PESTICIDE ASSESSMENT GUIDELINES
SUBDIVISION F
HAZAFD EVALUATION:
Hl*WE AND DOMESTIC ANIMALS
Series 83-3
Rat or Rabbit Developmental Toxicity Study
ADDENDUM 1 ON DATA REPORTING
Prepared by:
Science Mission Support Staff
Toxicology Branch
Hazard Evaluation Division
Project Manager:
Elizabeth M.K. Leovey, Ph.D
Hazard Evaluation Division
Office of Pesticide Programs
U.S. Environmental Protection Agency
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Subdivision F - Rat or Rabbit Developmental Toxicity Study
Table of Contents of Mdendum
Discussion - Page
Introduction i
Response to Public Ccraments i
Guideline
Cover Page 1
Table of Contents 1
Body of the Report 1
I. Summary 1
II. Introduction 1
III. Materials/Methods 1
IV. Results 2
V. Discussion 3
VI. Bibliography 3
VII. Verification 3
VIII. Archives 3
IX. Tables/Figures 3
X. Appendix(es) 3
Example Formats for
Tables 1-8 4
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PESTICIDE ASSESSMENT GUIDELINES
HUMANS AND DOMESTIC ANIMALS
Rat or Rabbit Developmental Toxicity Study
Subdivision F, Series 83-3
DATA REPORTIN3
INTRODUCTION
A. Purpose
This study is designed to determine the potential of the test substance
to induce structural and/or other abnormalities to the fetus which may
arise from exposure of the mother during pregnancy.
B. Objective .
The objective of this Guideline is to provide an example of an acceptable
format for reporting materials, methods and results of developmental
toxicity studies. These recommendations are intended to show how the
requirements in §80-4 and §83-3(h) of this Subdivision and requirements
in the Agency's Good Laboratory Practice Standards (40 CFR Part 160) may
be organized in a Final Report that can be reviewed effectively and
expeditiously in accordance with "Proposed Guidelines for the Health
Assessment of Suspect Developmental Toxicants and Request for Comments"
(Federal Register, Vol. 49, No. 227, November 23, 1984, pages 46324-46331)
and the Hazard Evaluation Division's Standard Evaluation Procedure for
Teratology studies (EPA-540/9-85-018 published for the Agency in June,
1985). While following this Guideline is not mandatory, data submitters
are encouraged to submit complete reports which can be efficiently
reviewed by the Agency.
RESPONSE TO PUBLIC COMMENTS
Comments were received frcm 17 sources including testing facilities,
pesticide manufacturers, a manufacturer's organization, a scientific
association, and one state government agency. There were three general
concerns described by most conroenters. Specific comments on the Teratology
Data Reporting Guideline (Addendum to 583-3 of Subdivision F of the
Pesticide Assessment Guidelines) were related to those three issues.
A. Garments
The proposed Data Reporting Guideline (DRG) was described as incon-
sistent with guidelines published in Subdivision F (§80-4 and §83-3), 40
CFR Part 160, Good Laboratory Practice Standards, and "Proposed Guidelines
for the Health Assessment of Suspect Developmental Toxicants and Request for
Conments" (Federal Register, Vol. 49, No. 227, November 23, 1984, pages 46324-
46331). Corenenters also expressed concern that the Teratology DRGs repre-
sented an extension of previous Guidelines to include conduct of the study.
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The second concern most conmenters had about the proposed ERG was
that it does not acccroodate computerized systems developed by various
laboratories for presenting and analyzing data from Developmental Tbxicity
studies.
Finally, connenters were concerned that the proposed format may represent
a trend toward a clerical, statistical, or "checklist" approach to interpre-
tation of results rather than one that depends upon scientific judgement.
B. Response
The Agency has reconsidered the proposed Data Reporting Guideline
with respect to Subdivision F reporting requirements (§80-4 and §83-3),
Good Laboratory Practice Standards (40 CFR, Part 160), and the proposed
Health Assessment Guidelines for Suspect Developmental Toxicants, and
appropriate revisions have been made.
In order to acccroodate the variety of formats used for conputerized data
recording and analysis, the use of example tables has been limited to a
generalization of commonly submitted formats. The examples should not be
considered as r-equired or preferred, but they represent the most frequently
submitted and accepted format.
In addition, the revised Data Reporting Guideline includes citations
of appropriate sections of the three Aqency sources mentioned above to
avoid extending or repeating existing requirsnents or Guidelines.
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&IDELINE
The Final Report should contain the following items.
COVER PAGE
Cover page and additional documentation requirements (ie. requirements
for data submission, Good Laboratory Practice statement, and procedures
for claims of confidentiality of data), if relevant to the study report,
must precede the content of the study formatted below. These currently
proposed requirements are described in 49 FR (188) 37596 (9/26/84).
TABLE OF CONTENTS
This item should be a concise listing of the essential elements of the
Final Report including the page numbers for each. Essential elements
should include a Sunmary, an Introduction, the Materials and Methods
section, Results, Discussion, Bibliography, Tables, Figures, Appendices,
and key subsections as appropriate.
