UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

WASHINGTON, D.C.  20460

OFFICE OF

PREVENTION, PESTICIDES, AND

         TOXIC SUBSTANCES

TXR No.:	0053835

MEMORANDUM

DATE:		November 3, 2005

SUBJECT:	FOMESAFEN: Second Report of the Cancer Assessment Review
Committee

PC Code:  123802

FROM:	Jessica Kidwell, Executive Secretary

		Cancer Assessment Review Committee

		Health Effects Division (7509C)

TO:		Robert Mitkus/Lisa Austin/William Greear, Toxicologists (RAB1)

		Toiya Goodlow/Mike Metzger, Risk Assessors (RRB1)

		Health Effects Division (7509C)

		James Stone/Joanne Miller, Product Manager

		Herbicide Branch, Registration Division (7505C)

The Cancer Assessment Review Committee met on September 14, 2005 to
re-evaluate the carcinogenic potential of Fomesafen.  Attached please
find the Final Cancer Assessment Document.

cc:	J. Pletcher

	Y. Woo

EVALUATION OF THE CARCINOGENIC POTENTIAL OF

FOMESAFEN (SECOND REVIEW)

PC Code 123802

November 3, 2005

CANCER ASSESSMENT REVIEW COMMITTEE

HEALTH EFFECTS DIVISION

OFFICE OF PESTICIDE PROGRAMS

DATA PRESENTATION:
______________________________________________

             Robert Mitkus/Lisa Austin/William Greear, Toxicologists

DOCUMENT PREPARATION:
_____________________
       Jessica Kidwell, Executive Secretary

COMMITTEE MEMBERS IN ATTENDANCE:(Signature indicates concurrence with
the assessment unless otherwise stated).

Karl Baetcke					                                 _____________________

Lori Brunsman, Statistician
    _________

William Burnam, Chair			_
_________

Marion Copley				                                 _____________________

Kit Farwell					___     						__

Abdallah Khasawinah				______
   ___

Nancy McCarroll				      					________

Tim McMahon				      					________

Esther Rinde					     				(Note pg. 49)___

Jess Rowland					         					________

Linda Taylor					     					________

Yin-Tak Woo					      					________

NON-COMMITTEE MEMBERS IN ATTENDANCE: (Signature indicates concurrence
with the 								pathology report)

John Pletcher, Consulting Pathologist	        					________

OTHER ATTENDEES:   Margy Takacs, Karen Whitby (HED/RAB1), Whang Phang
(HED/RRB1), P.V. Shah (HED/RAB1), Pramod Terse (HED/RAB1), Mike Metzger
(RRB1), Toiya Goodlow (HED/RRB1)

TABLE OF CONTENTS

 TOC \f

EXECUTIVE SUMMARY	1

I.  INTRODUCTION	3

II.  BACKGROUND INFORMATION	3

III. MODE OF ACTION ANALYSIS OF
FOMESAFEN………………………………………..6

IV. WEIGHT-OF-THE-EVIDENCE
CONSIDERATIONS…………………………………….35

V.   CLASSIFICATION OF CARCINOGENIC
POTENTIAL………………………………….37

VI. QUANTIFICATION OF CARCINOGENIC
POTENTIAL……………………………….. 37

VII.
REFERENCES……………………………………………………
………………………..38

APPENDIX
1……………………………………………………………
……………………….41

APPENDIX
2……………………………………………………………
……………………….45

APPENDIX
3……………………………………………………………
……………………….47

APPENDIX
4……………………………………………………………
……………………….48

EXECUTIVE SUMMARY

On September 14, 2005, the Cancer Assessment Review Committee (CARC) of
the Health Effects Division of the Office of Pesticide Programs met to
re-evaluate the cancer classification of fomesafen.  Fomesafen was
previously classified as a Category C oncogen (possible human carcinogen
with limited evidence of carcinogenicity in animals in the absence of
human data) in 1986.  This classification was based on liver tumors
(adenomas, carcinomas, and adenomas/carcinomas combined) at several dose
levels in both sexes of CD-1 mice, some limited evidence for
mutagenicity, and SAR.  Subsequently, the registrant submitted new
information in support of a proposal of activation of peroxisome
proliferator-activated receptor alpha (PPAR) as the mode of action
for fomesafen-induced liver tumors.

The CARC met on 9/14/05 to evaluate this hepatocarcinogenic mode of
action for fomesafen.  Robert Mitkus and Lisa Austin of Registration
Action Branch 1 presented previous and recent data submitted by the
registrant proposing activation of peroxisome proliferator-activated
receptor alpha (PPAR) (also referred to as peroxisome proliferation)
as the mode of action for fomesafen.  The evidence included the
following:

	1.	Individual, new studies that address certain aspects in the proposed
mode of action for mouse liver tumors (Brady 1998; Moffat 1998; Moffat
and Townsley 2004; Peffer 2004; Roberts 2001)

	2.	Additional, new studies on genotoxicity and an overall assessment of
genotoxicity. (Elliott 1998; Howard and Richardson 1998; Mellano and
Berruto 1984; Sheldon et al. 1988)

 activation and toxicokinetics.

 tc "V. 	SUMMARY"

 activation and toxicokinetics.  The quantification of risk is not
required.

