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Section C
Biomarkers of Early Biological Effect and
Altered Structure and Function
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The Toxicological Process
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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
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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
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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.
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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.
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p53 Protein Binding to DNA
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Mutational “Codon Hot Spots” in the p53 Gene
Colon
175, 248, 273, 282
Brain
273
Liver
249
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Generation of Oligonucleotides for ESI-MS
Source: Laken et al. (1998). Nature Biotechnology, 16, 1352–1356.
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Short Oligonucleotides for Analysis of Mutations
! Short oligonucleotides generated for analysis of p53 codon 249
mutations (SOMA)
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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, 33–35.
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Markers of Early Biological Effect
! Altered enzymatic activities
- Elevated protoporphyrin (Pb)
- Decreased acetylcholinesterase (organophosphates)
- Elevated xenobiotic metabolizing enzymes (TCDD)
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Effects of Lead on Heme Synthesis
! Effects of lead on heme synthesis in the mammalian erythrocyte
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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)
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Breath Biomarkers
Biomarker
Tissue injury
Mechanism
ethane, ethylene, 1-pentane
injury caused by reactive
oxygen species
lipid peroxidation
isoprene
repair of injury caused by ROS
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
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Criteria for Selecting Biomarkers
! Practical
- Simple, inexpensive, high throughput
! Analytical
- Reliability: repeatability
- Precision: sharply measured
- Accuracy: measures “true” level
- Validity: measures “true” change
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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
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