BODY OF THE REPORT
This item shall include all information required in S80-4(b)(2),
§80-4(c), and §83-3(h) of this subdivision. Additional reccmnendations
are provided in paragraphs identified as III and IV of this addendum.
I. SUMMARY
As per S80-4 (b)(l) of this Subdivision, this section of the test
report shall contain a summary and analysis of the data, and a state-
ment of the conclusions drawn fron the analysis. The summary should
highlight any and all positive data or observations, and any deviations
fron control data which may be indicative of toxic effects. The
summary should be presented in sufficient detail to permit independent
evaluation of the results.
II. INTRODUCTION (include the objectives of the study)
III. MATERIALS/METHODS
This section of the Final Report shall include information required
by §80-4 of this Subdivision, and it may also contain the following
descriptions specific to developmental toxicity studies:
A. Test Animals. Include pre-test mating or insemination procedures;
B. Observations. Include descriptions of the type, frequency and
duration in accordance with §83-3(g)(7) for maternal toxicity and
developmental toxicity end points as required in §83-3{g) as well
as descriptions of the methods for fetal examinations (e.g.,
Staples' technique, Wilson's technique, etc).
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C. Rationale for deviations fran standard protocol. A description
of the rationale for deviations frcm the protocol recommended in
§83-3 should be provided along with rationale for such changes;
D. Evaluation Procedures. The Final Report should describe:
1. Definitions used by the investigators for terms such as mal-
formation, variation, alterations, etc., which are used to
classify fetal changes;
2. Observations used to calculate percentages, weight changes, or
other indices (e.g., number of affected litters = no. with
resorptions + no. with dead fetuses + no. with malformed
fetuses; see paragraph (d) of this addendum for additional
examples);
3. Methods for adjusting data (censoring, pooling, or transforming
results) prior to statistical analysis; and
4. Statistical procedures shall be described according to the
recommendations of §80-4(c) of this Subdivision.
IV. RESULTS
Example formats in this section are generalized frcm those commonly
submitted to the Agency. They do not represent required or preferred
formats, but they are provided as a guide.
A. Summary Tables. Paragraph 83-3(h) and §80-4 require results to be
summarized in tabular form. Example Formats 1-4 show common presen-
tations of summary data that are accepted by the Agency.
1. Section §83-3 of this Subdivision does not mention counting
implantation sites, but that parameter is described in the
Example Formats and in the proposed Health Assessment Guide-
lines mentioned in the Introduction §B to this addendum.
Reports received by the Agency routinely include that parameter,
and it is necessary for calculation of such indices as ore-
implantation loss or percent pre-implantation loss.
2. The proposed Health Assessment Guidelines cited in the Intro-
duction §B to this addendum indicate the observations and
indices to be used in the Agency's assessment of results from
developmental toxicity studies. With the addition of implan-
tation data to the observations required under §83-3, any or
all of the end points described in the proposed assessment
guidelines may be calculated frcm the required observations
and included in a Final Report as shown in Example Formats 1
through 4.
B. Individual Animal Data. These data may be presented as in Example
Formats 5 through 8 and appended in the Final Report.
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V. DISCUSSION
VI. BIBLIOGRAPHY
This item should contain a list of references cited in the body of the
report.
VII. VERIFICATION
This item shall contain information required by $80-4(b)(2). Each
test report shall be:
A. Signed by each of the senior scientific personnel, including the
laboratory director, responsible for performing and supervising
the testing, preparing, reviewing, and approval of the test
report; and
B. Certified by the applicant or an authorized agent of the applicant
as a complete and unaltered copy of the report provided by the
testing laboratory whether independent or owned, operated, or
contrpiled by the applicant.
VIII. ARCHIVES
This section of the Final Report shall contain all information required
in 40 CFR Part 160. 185.
IX. TABLES/FIGURES
X. APPENDIX(ES)
These should include individual animal data, historical control data,
pathology report, analytical nethod and results of analyses on the
test substance and test diet (if the test material is administered via
the diet), details of statistical analyses, protocol, and other infor-
mation as appropriate.
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Example Format 1
Exanple of frequently submitted summary table presentations
of seme maternal and developmental toxicity end points
Observation
Clinical sign A
Clinical sign B
. Dose (mg/kg/day)
__i —
X
*
*
X
N
Y
X
•
*
X
N
5Y
X
*
*
X
N
10Y
X
•
•
X
N
Clinical sign Z
Number of animals
Number pregnant; number died before,
during, and after dosing; totally re-
sorbed litters; abortions; litters with
dead fetuses; .number not pregnant; live
litters at cesarean section:
Number of litters with resorptions;
Corpora lutea/dam; implantations/dam;
pre- implant at ion losses/litter*;
resorpt ions/litter (early, late, and
total)*; mean % resorptions or dead
conceptuses/litter*: live fetuses/
litter*; live male fetuses/litter*;
number live female fetuses/litter*;
sex ratio (males/females) /litter*;
dead fetuses/litter*; uterine weight
(g); mean lire fetal weight (g);
Mean Mean Mean Mean
S. D. S. D. S. D. S. D.
N N N N
*These parameters should also be presented on a total per group basis
07
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Example Format 2
submitted summary table format for
Observation
Body weight (g) at
Day 0
Each treatment
day when weights S. D.
are obtained
Day of sacri-
fice
Gravid uterine
weight (g)
Corrected body
weight (subtrac
gravid uterine
weight fron day
of sacrifice
weight)
Body weight
change, food
and water con-
sunption
nose (maAq/day)
-- Y 5Y
-
Mean
S. D.
N
Mean Mean
S. D. S. D.
N N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean15
S. D.
N
-• w.