I.     INTRODUCTION

 tc "

I.     INTRODUCTION"

On September 14, 2005, the Cancer Assessment Review Committee (CARC) of
the Health Effects Division of the Office of Pesticide Programs met to
evaluate the hepatocarcinogenic mode of action of fomesafen.  This was a
re-evaluation of the cancer classification of fomesafen in light of new
data submitted by the registrant, Syngenta Crop Protection, Inc.

II.   BACKGROUND INFORMATION tc "II.   BACKGROUND INFORMATION"

Fomesafen is used as a post-emergence herbicide.

HISTORY

	The Toxicology Branch Peer Review Committee met on July 24, 1986 to
evaluate the carcinogenic potential of fomesafen (Memo, J. Quest, Peer
Review of Fomesafen, 8/27/86).  A summary of the findings is presented
below.

Mouse Carcinogenicity Study With Fomesafen

Citation: Colley J et al. (1983). Fomesafen 2-year feeding study in
mice. Huntingdon Research Centre, UK. Study No. CTL/C/1207A. Sponsor:
ICI Americas, Inc. Unpublished. MRID 131491.

Fomesafen was administered in the diet to Charles River CD-1 mice for 2
years at dose levels of 0 ppm (128 mice/ sex), 1 ppm (64/mice/sex), 5
ppm (64 mice/sex, 100 ppm (64 mice/sex) and 1000 ppm (64 mice/sex).  An
interim sacrifice period was conducted at 52 weeks involving 24 mice/sex
in control and 12 mice/sex in the treatment groups.  The study was
conducted by Huntingdon Research Center, England.  The tumor table below
(Table 1) is taken from the 1986 Peer Review Report.

Table 1. Tumor Incidence in Mice Following Administration of Fomesafen

Liver Tumors

Sex

0  ppm

1 ppm

5 ppm

100 ppm

1000 ppm

Adenoma	Male	13/110 (12%)	19/56 (34%)b	5/52 (10%)	17/54 (31%)a	14/36
(39%)b

Carcinoma	Male	17/127 (13%)	7/63 (11%)	12/64 (19%)	10/64 (16%)	28/64
(44%)b

Combined	Male	30/127 (24%)	26/63 (41%)	17/64 (26%)	27/64 (42%)	42/64
(66%)b

Adenoma	Female	3/128 (2%)	1/63 (1%)	1/62 (1%)	8/62 (13%)a	12/48 (25%)b

Carcinoma	Female	0/128 (0%)	1/64 (1%)	2/64 (3%)	2/64 (3%)	16/64 (25%)b

Combined	Female	3/128 (2%)	2/64 (3%)	3/64 (5%)	10/64 (16%)a	28/64 (44%)b

  a=p<0.01 compared to controls, Fisher’s Exact Test

b=p<0.001 compared to controls, Fisher’s Exact Test

Rat Carcinogenicity Study With Fomesafen

Citation: Milburn G et al. (1984). Fomesafen: 2-year feeding study in
rats. Central Toxicology Laboratory, UK. Study No. CTL/P/863. Sponsor:
Imperial Chemical Industries PLC. Unpublished. MRID 142125.

Fomesafen was administered in the diet to Wistar (Alderley Park) albino
SPF rats for 106 weeks.   Groups of 52 males and 52 females were fed 0,
1, 5, 100 or 1000 ppm for 106 weeks (terminal sacrifice) and additional
sacrifice.  The study was conducted by ICI Central Toxicology
Laboratory.

	No treatment-related tumors were seen in male or female Wistar rats.

Weight-of-the-Evidence Considerations

The weight-of-the-evidence considerations from the 1986 Peer Review
meeting were as follows:

1. Fomesafen when administered in the diet to Charles River CD-1 mice
was associated with significantly elevated incidences of liver tumors
(adenomas, carcinomas, and adenomas/carcinomas combined) in males and
females.  There was evidence, however, for a progression of benign
tumors to malignancy, and for a reduction in the latency period for the
time-to-tumor appearance.  There was no evidence for the occurrence of
hyperplastic changes in the livers of treated mice.

2. Liver tumors were seen in male mice at dose levels (i.e., 100 and
1000 ppm) that seemed to exceed a MTD level, and also at a dose level
(i.e., 1 ppm) that was below a MTD level.  In addition, liver tumors
were seen in female mice at a dose level (i.e., 1000 ppm) that seemed to
exceed a MTD level, and also at a dose level (i.e., 100 ppm) that
approximated a MTD level.

