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Assessing Carcinogenic Potential of Chemicals Using OncoLogic Cancer Expert System. Yin-tak Woo, Ph.D., DABT Office of Pollution Prevention and Toxics U.S. Environmental Protection Agency Washington, DC 20460 May 19, 2010. Outline of the Presentation.

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Assessing carcinogenic potential of chemicals using oncologic cancer expert system l.jpg

Assessing Carcinogenic Potential of Chemicals UsingOncoLogic Cancer Expert System

Yin-tak Woo, Ph.D., DABT

Office of Pollution Prevention and Toxics

U.S. Environmental Protection Agency Washington, DC 20460

May 19, 2010


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Outline of the Presentation

  • Scientific background in development of the OncoLogic system

  • Brief description of the OncoLogic system

  • Recent development in updating and expanding OncoLogic system


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SAR/QSAR: Background & Issues

  • SAR/QSAR: activity = f (structure)

  • Given sufficient data/knowledge on related compounds  screen well defined endpoint

  • Evolution of SAR/QSAR: from human intuition to cyber sophistication

  • Impact of commercial software

  • User base: from domain expert to nonscientists

  • Pressure of reduction in experimentation


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Approaches to SAR/QSAR

  • Statistical vs. Mechanistic

  • Local/Homogeneous/Congeneric vs. Global/General/Heterogeneous

  • Active/Inactive vs. Ranking vs. Potency


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Criteria for assessing scientific soundness

  • Selection of endpoint

  • Knowledge base/training database/applicability domain

  • Methodology and descriptor selection

  • Model validation (predictive accuracy, internal, external, prospective)

  • Transparency and scientific rationale

  • Confidence/uncertainty analysis

  • Strengths, weaknesses and limitations


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Importance of mechanistic understanding in (Q)SAR modelling

  • selection of toxicological endpoint

  • selection of molecular descriptors

  • coverage of training database

  • consideration of database stratification

  • interpretation of outliers

  • consideration of human relevance

  • achieving the goal of statistical association with mechanistic backing


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ADME/Toxicokinetics considerationAbility of toxicant to reach target tissue/molecule

  • Chemical structure/phys-chem on ADME

  • Route of administration

  • Facilitating “carrier” molecule

  • Protective “carrier” molecule

  • Biological half-life


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Mechanistic/Toxicodynamics consideration

  • Electrophilic

  • Receptor-mediated

  • Disruption of homeostasis

  • Multiple mechanisms


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Future Trend of (Q)SAR

  • Critical evaluation of current methods

  • Expansion of publicly accessible databases/knowledge bases

  • Expansion of integrative approaches

  • Utilization of input from emerging predictive technologies


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Incorporating emerging predictive technologies

Chemical

In silico ADME,

ADME assays

ADME/Toxicokinetics

Consideration

T

O

X

I

C

G

G

E

N

O

M

I

C

S

Mechanistic/Toxicodynamics

Consideration

HTS assays

Structural

Functional

Molecular descriptors,

Structural features, etc

Screening assays,

Biomarkers, etc

Prediction


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Major References for (Q)SARas a Screening Tool

Woo YT, Lai DY (2003): Mechanism of action of chemical carcinogens and their role in SAR analysis and risk assessment. In: Quantitative Structure-Activity Relationship (QSAR) Models of Mutagens and Carcinogens, R. Benigni, ed., CRC Press, Boca Raton, FL

Doull D, Borzelleca J, Becker R, Daston G, DeSesso J, Fan A, Fenner-Crisp P, Holsapple M, Holson J, Llewellyn G, MacGregor J, Seed J, Walls I, Woo Y, Olin S (2007): Framework for use of toxicity screening tools in context-based decision-making. Food Chem.Toxicol. 45:759-796.



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Introduction to the Cancer Endpoint Background

  • Definitions

    • Uncontrolled dividing and growth of cells

    • Caused by mutations, ↑ cell proliferation, ↓ cell death, loss of homeostatic control, etc.

  • Two general mechanisms by which a chemical can induce cancer

    • Genotoxic (default)

      • Interaction with DNA to cause mutation(s) in genes

    • Non-genotoxic

      • Variety of mechanisms


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Introduction to the Cancer Endpoint Background(Cont.)