Mean"
S. D.
Meanb
S. D.
N
Meanb
S. D.
N
Meanb
S. D.
i M
( N
Mean
S. D.
N
^Throughout gestation, during treatment (increments of
time between measurements), post-treatment to sacrifice,
corrected body weight change throughout gestation (minus
gravid uterine weight or litter weight at sacrifice).
^Means that are statistically significantly different from
control group means should be highlighted.
08
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Example Format 3
Example of a frequently submitted summary table presentations
of fetal effects (ie., malformations, variations, alterations
or other designations used by various investigators)*
Observation 0_
External Alterations
No. fetuses or litters examined N
Pose (mqAq/day)
V 5Y 10Y-
Alteration A
Alteration B
X
X
N
X
X
N
X
X
N
X
X
X
Alteration Z
Soft Tissue Alterations
No. fetuses or litters examined N
Alteration A
Alteration B
X
X
N
X
X
N
X
X
N
X
X
Alteration 2 x
Skeletal Alterations
No. fetuses or litters examined N
Alteration A
Alteration B
X
X
N
X
X
N
X
X
N
X
X
Alteration Z x A A
*These results are presented on a per litter and/or a per group basis.
They are also expressed as percentages in some reports.
09
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Exanple Format 4
Presentation of sane optional maternal toxicity end points
such as liver weight8 and weight ratios (% body weight)
.. . . Liver wt as %
Dosee
(mg/kg/day)
0
y
5Y
. ."•'
10Y
Body
Day 0
Mean
S. D.
N
Mean
S. p.
N
Mean
S. D.
N
Mean
S. D.
N
weight (g) at
Sacrifice
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
body
Day 0
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
Mean
S. D.
N
wt on
Sacrifice
Meanb
S. D.
N
Meanb
S. D.
N
Mean13
S. D.
N
Mean0
S. D.
N
aMay include other organs as appropriate.
tMeans that are statistically significantly different from control
group means should be highlighted.
Example Format 5
Example of a format for reporting individual
survival and toxic signs results
Dose Animal Day of death Toxic signs
group no. or sacrifice Onset day Duration Description
Y - 1 X X X Brief description of
changes in behavior,
appearance, or other
clinical signs.
2 X XX
3 X XX
10
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Example Format 6
An example of a format for reporting jjrdividual
maternal body weight data
Dose
group
Y
Animal
no.
1
2
3
Body
0
X
X
X
weight
6**
X
X
X
(g) on
g***
x
x
X
Gestation Day*
15t
• * * A
• * * X
» • » X
20tt
X
X
X
N
Mean Mean Mean ... Mean Mean
S. D. S. D. S. D. ... S. D. S. D.
N N N .., N N
*For food consumption data these would be increments such as
Days 0-6 (prior to treatment); 6-9, 9-12, 6-16, etc (during
treatment); 6-16 (during treatment); 0-20 (throughout gestation);
etc.
**First treatment day
tLast treatment day
***Weighing days during treatment
ttDay of sacrifice
Example Format 7
Example format for presentation of individual animal data
for pregnancy status and litter results
Dose
Group
Y
Animal
no.
1
2
3
n
Corpora
lutea
X
X
X
*
*
X
implan-
tations
X
X
X
X
P re- implanta-
tion losses
X
X
X
X
Resorptions
Early
X
X
X
X
Late
X
X
X
*
*
*
X
Total
X
X
X
*
X
Dose
Group
Y
Animal
no.
1
2
3
•
»
•
n
Dead
fetuses
X
X
X
•
*
•
X
Live fetuses
Hales
X
X
X
•
•
•
X
Females
X
X
X
•
•
•
X
Total
X
X
X
•
•
•
X
Average fetal
weight (g)
X
X
X
•
•
•
X
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Dose
Example Format 8
Example format for presentation of individual animal data
for external, soft tissue, and skeletal fetal defects
Description of defect(s)
External alterations
Skeletal alterations (if skeleton was examined
Soft tissue alterations (if fetus examined for
soft tissue effects)
External alterations
Skeletal alterations (if skeleton was examined
Soft tissue alterations (if fetus examined for
soft tissue effects)
B
External alterations
Skeletal alterations (if skeleton was examined
Soft tissue alterations (if fetus examined for
soft tissue effects)
n
B
External alterations
Skeletal alterations (if skeleton was examined
Soft tissue alterations (if fetus examined for
soft tissue effects)
External alterations
Skeletal alterations (if skeleton was examined
Soft tissue alterations (if fetus examined for
soft tissue effects)
External alterations
Skeletal alterations (if skeleton was examined
Soft tissue alterations (if fetus examined for
soft tissue effects)
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