specifically exercise and goal-setting activity, were explored after participating in SMAs for 4 months.

Data Sources: The study employed a pretest-posttest quasi-experimental design. Participants completed a questionnaire of their self-managing behaviors and a behavioral action plan at each SMA. The SMAs were facilitated in English, Spanish, and bilingually (English and Spanish) with a total of 37 participants.

Conclusions: Descriptive analysis showed a significant increase in exercise time with a mean increase of 86 min per week at post-SMA (p= .002, 95% confidence interval [CI]). Each participant identified a measurable goal, and 97% of participants reported achieving or almost achieving their goals. Males reported a significantly (p= .002, 95% CI) larger increase in exercise time than women. Variance of self-managing behaviors among the English, Spanish, and bilingual SMAs was statistically not significant.

Implications For Practice: Though much evidence exists demonstrating that SMAs provide effective quality care, literature is lacking in examining patients' self-managing behaviors after participation in language-specific SMAs. Understanding patients' response to programs that address the needs of the individual leads to more effective programs.
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http://dx.doi.org/10.1111/j.1745-7599.2011.00660.xDOI Listing
January 2012

Aristolochic acid I metabolism in the isolated perfused rat kidney.

Chem Res Toxicol 2012 Jan 14;25(1):130-9. Epub 2011 Dec 14.

Department of Biological Sciences, Florida International University , Miami, Florida 33199, United States.

Aristolochic acids are natural nitro-compounds found globally in the plant genus Aristolochia that have been implicated in the severe illness in humans termed aristolochic acid nephropathy (AAN). Aristolochic acids undergo nitroreduction, among other metabolic reactions, and active intermediates arise that are carcinogenic. Previous experiments with rats showed that aristolochic acid I (AA-I), after oral administration or injection, is subjected to detoxication reactions to give aristolochic acid Ia, aristolactam Ia, aristolactam I, and their glucuronide and sulfate conjugates that can be found in urine and feces. Results obtained with whole rats do not clearly define the role of liver and kidney in such metabolic transformation. In this study, in order to determine the specific role of the kidney on the renal disposition of AA-I and to study the biotransformations suffered by AA-I in this organ, isolated kidneys of rats were perfused with AA-I. AA-I and metabolite concentrations were determined in perfusates and urine using HPLC procedures. The isolated perfused rat kidney model showed that AA-I distributes rapidly and extensively in kidney tissues by uptake from the peritubular capillaries and the tubules. It was also established that the kidney is able to metabolize AA-I into aristolochic acid Ia, aristolochic acid Ia O-sulfate, aristolactam Ia, aristolactam I, and aristolactam Ia O-glucuronide. Rapid demethylation and sulfation of AA-I in the kidney generate aristolochic acid Ia and its sulfate conjugate that are voided to the urine. Reduction reactions to give the aristolactam metabolites occur to a slower rate. Renal clearances showed that filtered AA-I is reabsorbed at the tubules, whereas the metabolites are secreted. The unconjugated metabolites produced in the renal tissues are transported to both urine and perfusate, whereas the conjugated metabolites are almost exclusively secreted to the urine.
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http://dx.doi.org/10.1021/tx200333gDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3276251PMC
January 2012

Aristolactam-DNA adducts are a biomarker of environmental exposure to aristolochic acid.

Kidney Int 2012 Mar 9;81(6):559-67. Epub 2011 Nov 9.

School of Medicine, University of Zagreb and Department for Nephrology, Arterial Hypertension and Dialysis, University Hospital Center Zagreb, Zagreb, Croatia.

Endemic (Balkan) nephropathy is a chronic tubulointerstitial disease frequently accompanied by urothelial cell carcinomas of the upper urinary tract. This disorder has recently been linked to exposure to aristolochic acid, a powerful nephrotoxin and human carcinogen. Following metabolic activation, aristolochic acid reacts with genomic DNA to form aristolactam-DNA adducts that generate a unique TP53 mutational spectrum in the urothelium. The aristolactam-DNA adducts are concentrated in the renal cortex, thus serving as biomarkers of internal exposure to aristolochic acid. Here, we present molecular epidemiologic evidence relating carcinomas of the upper urinary tract to dietary exposure to aristolochic acid. DNA was extracted from the renal cortex and urothelial tumor tissue of 67 patients that underwent nephroureterectomy for carcinomas of the upper urinary tract and resided in regions of known endemic nephropathy. Ten patients from nonendemic regions with carcinomas of the upper urinary tract served as controls. Aristolactam-DNA adducts were quantified by (32)P-postlabeling, the adduct was confirmed by mass spectrometry, and TP53 mutations in tumor tissues were identified by chip sequencing. Adducts were present in 70% of the endemic cohort and in 94% of patients with specific A:T to T:A mutations in TP53. In contrast, neither aristolactam-DNA adducts nor specific mutations were detected in tissues of patients residing in nonendemic regions. Thus, in genetically susceptible individuals, dietary exposure to aristolochic acid is causally related to endemic nephropathy and carcinomas of the upper urinary tract.
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http://dx.doi.org/10.1038/ki.2011.371DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3560912PMC
March 2012

17β-estradiol protects the lung against acute injury: possible mediation by vasoactive intestinal polypeptide.