3. Fomesafen was not carcinogenic when administered in the diet to
Wistar albino rats at doses ranging from 1 to 1000 ppm.  The highest
dose tested in males (1000 ppm) in the chronic rat bioassay exceeded a
MTD level, whereas this dose in females approximated the MTD.

4. The primary target organ of toxicity of Fomesafen in various
non-chronic studies in several species other than mice (e.g., subchronic
tests in rats and dogs, reproduction study in rats) was also the liver.
Evidence of hepatocellular toxicity was also seen in the rat chronic
bioassay of Fomesafen (where no tumors were observed).

5. No metabolism studies of Fomesafen were performed in mice.  Studies
conducted in rats, however, demonstrated a preferential concentration of
the compound in the liver to the exclusion of other tissues.  This
finding is consistent with the liver being a target organ for fomesafen
toxicity.

6. Fomesafen is not considered to be mutagenic.

7. Fomesafen bears a structural resemblance to other substances (e.g.,
lactofen, acifluorfen) that have been demonstrated to be hepatocellular
carcinogens in mice.

8. Fomesafen did not evoke adverse reproductive effects in rats or mice
and was not teratogenic in rats or rabbits.

1986 Classification of Carcinogenic Potential

The Committee concluded that the data available for Fomesafen provided
limited evidence of carcinogenicity for the chemical in male and female
CD-1 mice.  According to EPA proposed guidelines (CFR, November 23,
1984), the Committee classified Fomesafen as a Category C oncogen
(possible human carcinogen with limited evidence of carcinogenicity in
animals in the absence of human data).  That is, Fomesafen produced
liver tumors (adenomas, carcinomas, and adenomas/carcinomas combined) at
several dose levels in both sexes of CD-1 mice in a single experiment.
In addition, there was some limited evidence for the mutagenicity of
Fomesafen, and the compound was structurally related to known oncogens.
The Committee also considered criteria for classifying a carcinogen in
the B2 category, but Fomesafen did not fully meet the criteria specified
for this classification.  The criteria included the following: 1)
fomesafen did not produce tumors in multiple species or strains; 2) it
did not produce tumors in multiple experiments; and 3) it did not
produce tumors to an unusual degree with regard to incidence or tumor
site.   There was a reduced latency period for the time to appearance of
tumors in mice, but the Committee did not consider this finding to be of
sufficient weight to elevate Fomesafen from the C to the B2 category,
particularly since the mouse liver tumor was the only carcinogenic
response observed.

REGULATORY HISTORY

Subsequently, the registrant (Syngenta) submitted a registration (DP
Barcode No. D311247) requesting the cancer reclassification for
fomesafen (from Thomas J. Parsley, Syngenta to Ms. Joanne Miller,
Product Manager, RD,  MRID No. 46527200 dated April 15, 2005).  The
registration included new studies as well as summarized earlier data
submissions, which provided the information to support the proposal of
activation of PPAR as the mode of action of Fomesafen in inducing
liver tumor in mice.

III. 	MODE OF ACTION ANALYSIS OF FOMESAFEN

Introduction

There have been two approaches that the Health Effects Division (HED) of
OPP has used to evaluate a putative mode of action involving peroxisome
proliferation.  The first is based on Cattley et al. (1998), in which a
“minimum database” is established to demonstrate that
hepatocarcinogenesis takes place by way of a mode of action involving
peroxisome proliferation.  The three key criteria needed to establish
such a database are hepatomegaly (enlarged liver), peroxisome
proliferation, and increased DNA synthesis following exposure to a
non-genotoxic hepatocarcinogen.  These endpoints are measured by
increases in relative liver weight, number or volume of hepatic
peroxisomes, and hepatic BrdU labeling, respectively (Cattley et al.
1998).  The HED Mechanism of Toxicity Assessment Review Committee
(MTARC) utilized this approach in its evaluations of the herbicides
diclofop-methyl, acifluorfen, and lactofen (TXR#s 0014172, 0052006, and
0051907, respectively).

The second approach is articulated in the recent Final Guidelines for
Carcinogen Risk Assessment (USEPA 2005), which follows the
decision-making framework developed by the International Programme on
Chemical Safety (IPCS) described in detail by Sonich-Mullin et al.
(2001).  While Syngenta Crop Protection, Inc. has submitted its own mode
of action analysis that uses the IPCS framework, the purpose of this
section is to present and evaluate the original data submitted by
Syngenta Crop Protection, Inc. in support of its postulated mode of
action for fomesafen-induced hepatocarcinogenesis.  These data will be
evaluated using the Agency Guidelines for Carcinogen Risk Assessment
(2005), as well as the work of Klaunig et al. (2003).  Table 2 lists the
key events necessary, according to current scientific understanding, to
establish that a chemical causes hepatocarcinogenesis by way of a mode
of action involving peroxisome peroliferation.

 ligands (Permission requested from the publisher).