  • Carcinongesis is a multistage/multistep process

    • Initiation

      • Mutation converts normal to preneoplastic cells

    • Promotion

      • Expansion of preneoplastic cells to benign tumors

    • Progression

      • Transformation of benign to invasive malignant tumors

  • A potent carcinogen acts directly on all three stages

  • A weak carcinogen acts directly on one stage and indirectly on other


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Initiation Background

Promotion

Progression

Main event(s)

Direct DNA

binding

Indirect DNA

damage

Clonal expansion

Cell proliferation

Apoptosis

Differentiation

Overcoming suppressions (e.g., p53, immune, angiogenesis)

Key mechanistic

consideration

Electrophile, resonance stabilization, nature of DNA adduct

Receptor, cytotoxicity,

gene expression

Free radical, receptor, gene suppression

Signal transduction, homeostasis

SAR/QSAR mechanistic descriptors

Electrophilicity, HOMO/LUMO, delocalization energies, ……

2D, 3D, docking, biopersistence, methylation, ….

Reduction potential, 2D, 3D, ……


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Difficulties of (Q)SAR of carcinogenicity Background

  • Complex, mechanism-dependent (Q)SAR

  • Local vs. global models

  • Data scarcity and variability

  • Feedback and validation issues

  • Need for integrative approach


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Historical development of OncoLogic Background

  • TSCA and New Chemicals Program (PMN)

  • Structure-Activity Team approach

  • Need to provide guidance to industries

  • OncoLogic Team (Joseph Arcos, Mary Argus, David Lai, Yin-tak Woo)

  • LogiChem coop version

  • Current version

  • Future developments


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OncoLogic: A mechanism-based expert system for predicting carcinogenic potential

  • Developed by domain experts in collaboration with expert system developer

  • Knowledge from SAR on >10K chemicals

  • Class-specific approach to optimize predictive capability

  • Consider all relevant factors including biological input when possible

  • Predictions with scientific rationale and semiquantitative ranking


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Major Sources of Data/Insight Used to Develop Cancer Knowledge Rules

  • The OncoLogic Team and members of SAT

  • Chemical Induction of Cancer monograph series

  • IARC monograph series

  • NCI/NTP technical reports

  • Survey of compounds which have been tested for carcinogenic activity, PHS Publ. 149

  • Non-classified EPA submission data from various EPA program offices

  • Current literature and ad hoc expert panels


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Profile of most potent carcinogens Knowledge Rules

  • Ability to reach target tissue

  • Reasonable lifetime of ultimate carcinogen

  • Persistent and site-specific interaction with target macromolecule

  • Ability to affect all three stages of carcinogenesis


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Development of rules for each class Knowledge Rules

  • Gather all available data and information

  • Brainstorming to determine key factors

  • Determine need for subclassification

  • Assign concern levels to known carcinogens

  • Determine mechanism-based modification factors for substituents

  • Develop rationale for conclusion


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Concern Levels Knowledge Rules


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Critical Factors for SAR Knowledge RulesConsideration

  • Electronic and Steric Factors

    • Resonance stabilization

    • Steric hindrance

  • Metabolic Factors

    • Blocking of detoxification

    • Enhancement of activation


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Critical Factors for SAR Knowledge RulesConsideration

  • Mechanistic Factors

    • Electrophilic vs. receptor- mediated

    • Multistage process

  • Physicochemical Factors

    • Molecular weight

    • Physical state

    • Solubility

    • Chemical reactivity


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OncoLogic® Knowledge RulesFactors Affecting Carcinogenicity of Aromatic Amines

  • Number of aromatic ring(s)

  • Nature of aromatic ring(s) - homocyclic vs. heterocyclic - nature and position of heteroatoms

  • Number and position of amino or amine-generating groups(s) - position of amino group relative to longest resonance pathway - type of substituents on amino group

  • Nature, number, position of other ring substituent(s) - steric hindrance - hydrophilicity

  • Molecular size, shape, planarity


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R-NO Knowledge Rules2 R-NO R-NH-OH R-NH2

R-NH-OAc [R-N(CH3)2]

Some Hydrocarbon Moieties Present in Carcinogenic Aromatic Amines


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- Knowledge Rules

+

+

+

Carbonium ion

Amidonium ion

Molecular Mechanism for Generation of Resonance-stabilized Reactive Intermediates from N-acyloxy Aromatic Amines


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5’ Knowledge Rules

6’

6

5

4’

4’

3’

2’

2

3

Very active if:

OC•CH3

--N

OH

Active if:

--NH•OC•CH3

--N(CH3)2

--NO2

--OCH3

Inactive if:

--F

R

|

--CH--

Transition to

diphenylmethane

and triphenyl-

methane amines

--CH==CH--

Transition to amino-

stilbenes

--N==N--

Transition to amino

azo dyes

Very active if:

--F

Active if:

--NH2

--NH•OC•CH3

--NO2

Weakly active if:

--C6H5

Inactive if:

--CH3

--Cl

--Br

Active if:

--S--

Inactive if:

--NH--

For 4-aminobiphenyl:

Very active if:

3-methyl

3,2’-dimethyl

3,3’-dimethyl

3-fluoro

3’-fluoro

Active if:

3-chloro

3-methoxy

3,2,5’-trimethyl

3,2’,4’,6’-tetramethyl

Weakly active if:

3-hydroxy

Inactive if:

2-methyl

2’-methyl

2’-fluoro

3-amino

For benzidine:

Very active if:

2-methyl

3,3’-dihydroxy

3,3’-dichloro

Weakly active if:

3,3’-dimethyl

3,3’-dimethoxy

Inactive if:

2,2’-dimethyl

3,3’-bis-oxyacetic acid

Synoptic Tabulation of Structural Requirements for Carcinogenic Activity of 4-Aminobiphenyl and Benzidine Derivatives


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C = Clear evidence of carcinogenicity Knowledge Rules

S = Some evidence of carcinogenicity

N = No evidence of carcinogenicity

NT = Not tested

+ = At least one test = C or S

Eq = No C or S, and E must appear at least once

-- = No C, S, or E

OncoLogic® Prediction vs. NTP BioassaysAromatic Amines and Related Compounds


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Examples of how “Knowledge Rules” can be used in chemical design

Strategies to Designing Safer Chemicals:

  • Steric hindrance

  • Nonplanarity

  • Electronic insulation

  • Hydrophilicity

OncoLogic Cancer Concern = High










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C = Clear evidence of carcinogenicity Carcinogenic Potential (Cont.)

S = Some evidence of carcinogenicity

N = No evidence of carcinogenicity

NT = Not tested

+ = At least one test = C or S

Eq = No C or S, and E must appear at least once

-- = No C, S, or E

OncoLogic® Prediction vs. NTP BioassaysAromatic Amines and Related Compounds


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Conclusions from NTP Cancer Bioassays Predictive Exercises Carcinogenic Potential (Cont.)

  • Most of the best performers are predictive systems that incorporate human expert judgment and biological information

  • OncoLogic was one of the best performers among more than 15 methods


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Final results of 2 Carcinogenic Potential (Cont.)nd NTP predictive exercise(from Benigni and Zito, Mutat. Res. 566, 49, 2004)


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FDA Validation of Genetic Toxicity and SAR Carcinogenic Potential (Cont.)

Methods for Predicting Carcinogenicity*

*from Mayer et al.: SAR analysis tools: validation and applicability in predicting carcinogens. Regulatory Toxicol. Pharmacol. 50: 50-58, 2008


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Sensitivity and Specificity of the Genetic Toxicity Carcinogenic Potential (Cont.)

and SAR Methods for Predicting Carcinogenicity


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FDA Validation of Genetic Toxicity and SAR Carcinogenic Potential (Cont.)

Methods for Predicting Potent Carcinogenicity


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OncoLogic® - Benefits Carcinogenic Potential (Cont.)

  • Allow non-experts to reach scientifically supportable conclusions

  • Expedites the decision making process

  • Allows sharing of knowledge

  • Reduces/eliminates error and inconsistency

  • Formalize knowledge rules for cancer hazard identification (SAT-style)

  • Bridge expertise of chemists and toxicologists for most effective hazard evaluation

  • Provide guidance to industries on elements of concern for developing safer chemicals


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Some Notable Uses of OncoLogic Carcinogenic Potential (Cont.)

  • OPPT (new chemicals, design for environment, green chemistry, existing chemicals)

  • Guidance to industries (Sustainable Future program)

  • OW (disinfection byproduct prioritization) and other EPA program offices

  • FDA (food contact notification) and other governmental agencies


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Developed by recognized domain experts Carcinogenic Potential (Cont.)

Knowledge not just data

Local models with strong mechanistic basis

Integrates biological input when possible

Semiquantitative ranking with scientific rationale

Proven performance in prospective and external validations

Industrial chemicals

Users need to have some organic chemistry background

Coverage limited by available knowledge

No batch mode

Some updates are needed

Current coverage mainly on established carcinogen classes

Limited receptor-based SAR

Pharmaceuticals?