Endocrinology 2011 Dec 18;152(12):4729-37. Epub 2011 Oct 18.

Department of Medicine, State University of New York, Stony Brook, New York 11794-8172, USA.

Beyond their classical role as a class of female sex hormones, estrogens (e.g. 17β-estradiol) exert important biological actions, both protective and undesirable. We have investigated the ability of estradiol to protect the lung in three models of acute injury induced by 1) oxidant stress due to the herbicide paraquat; 2) excitotoxicity, caused by glutamate agonist N-methyl-d-aspartate; and 3) acute alveolar anoxia. We also assessed the role of estrogen receptors (ER) ERα and ERβ and the neuropeptide vasoactive intestinal peptide (VIP) in mediating this protection. Isolated guinea pig or rat lungs were perfused in situ at constant flow and mechanically ventilated. The onset and severity of lung injury were monitored by increases in pulmonary arterial and airway pressures, wet/dry lung weight ratio, and bronchoalveolar lavage fluid protein content. Estradiol was infused into the pulmonary circulation, beginning 10 min before induction of injury and continued for 60-90 min. Lung injury was marked by significant increases in the above measurements, with paraquat producing the most severe, and excitotoxicity the least severe, injury. Estradiol significantly attenuated the injury in each model. Both ER were constitutively expressed and immunohistochemically demonstrable in normal lung, and their selective agonists reduced anoxic injury, the only model in which they were tested. As it protected against injury, estradiol rapidly and significantly stimulated VIP mRNA expression in rat lung. Estradiol attenuated acute lung injury in three experimental models while stimulating VIP gene expression, a known mechanism of lung protection. The up-regulated VIP expression could have partially mediated the protection by estrogen.
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http://dx.doi.org/10.1210/en.2011-1631DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3230060PMC
December 2011

Physiological and molecular characterization of aristolochic acid transport by the kidney.

J Pharmacol Exp Ther 2011 Aug 5;338(2):588-97. Epub 2011 May 5.

Department of Pharmacological Sciences, Stony Brook University, BST-8, 152, Stony Brook, NY 11794, USA.

Consumption of herbal medicines derived from Aristolochia plants is associated with a progressive tubulointerstitial disease known as aristolochic acid (AA) nephropathy. The nephrotoxin produced naturally by these plants is AA-I, a nitrophenanthrene carboxylic acid that selectively targets the proximal tubule. This nephron segment is prone to toxic injury because of its role in secretory elimination of drugs and other xenobiotics. Here, we characterize the handling of AA-I by membrane transporters involved in renal organic anion transport. Uptake assays in heterologous expression systems identified murine organic anion transporters (mOat1, mOat2, and mOat3) as capable of mediating transport of AA-I. Kinetic analyses showed that all three transporters have an affinity for AA-I in the submicromolar range and thus are likely to operate at toxicologically relevant concentrations in vivo. Structure-activity relationships revealed that the carboxyl group is critical for high-affinity interaction of AA-I with mOat1, mOat2, and mOat3, whereas the nitro group is required only by mOat1. Furthermore, the 8-methoxy group, although essential for toxicity, was not requisite for transport. Mouse renal cortical slices avidly accumulated AA-I, achieving slice-to-medium concentration ratios >10. Uptake by slices was sensitive to known mOat1 and mOat3 substrates and the organic anion transport inhibitor probenecid, which also blocked the production of DNA adducts formed with reactive intracellular metabolites of AA-I. Taken together, these findings indicate that OAT family members mediate high-affinity transport of AA-I and may be involved in the site-selective toxicity and renal elimination of this nephrotoxin.
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http://dx.doi.org/10.1124/jpet.111.180984DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3141898PMC
August 2011

Cytochrome P450 1A2 detoxicates aristolochic acid in the mouse.

Drug Metab Dispos 2010 May 17;38(5):761-8. Epub 2010 Feb 17.

Department of Pharmacological Sciences, State University of New York, One Nicolls Road, Stony Brook, NY 11794-8651, USA.