Summary description of hypothesized mode of action

Oral exposure of mice to fomesafen causes activation of peroxisome
proliferator-activated receptor alpha (PPAR) in hepatocytes, leading
to disruption of cell proliferation and apoptosis, followed by selective
clonal expansion of initiated cells.

Key events

In addition to exposure to fomesafen, activation of PPAR, increased
cell proliferation (as measured by DNA synthesis), decreased apoptosis,
clonal expansion, and the appearance of liver tumors, two other events
are viewed as key to the mode of action of fomesafen-induced
hepatocarcinogenesis in mice.  These are peroxisome proliferation,
defined as an increase in the volume density of peroxisomes and an
increase in peroxisomal enzyme activity (IARC 1995; Klaunig et al.
2003), and increased expression of peroxisomal genes (e.g.,
CN-insensitive palmitoyl CoA oxidase or catalase).

Temporal Association

 wild-type mice after 24 hours of in vitro exposure to fomesafen.
These effects were not observed in hepatocytes extracted from
PPAR-null mice, thereby supporting the role of the receptor in
mediating these effects.  However, the corresponding concentration of
fomesafen needed in an in vivo feeding study to achieve the same in
vitro tissue concentration after 24 hrs. was not calculated.  For this
reason, activation of PPAR was not included in the temporal sequence
of key events.

    Oral Exposure of mice to Fomesafen

1 Week 		Increased absolute/relative liver weight, liver hypertrophy,
hepatocytic palmitoyl CoA oxidation, cell proliferation (DNA synthesis);
(≥100 ppm)

4 Weeks	Increased absolute/relative liver weight, liver hypertrophy,
hepatocytic peroxisomal volume density and palmitoyl CoA oxidation
(≥10 ppm), cell proliferation (DNA synthesis); (≥100 ppm)

8 Weeks	Increased absolute/relative liver weight, hepatocytic
hypertrophy, hepatocytic palmitoyl CoA oxidation, cell proliferation
(DNA synthesis); (≥100 ppm)

52 Weeks	Increased absolute liver weight, liver hypertrophy; (≥100
ppm)

79/89 Weeks	Increased absolute liver weight, liver hypertrophy, liver

(M/F)				tumors; (≥100 ppm)

Figure 1. Temporal sequence of events in the postulated mode of action
of fomesafen-induced liver tumor formation.

Dose-response Concordance

 and peroxisome proliferation.  However, a dose-dependent increase in
the incidences of adenomas and combined adenomas/carcinomas was observed
after 104 weeks in both sexes from 10-1000 ppm fomesafen, as well as
increased mean absolute liver weights in both sexes at the 52-week
interim sacrifice time point at 10-1000 ppm fomesafen.  A clear
threshold in the development of carcinomas was observed in both sexes at
1000 ppm fomesafen.  Endpoints that exhibited dose-dependent increases
from 0-1000 ppm fomesafen (the entire dose range) were mean absolute
liver weights at 52-weeks and termination (week 79 or 104, depending on
dose) in males only and enlarged liver (hepatomegaly) in males only at
termination.

The incidence of adenomas and combined adenomas/carcinomas in males at 1
ppm fomesafen was higher than control values, but also higher than at 10
ppm.  In Peffer (2004), the study author stated that this increase was
“isolated… [and] sporadic” due to a high background incidence of
adenomas in CD-1 mice.  However, the historical control range for
adenomas in

Table 3. Key events measured in the 2-year bioassay in CD-1 mice
(Colley et al. 1983)

Concentration (ppm)

[Dose (mg/kg BW/day; M/F)]	Absolute Liver Weights (52-wk. interim
sacrifice)1	Absolute Liver Weights [79 (M) or 89 (F) weeks]1,2	Absolute
Liver Weights (104 weeks)1,2	Enlarged liver

(52-wk. interim sacrifice)3,4	Enlarged liver

(Termination)3,4,5	Neoplasms (Quest 1986)3

	Adenoma	Carcinoma	Combined

	M	F	M	F	M	F	M

	F	M

	F	M	F	M	F	M	F

0 [0/0]	2.2±0.3	1.8±0.4	3.5±2.6	1.7±0.5	2.7±0.9	1.9±0.5	0/23	0/24
7/46 (15)	6/50 (12)	13/110 (12)	3/128 (2)	17/127 (13)	0/128 (0)	30/127
(24)	3/128 (2)

1 [0.10/0.10]	2.3±0.6	1.9±0.3	ND	ND	2.8±1.2	1.7±0.3	0/12	0/10	4/27
(15)	3/19 (16)	19/56 (34)***,6	1/63

(2)	7/63 (11)	1/64

(2)	26/63 (41)	2/64

(3)

