Section C. Biomarkers of Early Biological Effect and Altered Structure and Function
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2 Section C Biomarkers of Early Biological Effect and Altered Structure and Function The material in this video is subject to the copyright of the owners of the material and is being provided for educational purposes under rules of fair use for registered students in this course only. No additional copies of the copyrighted work may be made or distributed.
3 The Toxicological Process 3
4 Markers of Early Biological Effect! Assessment of molecular sequelae of the interaction of the active form of xenobiotic with its molecular target! Genetic alterations (mutation) in target and reporter genes - Mutated oncogenes, hprt, thymidine kinase, glycophorin A - Loss of tumor suppressor genes - Gene rearrangements 4
5 Markers of Early Biological Effect! Oncogenes and suppressor genes - Gatekeepers: control net cellular proliferation (division), e.g., APC, K-ras, p53 - Caretakers: maintain genomic integrity, e.g., hmsh2, hmlh1 5
6 p53 Tumor Suppressor Gene! Gene is located on chromosome 17p13; nuclear 53kD phosphoprotein! p53 is the gene most frequently found to be mutated in human tumors. Mutation of the gene results in a p53 protein with an altered structure and function. 6
7 p53 Tumor Suppressor Protein! First described in the 1970s as a cellular protein that coprecipitated with the large T antigen of simian virus 40 (SV40) and whose synthesis was enhanced in chemically transformed tumors! p53 protein binds to DNA and regulates the expression of cell cyclerelated genes. Wild type protein blocks cell cycle progression while mutant protein promotes cell proliferation. 7
8 p53 Protein Binding to DNA 8
9 Mutational Codon Hot Spots in the p53 Gene Colon 175, 248, 273, 282 Brain 273 Liver 249 9
10 Generation of Oligonucleotides for ESI-MS Source: Laken et al. (1998). Nature Biotechnology, 16,
11 Short Oligonucleotides for Analysis of Mutations! Short oligonucleotides generated for analysis of p53 codon 249 mutations (SOMA) 11
12 SOMA of DNA from Tumors and Plasma Pairs! SOMA of DNA from tumors and plasma pairs from two patients with HCC for G T mutation at codon 249 of p53 gene Source: Jackson et al. (2001). Cancer Research, 61,
13 Markers of Early Biological Effect! Altered enzymatic activities - Elevated protoporphyrin (Pb) - Decreased acetylcholinesterase (organophosphates) - Elevated xenobiotic metabolizing enzymes (TCDD) 13
14 Effects of Lead on Heme Synthesis! Effects of lead on heme synthesis in the mammalian erythrocyte 14
15 Markers of Altered Structure/Function! Assessment of functional and/or morphological changes or cells or organs following toxicant-cell interactions! Serum markers: elevated serum GSTs, ALA, SDH (liver toxicity), SGOT, creatinine kinase (myocardial infarction)! Urine markers: protein or glucose in urine! Imaging of the brain or heart (echocardiography) 15
16 Breath Biomarkers Biomarker Tissue injury Mechanism ethane, ethylene, 1-pentane isoprene injury caused by reactive oxygen species repair of injury caused by ROS lipid peroxidation biosynthesis of cholesterol and prenylated proteins acetone, 2-propanol ketonemia ketone bodies ethanol mechanism of NASH? gut bacterial synthesis acetaldehyde altered liver redox status and fatty acid metabolism oxidation of ethanol methanol unknown hydrolysis of demethylation products carbonyl sulfide, methyl mercaptan, dimethyl sulfide, dimethyl disulfide hepatotcellular injury incomplete metabolism of methionine ammonia, methylamine, dimethylamine renal injury uremia 16
17 Criteria for Selecting Biomarkers! Practical - Simple, inexpensive, high throughput! Analytical - Reliability: repeatability - Precision: sharply measured - Accuracy: measures true level - Validity: measures true change 17
18 Criteria for Selecting Biomarkers! Biological - Dynamism: biomarker can be modulated by eliminating exposure or by chemopreventive agents - Surrogacy: amount of effect of a chemopreventive agent explained by modulation of biomarker by agent 18
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