Strengths Limitations


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Overall Structure of The Cancer Expert System Carcinogenic Potential (Cont.)


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Running OncoLogic® Carcinogenic Potential (Cont.)

  • Two methods to predict carcinogenicity

    • SAR Analysis

      • Knowledge rules

    • Functional Analysis

      • Uses results of specific mechanistic/non-cancer studies


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SAR Analysis Carcinogenic Potential (Cont.)

  • Four modules

    • Organics

    • Metals

    • Polymers

    • Fibers

  • Different method used to evaluate each type


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Running OncoLogic Carcinogenic Potential (Cont.)® : Organics Module

  • Organics

    • Enter information on chemical identity

    • Choose appropriate chemical class

    • Enter chemical name, CAS#, or chemical structure


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Running OncoLogic®: Carcinogenic Potential (Cont.)Organics Module

  • Select chemical class

    • 48 total

    • Description in Manual

    • Hit “F1” to view sample structures

  • Absence of structure in OncoLogic provides suggestive, but not definitive, evidence of lack of major cancer concern. Functional arm should be used if possible.


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Running OncoLogic®: Carcinogenic Potential (Cont.)Organics Module

  • Pick Correct Backbone Structure if Provided

  • Draw chemical


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Running OncoLogic®: Carcinogenic Potential (Cont.)Organics Module


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Running OncoLogic®: Carcinogenic Potential (Cont.)Organics Module

Perform evaluation


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OncoLogic® Justification for Carcinogenic Potential (Cont.)Organics Module

OncoLogic®(R) Justification Report

CODE NUMBER: Isodecyl Acrylate Example

SUBSTANCE ID: 1330-61-6

The final level of carcinogenicity concern for this acrylate when

the anticipated route of exposure is inhalation or injection is

MARGINAL.

JUSTIFICATION:

An acrylate is a potential alkylating agent which may bind, via

Michael addition, to key macromolecules to initiate/exert

carcinogenic action. The alkylating activity of acrylates can be

substantially inhibited by substitution at the double bond,

particularly by bulky or hydrophilic groups..........................


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Other Chemicals Carcinogenic Potential (Cont.)

  • In addition to SAR analysis, OncoLogic includes evaluations of approximately 90 specific chemicals that do not fit into any OncoLogic class


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Other Chemicals (Cont.) Carcinogenic Potential (Cont.)

  • Locate chemical by CAS number or by name


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Running OncoLogic®: Carcinogenic Potential (Cont.)Metals

  • Similar to running the organics module

  • Pick the metal to be evaluated

    • OncoLogic® will then either ask a series of questions needed to evaluate the chemical or provide a database of related compounds


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Information Needed to Run the Carcinogenic Potential (Cont.)Metals Module

  • Nature/form of the metal / metalloid

    • Organometal, metal powder

  • Type of chemical bonding (e.g., organic, ionic)

  • Dissociability / solubility

    • Valence / oxidation state

  • Crystalline or amorphous

  • Exposure scenario

  • Breakdown products (e.g., organic moieties)


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Running OncoLogic® Carcinogenic Potential (Cont.)Polymers

  • Polymer must consist of covalently linked repeating units and have a number average molecular weight >1000

  • OncoLogic® asks a series of questions designed to aid in evaluation of carcinogenicity of the polymer


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Polymers Module Carcinogenic Potential (Cont.):Information Needed to Evaluate Polymers

  • Percentage of polymer with molecular weight <500 and <1000

  • Percent of residual monomer

  • Identification of Reactive Functional Group(s)

  • Solubility

  • Special features

    • Polysulfation, "water-swellability"

  • Exposure route

  • Breakdown products (e.g., hydrolysis)


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Fibers Carcinogenic Potential (Cont.)Module

Evaluations are based on physical dimensions and physicochemical properties

Physical dimensions

Diameter, length, aspect ratio

Physicochemical properties

High density charge, flexibility, durability, biodegradability, smooth and defect-free surface, longitudinal splitting potential

Presence of high MW polymer, low MW organic moiety, metals/metalloids


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Fibers Module Carcinogenic Potential (Cont.) (Cont.)

Relevant manufacturing / processing / use information

Crystallization, thermal extrusion, naturally occurring, unknown method


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The Carcinogenic Potential (Cont.)Functional Arm of OncoLogic®


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Functional Arm (Cont.) Carcinogenic Potential (Cont.)

  • OncoLogic® can use results from some shorter-term tests to support a cancer concern.