10 [0.99/1.08]	2.5±0.4	1.8±0.4	ND	ND	2.9±0.6	2.0±0.6	0/11	0/12	5/19
(26)	4/18 (22)	5/52 (10) 	1/62

(2)	12/64 (19)	2/64

(3)	17/64 (27)	3/64

(5)

100 [9.96/10.67]	4.0±0.5***	2.5±0.8*	ND	ND	4.3±1.4**	2.9±1.3**	5/9
(56)	3/10 (30)	12/15 (80)	19/25 (76)	17/54 (31)**	8/62 (13)**	10/64 (16)
2/64

(3)	27/64 (42)	10/64 (16)**

1000 [121.08/115.04]	8.1±4.0***	4.9±2.6***	9.2±2.6***	4.6±2.2***	ND
ND	9/9 (100)	7/11 (64)	11/11 (100)	15/15 (100)	14/36 (39)***	12/48
(25)***	28/64 (44)***	16/64 (25)***	42/64 (66)***	28/64 (44)***

ND - No data

* p<0.05

** p<0.01

*** p<0.001

1 There was no statistically significant increase in body weights at any
dose

2 Termination took place at weeks 79 (males) or 89 (females) for the
1000 ppm group (and an equal number of control mice) and at week 104 for
groups treated with 1-100 ppm

3 Percent incidence in parentheses

4 Statistics not performed

5 Termination took place after 79 (M) or 89 (F) weeks of treatment for
the high-dose group and after 106 weeks for all other groups

6 This increase is outside of the laboratory’s historical control
range: 1.6%-23.4% (Calderbank 1988)

male CD-1 mice was reported by the study author as 1.6%-23.4%
(Calderbank 1988).  The incidence of adenomas in males at 1 ppm was 34%
and clearly outside of the historical control range.  The effect was
also significantly increased over controls (P<0.001).

Essential to any mode of action analysis is establishing whether the
dose-response relationship for any key step in the postulated mode of
action parallels that of other key steps.  This is known as establishing
dose-response concordance.  Ideally, key events should occur in a
dose-dependent manner such that “the key [precursor] events forecast
the appearance of tumors at a later time or higher dose” (USEPA 2005).
 The significantly increased incidence of adenomas and the
non-significantly increased incidence of combined adenomas/carcinomas at
1 ppm in males weakens the proposed mode of action, because tumors
appear at 1 ppm, when the incidences of all measured precursor events
are at or near control values.

	In general, the measured precursor events occur at the same doses as
the tumor response (except for 1 ppm, as mentioned above).  This fact
strengthens the dose-response concordance for the 2-year feeding study
in CD-1 mice.  For example, significantly increased (P<0.01) tumor
responses occurred in either sex at concentrations of 100 or 1000 ppm
only.  These are the same concentrations at which significant (P<0.05)
increases in mean absolute liver weight and enlarged liver (statistics
not performed) were observed.

	Tables 4a-c summarize the key events measured in two short-term (28-
and 56-day) studies carried out in CD-1 mice exposed to 0, 1, 10, 100,
1000, or 3000 ppm fomesafen.  Although the length of the studies was too
short to measure a tumor response and not all concentrations were tested
in each study, two of the other causal key events of the peroxisome
proliferation mode of action were measured and observed in CD-1 mice in
both studies, i.e., changes in cell proliferation and apoptosis, as well
as peroxisome proliferation (volume density and peroxisomal enzyme
activity).  Activation of PPAR was not measured in either study.
Increased liver weight and liver hypertrophy, which are non-causal key
events associated with peroxisome proliferation, were observed as early
as 7 days post-treatment at 100 ppm.

e density of peroxisomes was observed after 28 days of treatment
(peroxisome proliferation was not measured after 7 or 56 days).  The
effect was seen at ≥100 ppm fomesafen.  A dose-dependent increase in
palmitoyl CoA oxidation (PCO), a marker of peroxisomal enzyme activity
and gene expression, was also observed after 7, 28, and 56 days of
treatment with fomesafen.  Statistically significant increases were
observed at 10-1000 ppm fomesafen on day 28 and at 100-3000 ppm on days
7 and 56 (Table 4c).  In the first experiment, 3000 ppm fomesafen was
not tested after 28 days of treatment, whereas in the second experiment,
in which effects were measured after 56 days, 10 ppm was not tested.