Aristolochic acids (AAs) are plant-derived nephrotoxins and carcinogens responsible for chronic renal failure and associated urothelial cell cancers in several clinical syndromes known collectively as aristolochic acid nephropathy (AAN). Mice provide a useful model for study of AAN because the renal histopathology of AA-treated mice is strikingly similar to that of humans. AA is also a potent carcinogen in mice with a tissue spectrum somewhat different from that in humans. The toxic dose of AA in mice is higher than that in humans; this difference in susceptibility has been postulated to reflect differing rates of detoxication between the species. Recent studies in mice have shown that the hepatic cytochrome P450 system detoxicates AA, and inducers of the arylhydrocarbon response protect mice from the nephrotoxic effects of AA. The purpose of this study was to determine the role of specific cytochrome P450 (P450) enzymes in AA metabolism in vivo. Of 18 human P450 enzymes we surveyed only two, CYP1A1 and CYP1A2, which were effective in demethylating 8-methoxy-6-nitro-phenanthro-(3,4-d)-1,3-dioxolo-5-carboxylic acid (AAI) to the nontoxic derivative 8-hydroxy-6-nitro-phenanthro-(3,4-d)-1,3-dioxolo-5-carboxylic acid (AAIa). Kinetic analysis revealed similar efficiencies of formation of AAIa by human and rat CYP1A2. We also report here that CYP1A2-deficient mice display increased sensitivity to the nephrotoxic effects of AAI. Furthermore, Cyp1a2 knockout mice accumulate AAI-derived DNA adducts in the kidney at a higher rate than control mice. Differences in bioavailability or hepatic metabolism of AAI, expression of CYP1A2, or efficiency of a competing nitroreduction pathway in vivo may explain the apparent differences between human and rodent sensitivity to AAI.
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http://dx.doi.org/10.1124/dmd.110.032201DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2872943PMC
May 2010

Akt and c-Myc differentially activate cellular metabolic programs and prime cells to bioenergetic inhibition.

J Biol Chem 2010 Mar 17;285(10):7324-33. Epub 2009 Dec 17.

Department of Molecular Genetics and Microbiology, Stony Brook University, Stony Brook, New York 11794, USA.

The high glucose consumption of tumor cells even in an oxygen-rich environment, referred to as the Warburg effect, has been noted as a nearly universal biochemical characteristic of cancer cells. Targeting the glycolysis pathway has been explored as an anti-cancer therapeutic strategy to eradicate cancer based on this fundamental biochemical property of cancer cells. Oncoproteins such as Akt and c-Myc regulate cell metabolism. Accumulating studies have uncovered various molecular mechanisms by which oncoproteins affect cellular metabolism, raising a concern as to whether targeting glycolysis will be equally effective in treating cancers arising from different oncogenic activities. Here, we established a dual-regulatable FL5.12 pre-B cell line in which myristoylated Akt is expressed under the control of doxycycline, and c-Myc, fused to the hormone-binding domain of the human estrogen receptor, is activated by 4-hydroxytamoxifen. Using this system, we directly compared the effect of these oncoproteins on cell metabolism in an isogenic background. Activation of either Akt or c-Myc leads to the Warburg effect as indicated by increased cellular glucose uptake, glycolysis, and lactate generation. When cells are treated with glycolysis inhibitors, Akt sensitizes cells to apoptosis, whereas c-Myc does not. In contrast, c-Myc but not Akt sensitizes cells to the inhibition of mitochondrial function. This is correlated with enhanced mitochondrial activities in c-Myc cells. Hence, although both Akt and c-Myc promote aerobic glycolysis, they differentially affect mitochondrial functions and render cells susceptible to the perturbation of cellular metabolic programs.
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http://dx.doi.org/10.1074/jbc.M109.035584DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2844180PMC
March 2010

Chromatin structure regulation in transforming growth factor-beta-directed epithelial-mesenchymal transition.

Cells Tissues Organs 2007 ;185(1-3):162-74

Department of Dermatology, New York University School of Medicine, New York, NY 10016, USA.

Epithelial-mesenchymal transitions (EMTs) occur in organogenesis throughout embryonic development and are recapitulated during epithelial tissue injury and in carcinoma progression. EMTs are regulated by complex, precisely orchestrated cell signaling and gene expression networks, with the participation of key developmental pathways. Here we review context-dependent modules of gene regulation by hairy/enhancer-of-split-related (H/E(spl)) repressors downstream of transforming growth factor-beta (TGF-beta)/Smad and Notch signals in EMT and in other phenotype transitions such as differentiation and cancer. Based on multiple models of disease-related EMT, we propose that Polycomb group epigenetic silencers and histone-lysine methyl-transferases EZH1 and EZH2 are candidate targets of H/E(spl)-mediated transcriptional repression, in a process accompanied by replacement of modified core histone H3 with de novo synthesized histone variant H3.3B. Finally, we discuss the potential significance of this scenario for EMT in the light of recent findings on gene regulation by histone modifications and chromatin structure changes.
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http://dx.doi.org/10.1159/000101317DOI Listing
July 2007