  • Results indicate whether chemical may be an initiator, promoter, or progressor


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Initiator Carcinogenic Potential (Cont.)

M/HM

M/HM

HM/H

Promoter

Progressor

LM/M

Use of Non-Cancer Data:Functional Arm (Cont.)

  • Functional Arm predicts whether the chemical is likely to be a tumor initiator, promoter, and/or progressor

    • Possible relevance or contribution to the carcinogenesis process is indicated in the figure below


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Major References on OncoLogic® Carcinogenic Potential (Cont.)

Woo, Y.-T., Lai, D.Y., Argus, M.F. and Arcos, J.C. Development of Structure Activity Relationship Rules for Predicting Carcinogenic Potential of Chemicals. Toxico. Lett. 79: 219-228, 1995.

Woo, Y.-T., Lai, D.Y., Argus, M.F. and Arcos, J.C. Carcinogenicity of Organophosphorous Pesticides/Compounds: An analysis of their Structure Activity Relationships. Environ. Carcino. & Ecotox. Revs. C14(1), 1-42, 1996.

Lai, D.Y., Woo, Y,-T., Argus, M.F. and Arcos, J.C.: Cancer Risk Reduction Through Mechanism-based Molecular Design of Chemicals. In:"Designing Safer Chemicals" (S. DeVito and R. Garrett, eds.), American Chemical Society Symposium series No. 640, American Chemical Society Washington, DC. Chapter 3, pp.62-73, 1996.

Woo, Y.-T. et al.: Mechanism-Based Structure-Activity Relationship Analysis of Carcinogenic Potential of 30 NTP Test Chemicals. Environ. Carcino. & Ecotox. Revs. C15(2), 139-160, 1997.

Woo, Y., Lai, D., Argus, M.F., and Arcos, J.C.: An Integrative Approach of Combining Mechanistically Complementary Short-term Predictive Tests as a Basis for Assessing the Carcinogenic Potential of Chemicals. Environ. Carcino. & Ecotoxicol. Revs. C16(2), 101-122, 1998.


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Major References - continued Carcinogenic Potential (Cont.)

Woo, Y.-T., and Lai, D.Y. : Aromatic Amino and Nitroamino Compounds and their Halogenated Derivatives. In: Patty’s Toxicology, 5th edn., E. Bingham, ed., Wiley, pp. 969-1106, 2001

Woo, Y-T., Lai, D., McLain, J., Manibusan, M., Dellarco, V.: Use of Mechanism-Based Structure-Activity Relationships Analysis in Carcinogenic Potential Ranking for Drinking Water Disinfection Byproducts. Environ. Health Persp. 110 (suppl. 1): 75-88, 2002.

Woo, Y.-T., and Lai, D.Y. : Mechanism of Action of Chemical Carcinogens and their Role in Structure Activity Relationships (SAR) Analysis and Risk Assessment. In: Quantitative Structure-Activity Relationship (QSAR) Models of Mutagens and Carcinogens. R. Benigni, ed., CRC Press, Boca Raton, FL., pp. 41-80, 2003.

Woo, YT, and Lai DY: OncoLogic: A mechanism-based expert system for predicting the carcinogenic potential of chemicals. In: Predictive Toxicology, C. Helma, editor, Taylor and Francis, Boca Raton, FL., pp. 385-413, 2005.


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Downloading and Contact Information Carcinogenic Potential (Cont.)

Training and limited technical questions

[email protected]

Scientific questions

[email protected]

Downloading

http://www.epa.gov/oppt/newchems/tools/oncologic.htm


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Recent development: OncoLogic 7.0 Carcinogenic Potential (Cont.)

  • Windows 7.0 and XP-compatible

  • Improved drawing package

  • Improved printer functionality

  • Drop-down menus of CAS number and chemical names within each class/subclass

  • Ready for downloading soon


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OncoLogic 8.0: Work in progress Carcinogenic Potential (Cont.)

  • Master list of chemicals

  • Limited expansion and updates

  • “Low-potential” classes with delineation of exceptions and precautionary notes

  • Nongenotoxic SA; input from HTS/TXG?

  • Integrative approaches

  • Nanomaterial?


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Difficulties of negative predictions Carcinogenic Potential (Cont.)

  • Scarcity and uncertainty of negative data

  • Less certain mechanistic basis of negativity

  • Difficult to exhaust all possible mechanisms

  • Need for supportive data (e.g, mode of action, pathways, threshold, spp. difference)

  • Regulatory caution of negative predictions for high exposure chemicals


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