Table 4a. Key events measured in the 28- and 56-day studies in CD-1
mice (Moffat 1998)

Absolute Liver Weights

Concentration (ppm)1	   Experiment 1	     Experiment 2	   Experiment 1
 Experiment 2

	Day 7	Day 7	Day 28	Day 56

	M	F	M	F	M	F	M	F

0	1.80±0.09	1.30±0.17	1.97±0.17	1.40±0.22	1.99±0.20	1.55±0.20
2.11±0.23	1.61±0.24

1	2.03±0.19	1.41±0.15	ND	ND	2.01±0.16	1.56±0.30	ND	ND

10	2.05±0.11	1.37±0.20	ND	ND	2.01±0.17	1.56±0.22	ND	ND

100	2.73±0.24***	1.60±0.36	2.63±0.34**	1.67±0.29	2.87±0.32**
1.79±0.32	2.80±0.34	2.10±0.63

1000	3.15±0.42***	2.14±0.38***	3.60±0.38***	2.25±0.52**
3.24±0.80***	2.63±0.51***	5.16±0.97***	2.72±0.36*

3000	ND	ND	3.73±0.28***	2.99±0.37***	ND	ND	5.92±0.74***	5.25±1.32***

Relative Liver Weights

Concentration (ppm)	   Experiment 1	     Experiment 2	   Experiment 1
Experiment 2

	Day 7	Day 7	Day 28	Day 56

	M	F	M	F	M	F	M	F

0	5.74±0.25	5.13±0.29	4.82±0.18	4.67±0.38	5.23±0.13	5.07±0.29
4.84±0.35	4.66±0.57

1	5.95±0.56	5.42±0.48	ND	ND	5.34±0.27	5.16±0.31	ND	ND

10	6.36±0.34	5.32±0.28	ND	ND	5.66±0.27	5.19±0.16	ND	ND

100	8.27±0.63***	6.17±0.90*	6.80±0.54***	5.56±0.70	7.41±0.76***
5.64±0.55	6.35±0.47	6.02±1.40

1000	9.76±0.76***	7.92±1.28***	9.17±0.46***	7.36±1.14**
9.32±1.22***	8.35±1.20***	11.22±1.70***	7.35±1.18*

3000	ND	ND	10.28±0.98***	9.63±1.30***	ND	ND	13.03±1.58***
14.46±2.47***

ND – no data

1Equivalent doses (mg/kg bodyweight/day) not provided

*p<0.05

** p<0.01

*** p<0.001

Table 4b. Key events measured in the 28- and 56-day studies in CD-1 mice
(Moffat 1998)

Liver Histology

Concentration (ppm)	Experiment 1	Experiment 2

	Day 7	Day 28	Day 56

	Centrilobular hypertrophy	Centrilobular hypertrophy	Panlobular
hypertrophy/eosinophilia	Centrilobular eosinophilia	Panlobular
eosinophilia	Hepatocyte hypertrophy	Panlobular vacuolation	Centrilobular
atrophy

	M	F	M	F	M	F	M	F	M	F	M	F	M	F	M	F

0	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-

1	-	-	-	-	-	-	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND

10	-	-	-	-	-	-	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND

100	++	++	+++	++	-	-	+++	+++	++	-	++	++	-	-	-	-

1000	+++	++++	+++	+++	++++	+++	-	++	+++	+++	+++	+++	++	-	-	-

3000	ND	ND	ND	ND	ND	ND	-	-	++++	++++	++++	+++	+++	++	+++	++

ND – no data

- not observed

++ slight

+++ moderate

++++ marked

Table 4c. Key events measured in the 28- and 56-day studies in CD-1
mice (Moffat 1998)

Concentration (ppm)	Peroxisomal Volume

(% cytoplasm)

	Palmitoyl CoA Oxidation

(nmol NAD+ reduced/mg protein)	Cell Proliferation

(% BrdU labeling)

	Experiment 1	Experiment 1	Experiment 1

	Day 28	Day 7	Day 28	Day 7	Day 28

	M	F	M	F	M	F	M	F	M	F

0	2.03±0.49	1.47±0.19	16.5±3.1	15.5±1.7	25.5±4.0	30.3±2.4
1.66±1.48	9.08±2.86	0.91±0.42	1.80±0.79

1	2.51±0.58	1.91±0.47	16.3±2.6	16.4±3.8	27.9±3.3	32.9±5.8
2.40±1.56	7.61±5.20	2.67±1.62	2.85±2.16

10	3.83±1.35	3.02±0.77	22.5±4.6	17.9±5.1	107.0±18.9***	84.6±16.3*
1.27±0.42	7.42±4.07	2.36±1.20	2.65±2.61

100	16.56±9.33***	13.76±12.76**	85.3±2.7***	61.1±20.4	219.3±40.5***
114.8±62.9**	7.95±4.09**	8.11±3.03	4.35±2.84	8.74±9.51*

1000	24.37±4.26***	18.43±8.06**	97.8±16.1***	119.7±67.9***
241.0±23.8***	253.5±39.1***	18.75±4.03***	23.15±14.03**
16.08±5.59***	3.53±2.50

3000	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND

Concentration (ppm)	Peroxisomal Volume

(% cytoplasm)

	Palmitoyl CoA Oxidation

(nmol NAD+ reduced/mg protein)	Cell Proliferation

(% BrdU labeling)

