# Presentation: 352917

## Toxicological Knowledge Base (a Definition)

**Response**

**Dose**

- “**An *****in computero****** *****aggregated set of the most germane literature ****citations**** and biological activity ****datasets****, together with computational ****models which correlate those activities with chemical structure****, and, ultimately, ****models for risk assessment****.”**

- Activity

- Structure

- Correlation

## A circular process provides training sets DESIGNED to produce robust, living predictive models

**EDKB**

**Data validation**
**and selection**

**Diversity analysis**
**Training set design**

**New Data Needs &**
**Research Hypothesis**

**NCTR/EPA**
**Experiments**

**Data in **
**scientific **
**papers**

**Develop**
**Models**

**Validation:**
**1. Cross-validation**
**2. Testset challenge**

## EDKB OBJECTIVES

- Develop and validate predictive computational models for ER and AR by:
- developing appropriate data sets to train models
- examining a wide variety of models for performance and ease of use
- validating models by:
- internal validation
- external validation from non-randomly selected chemicals
- external validation from randomly selected chemicals from the 58,000 chemical EPSD

## Dataset Criteria for Development of SAR models

- Develop training data set by design for
-  structurally diverse chemical structures
- a wide range of binding values (RBAs)

## Status of ER Models

- 238 chemicals assayed (duplicate tubes; two replicates) over a six log RBA range
- Techniques developed for identification of potential binders by EDKB team
- Filters (Phase I) and 11 categorical SAR models (Phase II) finished
- Models validated against literature datasets

## Status of ER Models (2)

- Models biased toward false positives to minimize false negatives
- RBA predictions made for all 58,000 chemicals “in play”
- Handout of ER model publications
- ~ First two years-used literature RBAs
- Subsequently used NCTR dataset

## Androgen Receptor Competitive Binding Assay

- Two assays evaluated:
- Ventral prostate from castrated adults
- Could not obtain reproducible results
- PanVera androgen binding domain
- Good saturation and competitive binding results
- Assay subsequently validated for use
- Less expensive than ventral prostate
- Uses no animals

## Validation of PanVera Assay

- PanVera protein was diluted to several low concentrations and saturation assays were conducted with [3H]-R1881.
- Protein concentrations selected were in the range of the reported KD.
- A protein concentration in the stable KD range was used for subsequent competitive binding assays.

## AR Competitive Binding Results for Steroidal Androgens

## AR Competitive Binding Results for Some Steroidal Estrogens 

## AR Competitive Binding Results for Antiestrogens

## Status of AR Assay

- Up to May 9, 2001, 134 compounds were tested.
- To date,196 compounds have been tested

**Number of Chemicals**

## Activity Distribution for AR

**Number of Chemicals**

## Comparison of NCTR Assay with EPA Assay

## Primary Pharmacophore Model

## CoMFA Model

**CoMFA Model**

## De novo external validation of assays and models

- EPA funded $850K contract with Batelle Northwest for validation and comparison performance of NCTR and EPA assays and models
- Assays methods include:
- uterine cytosol for ER  (EPA and NCTR assays are similar)
- prostate assay for AR (EPA)
- PanVera assay for AR (NCTR)

## EDKB EXTERNAL FUNDING

**  ****SOURCE****	****	****	****    AMOUNT  ($)**

-    FDA OWH	1				185,000
-    FDA OWH 2				325,000
-    CMA CRADA 2				420,000
-    EPA GRANT 1				425,000
-    EPA IAG 3					_2,000,000_
-    TOTAL					3,355,000
-    1   Experimental collaboration with U Missouri
-      2  Two separate awards
-      3  $500,000 for FY 2001; $1,500,000 for FY 2002-2005

## What did the Buddhist say to the hot dog vendor?

## Make me one with everything