Aristolochic Acid Research

Toxicogenomics

Rosenquist

Individual variation in the response to drugs and to environmental exposures is an ongoing challenge to medicine. We are investigating the genetic factors contributing to susceptibility to the nephrotoxin and carcinogen, aristolochic acid (AA). Only 5% of humans exposed to AA develop symptoms resulting from renal toxicity. Of those people half will also develop transitional cell carcinoma of the upper urinary tract. This is a problem of some magnitude as Aristolochia have been used worldwide in traditional medicines for thousands of years and 50-300 million people, primarily in Asia, are currently exposed. Understanding the genetic factors underlying sensitivity to this toxin will allow the identification of people at risk and will give insight to the mechanism by which AA brings about disease.
Image: Toxicogenomics graph - fig 1

Metabolism of AA in mice - AA must be activated via nitro-reduction before reacting with DNA to form pro-mutagenic DNA adducts. Bio-inactivation is achieved by oxidative de-methylation.  Candidate proteins have been identified that participate in the metabolism of AA. We have used gene-targeted mouse strains to probe the contribution of specific proteins to these processes. Thus, we have shown that the cytochrome P450, CYP1A2, contributes significantly to the bio-inactivation of AA (Rosenquist, et al, 2010). However after induction of anti-xenobiotic pathways, including those induced by AA itself, CYP1A1 protein contributes to AA-inactivation as well. The study of bio-activation of AA is an ongoing project in the Laboratory of Chemical Biology and we are using a mouse models to address in vivo contributions of the specific genes identified.

Genetic Susceptibility to AA in mice - Inbred strains of mice vary in their response to AA. We have characterized these responses (Huang, et al, 2009; Shibutani, et al, 2010) and have developed a streamlined protocol to detect the acute renal toxicity following AA-exposure in mice (Rosenquist, 2011). We used this protocol to identify resistant and susceptible strains of mice for genetic analysis. Employing hybrid mice produced by breeding two such strains, we have defined four chromosomal regions called quantitative trait loci (QTL), designated Aanq1-4, that contribute to strain differences in AA-sensitivity (Rosenquist, 2011). At least one of these loci, Aanq1, also determines differences in kidney function between the untreated mice of each strain. We are fine-mapping each QTL to determine the specific gene that contributes to each. 
Image: Toxicogenomics graph - fig 2

Human genetic variants associated with AAN - Through collaborations with clinical investigators in Croatia, Belgium, and Taiwan, we have accumulated DNA samples from human populations exposed to AA, along with their relevant clinical histories. Our goal is to define specific gene-variants associated with the various manifestations of AA-induced disease. We have identified a small set of gene variants associated with aristolochic acid nephropathy. We use several approaches to validate these observations including a family-based genetic association screen among residents of AAN endemic villages in Croatia, as well as determining the consequence for AA-sensitivity of expressing specific gene variants in renal cells in culture. 
  • Rosenquist T.A., Grollman A.P. Mutational signature of aristolochic acid: Clue to the recognition of a global disease. DNA Repair 44:205-211, 2016. PMID27237586
  • Hoang M.L., Chen C.H., Chen P-C., Roberts N.J., Dickman K.G., Yun B.H., Turesky R.J., Pu Y-S., Vogelstein B., Papadopoulos N., Grollman A.P., Kinzler K.W., Rosenquist T.A. Aristolochic Acid in the etiology of renal cell carcinoma. Cancer Epidemiol Biomarkers Prev 113(35):9846-9851, 2016. PMID27555084
  • Yun B.H., Yao L., Jelaković B., Nikolić J., Dickman K.G., Grollman A.P., Rosenquist T.A., Turesky R.J. Formalin-fixed paraffin-embedded tissue as a source for quantitation of carcinogen DNA adducts: aristolochic acid as a prototype carcinogen. Carcinogenesis 35(9):2055-2061, 2014. PMID24776219
  • Hoang M.L., Chen C-H., Sidorenko V.S., He J., Dickman K.G., Yun B.H., Moriya M., Niknafs N., Douville C., Karchin R., Turesky R.J., Pu Y-S., Vogelstein B., Papadopoulos N., Grollman A.P., Kinzler K.W., Rosenquist T.A. Mutational Signature of Aristolochic Acid Exposure as Revealed by Whole Exome Sequencing. Sci Transl Med 5(197):197ra102, 2013. PMID23926200
  • Yun B., Rosenquist T., Sidorenko V., Iden C.R., Chen C.H., Pu Y.S., Bonala R., Johnson F., Dickman K.G., Grollman A.P., and Turesky R.J. Biomonitoring of aristolactam-DNA adducts in human tissues using ultra-performance liquid chromatography/ion-trap mass spectroscopy. Chem Res Toxicol 25:1119-31, 2012. PMID22515372
  • Rosenquist TA. Genetic loci that affect aristolochic acid nephrotoxicity in the mouse. Am J Physiol Renal Physiol. 300(6):F1360-7, 2011.  PMID21429970
  • Rosenquist TA, Einolf HJ, Dickman KG, Wang L, Smith A, Grollman AP. Cytochrome P450 1A2 detoxicates aristolochic acid in the mouse. Drug Metab Dispos. 38(5):761-8, 2010.  PMID20164109
  • Shibutani S, Bonala RR, Rosenquist T, Rieger R, Suzuki N, Johnson F, Miller F, Grollman AP.  Detoxification of aristolochic acid I by O-demethylation; less nephrotoxicity and genotoxicity of aristolochic acid Ia in rodents. Int J Cancer. 127(5):1021-7, 2010.  PMID20039324
  • Huang F, Clifton J, Yang X, Rosenquist T, Hixson D, Kovac S, Josic D. SELDI-TOF as a method for biomarker discovery in the urine of aristolochic-acid-treated mice. Electrophoresis 30(7):1168-74, 2009.  PMID19294690

 


 

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