	Experiment 2	Experiment 2	Experiment 2

	Day 56	Day 7	Day 56	Day 7	Day 56

	M	F	M	F	M	F	M	F	M	F

0	ND	ND	12.88±3.51	18.78±3.90	17.58±2.79	17.37±5.47	5.22±4.04
3.79±3.69	1.22±1.01	1.96±1.00

1	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND

10	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND

100	ND	ND	74.94±22.83***	50.08±29.28*	92.27±23.94***	63.13±28.09**
10.49±3.56	7.81±5.19	3.34±2.93	0.80±0.38

1000	ND	ND	117.18±9.52***	88.56±9.04***	146.94±37.61***
111.33±33.69***	19.18±9.59**	19.60±9.57**	8.45±3.12*	2.07±2.06

3000	ND	ND	114.76±15.58***	121.90±18.49***	139.66±14.57***
115.26±15.20***	33.60±8.29***	53.30±8.58***	16.23±9.49***
24.40±9.17***

ND - No data

*p<0.05

** p<0.01

*** p<0.001

n most cases, the key events peroxisome proliferation and increased
liver weight and size were statistically significantly increased at the
same doses i.e., ≥100 ppm fomesafen.  This fact strengthens the
dose-response concordance between these two key events, because these
precursor events occurred at the same doses as those at which adenomas
developed.  The results also strengthen the proposed mode of action,
because peroxisome proliferation and increased liver weights occurred at
a lower dose than that at which carcinomas developed.  In addition, the
statistically significant increase in PCO on day 28 at 10 ppm fomesafen
strengthens the proposed mode of action, because this precursor event
occurred at a lower concentration than that at which tumors developed
(≥100 ppm fomesafen).  PCO was not measured after 56 days, however, so
it is not known whether increased PCO activity was sustained.  It should
be noted that panlobular vacuolation and centrilobular atrophy were
observed after 56 days at ≥1000 and 3000 ppm fomesafen, respectively,
thereby suggesting a hepatotoxic effect at these two highest
concentrations.

	The percent increase in bromodeoxyuridine (BrdU) labeling was also
measured after 7, 28, and 56 days of treatment with fomesafen.  BrdU
labeling is a more sensitive indicator of cell proliferation than
macroscopic examination of hepatomegaly, because BrdU labeling measures
the number of cells in S-phase of the cell cycle at a given time,
whereas macroscopic examination cannot.  A dose-dependent increase in
%BrdU labeling was observed after 7, 28, and 56 days of treatment with
fomesafen.  After 28 days, BrdU labeling was increased at 100 ppm in
males, while after 56 days, cell proliferation was significantly
increased at ≥1000 ppm; BrdU labeling was not measured at 3000 ppm
after 28 days.  It should be noted that after 7 days of treatment with
100 ppm fomesafen, a significant increase (p<0.01) in %BrdU labeling was
observed in males.  In general, the data also indicate that cell
proliferation was sustained at 100 ppm in males after 28 days.  However,
the labeling index returned to control levels in females at 1000 ppm,
and %BrdU labeling at 100 ppm fomesafen was not significantly increased
above controls in males or females at 56 days.

According to the proposed mode of action and carcinogenesis in general,
one would expect a decreased, not increased, incidence of apoptosis to
be associated with an elevated tumor endpoint.  However, Figure 2
suggests an increased number of apoptotic bodies in the liver at 3000
ppm, relative to controls, after 56 days of treatment with fomesafen,
although a mean number of apoptotic bodies was not reported for either 0
or 3000 ppm fomesafen.  Due to the lack of animal data for other
timepoints and concentrations, a dose-response concordance analysis
cannot be carried out for this endpoint.

Figure 2. Key event (apoptosis) measured in the 56-day study in CD-1
mice (n=5) (Moffat 1998). AB=apoptotic bodies; 0.1 on x-axis (log scale)
= 0 ppm fomesafen

 WT mouse, PPAR-null mouse, and human hepatocytes were exposed to
0-500 M fomesafen in vitro.  Statistically significant (P<0.05)
decreases in mean percentages of apoptosis, relative to untreated cells,
were observed in PPAR WT mouse hepatocytes at 100 and 250 M
fomesafen (Table 4d).  Statistically significant (P<0.05) increases in
mean percentages of apoptosis were observed in PPAR-null mouse
hepatocytes at 10, 50, and 250 M fomesafen.  No appreciable change in
mean apoptosis levels was observed in human hepatocytes; however,
statistical analysis was not performed on the results for human
hepatocytes.

