# Presentation: 179412

## EDSP  Validation

- Gary E.Timm
- Senior Technical Advisor
- Office of Science Coordination and Policy
-  U.S. Environmental Protection Agency

## Framework

**EPA proposed a Two Tier screening process:**
**Screening (Tier 1)**
- Identifies substances for further testing
**Testing (Tier 2)**
- Identifies adverse effects and establishes dose-response relationship for hazard assessment

## TIER 1 SCREENING Detect potential for endocrine disruption 

**Screens should be**
**inexpensive, quick, simple**
**standardized and validated**
**more sensitive than specific**
**able to detect multiple endpoints**
**readily interpretable**
**Use information to **
**make initial judgments**
**direct and focus tier 2 tests****   **

## Modes of Action

**Hormone synthesis and clearance**
**Hormone storage and release**
**Hormone transport**
**Receptor binding**
**Altered post-receptor activation**

## PROPOSED SCREENING BATTERY (Tier 1)

**PROPOSED SCREENING BATTERY (Tier 1)**

***In vitro *****Screens**
**ER Binding / Reporter Gene Assay***
**AR Binding / Reporter Gene Assay***
**Steroidogenesis Assay with minced testis**
**In vivo Screens**
**Rodent 3-day Uterotrophic Assay (sc)**
**Rodent 20-day Pubertal Female Assay with Thyroid**
**Rodent 5-7 day Hershberger Assay**
**Frog Metamorphosis Assay**
**Fish Reproduction Screening Assay**
*** ****These assays are in the HTPS**

## ALTERNATE SCREENING ASSAYS 

**Rodent 20-day Pubertal Male  Assay with Thyroid **
**Placental aromatase**
**Rodent in utero through Lactation Assay**

## Screening Battery  Example 1 

- <date/time>

**Screening Battery **** Example 1**** **

**In vitro Screens**
**ER Binding or Reporter Gene Assay***
**AR Binding or Reporter Gene Assay***
**Placental aromatase +**
**Steroidogenesis Assay with minced testis #**
**In vivo Screens**
**Rodent 3-day Uterotrophic Assay (sc)**
**Rodent 20-day Pubertal Male Assay with Thyroid +**
**Rodent 20-day Pubertal Female Assay w/ Thyroid #**
**Rodent 5-7 day Hershberger Assay #**
**Frog Metamorphosis Assay**
**Fish Gonadal Recrudescence Assay**
**+ Assays in yellow replace **** assays in red #**** **

## Screening Battery  Example 2

**Screening Battery **** Example 2**

**In vitro Screens**
**ER Binding or Reporter Gene Assay***
**AR Binding or Reporter Gene Assay***
**Steroidogenesis Assay with minced testis #**
**In vivo Screens**
**Rodent 3-day Uterotrophic Assay (sc) #**
**In-utero Assay +**
**Rodent 20-day Pubertal Female Assay w/ Thyroid #**
**Rodent 5-7 day Hershberger Assay #**
**Frog Metamorphosis Assay**
**Fish Gonadal Recrudescence Assay**
**+ Assays in yellow relace ****assays in red #**

## TIER 2 TESTING Confirm and characterize endocrine effects 

**Tests should **
**determine if effects are a primary or secondary disturbance of  endocrine function**
**establish exposure/concentrations/timing and effects relationships  **
**be sensitive and specific**
**assess relevant endpoints**
**include life cycle of live-bearing and egg-laying species**
**include a dose range for full characterization of  effects **
**be conducted in accordance with GLP**
**be validated   **

## PROPOSED  TIER 2 TESTING  BATTERY

**PROPOSED  TIER 2 ****TESTING  BATTERY**

**Multigeneration reproduction and development studies**
**Rodents**
**Birds**
**Frogs**
**Fish **
**Shrimp**

## Validation ProcessStakeholder Involvement

- Standardization and Validation Task Force
- Technical group made up of major stakeholders
- 3 expert workgroups: in vitro, mammalian, ecotoxicity
- To be replaced by FACA
- EDSPVAC
- To be chartered by September 2000
- Will advise EPA on Standardization and validation issues

## Validation of Test MethodsICCVAM

- What is it?  Interagency Coordinating Committee for the Validation of Alternative Methods
- What does it do?
-  Establises criteria for validation and regulatory acceptance of alternative test methods
- Set up process for regulatory acceptance of alternative methods

## Validation Process

- 1.  Research/Method Development
- 2.  Demonstration of Relevance
- 3.  Standardization of Protocol
- 4.  Validation in multiple laboratories
- 5.  Scientific Peer Review

## Validation ProcessStatutory Requirements

- All screens and tests in the EDSP must be validated
- The FIFRA Scienctific Advisory Panel (SAP) must review the EDSP
- The SAP must peer review all test guidelines to be used under FIFRA

## Validation ProcessICCVAM Role

- ER and AR binding assays to be validated by ICCVAM
- no laboratory work required
- generic performance criteria to be established rather than specific methods
- All other assays to be validated in EPA process with extensive ICCVAM coordination

##  EPAValidation ProcessICCVAM Role

- Method development and preparation of Background Review Document (BRD)
- Pre-validation--demonstration of relevance and development of standardized protocol
- Determination of readiness for validation in consultation with ICCVAM
- Validation
- Peer review by SAP/SAB and review by ICCVAM

##  VALIDATION APPROACH

**Human health related assays are highest EPA priority (2- 3 year time frame ) **
***in vitro***** assays**** **
**Uterotrophic, Hershberger**
**Female and male pubertal assays**
**Rodent  *****in utero***** screen **
**Rodent multigeneration demonstration**
**Ecotoxicological assays EPA second order priority**
**(3 -5 year time frame )**
**frog metamorphosis assay **
**fish reproductive screen**
**fish, frog, and invertebrate reproduction and development studies **

## VALIDATION APPROACH

**Select chemicals specific to each assay, validate assay rather than battery**
**More chemicals in pre-validation to establish relevance of assay and standardize the protocol than in validation phase**
**Battery validation will be a paper exercise**
**	****	****	**

## Ongoing EPA Work

**Completed initial demonstration of relevance**
**Female Pubertal Development Assay (4/2000)**
**Male Pubertal Development Assay     (4/2000)**
**Need to refine and standardize protocols**
**Initial developmental protocol trials**
**Frog metamorphosis assay****	****	****       (8/99)**
**Fish Reproduction Screening Assay     (8/99)**
**Additional effort needed on frog assay before a determination of relvance/usefulness can be made**

## OECD ENDOCRINE DISRUPTOR TESTING AND ASSESSMENT WORKGROUP

**Framework for Screening and Testing**
**Standardizing/validating three screens:**
**Uterotrophic**
**Hershberger**
**28-Day repeat dose study (OECD Method 407)**
**Updating existing guidelines**
**Example: Mammalian multi-generation reproduction study (OECD Method 416)**
**Developing new guidelines**
**Avian multi-generation reproduction study**
**Fish partial life-cycle reproduction research**

## Status of Assays

- <date/time>

## Status of Assays

- <date/time>

## Tier I Validation Program Projections

- <date/time>

**Tier I Validation Program Projections**

## Tier I Validation Program Projections

- <date/time>

**Tier I Validation Program Projections**

## Tier I Validation Program Projections

- <date/time>

**Tier I Validation Program Projections**

## Summary

- <date/time>

- Priority Setting

- Tier I Validation

- Tier II Validation

- Phase I
- Screening