Table 4d. Key event (apoptosis) measured in PPAR WT and -null mouse
and human hepatocytes exposed to fomesafen for 24 hrs. in vitro (Roberts
2001)

Concentration (uM)	Apoptosis (%)

	Mouse (PPAR WT)	Mouse (PPAR-null)	Human (male, age 4) a

 0 	1.37 ± 0.31	1.70 ± 0.26	3.3 ± 0.1

10	1.53 ± 0.46	2.10 ± 0.26 *	3.1 ± 0.2

50	1.03 ± 0.12	2.17 ± 0.15 *	2.9 ± 0.3

100	0.83 ± 0.15 *	1.90 ± 0.36	3.1 ± 0.4

250	0.67 ± 0.12 *	2.89 ± 0.45 *	3.0 ± 0.6

500	0.87 ± 0.15	2.46 ± 0.89	2.9 ± 0.0

a Statistics not performed.

* Statistically different (p <0.05) from non-treated.

Based on the 2-year cancer bioassay and two short-term bioassays in
mice, there is an overall good dose-response concordance between the
precursor events of the proposed mode of action and the development of
tumors.  This is because all of the in vivo precursor events were first
observed at the same concentrations as those at which tumors developed,
i.e., 100 ppm fomesafen.  In addition, increased PCO at 10 ppm in both
sexes on day 28 further supports causality for the proposed mode of
action, because PCO was observed at a concentration of fomesafen lower
than that at which tumors developed.  Increased cell proliferation at
3000 ppm in females after 56 days, neither strengthens nor weakens the
proposed mode of action, because CD-1 mice were not treated with 3000
ppm fomesafen in the 2-year cancer bioassay.  In addition, the return of
cell proliferation rates to baseline at 1000 ppm in females after 28 and
56 days is consistent with the activity of a mitogen as well as the mode
of action of most other PPAR agonists, for which cell proliferation
returns to baseline levels after 2-4 weeks of treatment (Klaunig et al.
2003).  Based on the entire dose-response concordance analysis, a
reasonable inference of causation can be made regarding the key
precursor events and the tumor response in CD-1 mice.

Table 5 summarizes the key events measured in the 2-year carcinogenicity
bioassay carried out in Alderley Park (AP) rats (Milburn et al. 1984).
Although not all animals were examined for each endpoint, some of the
key events measured in the bioassay in CD-1 mice were also measured in
the rat feeding study.  A significant increase in peroxisome
proliferation (P<0.01), as measured by peroxisome volume as a proportion
of cell cytoplasm, and adjusted (relative-to-bodyweight) liver weight
was observed at 1000 ppm fomesafen after 52 and 104 weeks; however,
peroxisome proliferation was not dose-dependent at 104 weeks, and the
increase in adjusted liver weights was not dose-dependent at either 52
or 104 weeks.  In addition, no liver tumors were reported at any dose in
the rat bioassay.  Because of this, there is no dose-response
concordance between precursor events and tumors in this study in the
rat.

Table 5. Key events measured in the 2-year bioassay in AP rats (Milburn
et al. 1984)

Concentration (ppm)1	Peroxisome Proliferation

(52-wk. interim kill)2,3	Peroxisome Proliferation

(wk. 104)2,3	Adjusted (4) Liver Weights

(52-wk. interim kill)5	Adjusted (4) Liver Weights

(wk. 104)6	Neoplasms

	No liver tumors reported at any dose

	M	F	M	F	M	F	M	F

	0	7.1±3.0 (0/6)	10.9±4.2 (1/6)	5.8±1.1 (0/6)	6.2±1.8 (0/6)	19.5
11.9	22.0 (30)	14.7 (22)

	1	7.7±2.5 (0/6)	11.6±2.9 (0/6)	7.1±1.9 (0/6)	5.5±3.2 (0/6)	20.1
12.5	22.8 (23)	15.2 (24)

	5	11.7±6.7 (1/6)	9.7±5.3 (1/6)	19.0±17.8 (3/6)*	6.9±1.5 (0/6)	19.9
(10)	11.8	20.8 (26)	14.6 (21)

	100	12.3±10.3 (2/5)	12.1±2.8 (1/6)	9.1±2.6 (1/6)**	6.5±2.3 (0/6)
20.9	13.3 (11)	21.0 (34)	16.0 (17)

	1000	30.8±15.1 (6/6)**	48.3±36.7 (6/6)**	36.0±22.7 (5/6)**
40.5±30.0 (5/6)**	23.7 (10)**	13.7 (11)*	24.1 (36)*	16.4 (31)*

* p<0.05

** p<0.01

1 Doses in mg/kg bodyweight/day not reported

2 Expressed as the # of points in a 320-point morphometric analysis grid
coinciding with peroxisomes

3 Incidence of animals showing peroxisomal proliferation in parentheses

4 Adjusted for bodyweights

5 Based on 12 animals, unless noted in parentheses; means adjusted for
missing values

ker for increased protein synthesis), were observed at ≥1000 ppm
fomesafen.  Unlike the results for CD-1 mice, panlobular vacuolation and
centrilobular atrophy were not observed at any dose.

Table 6a. Key events measured in the 56-day study in AP rats (Moffat
1998)

Relative Liver Weights

Concentration (ppm)1	Day 7	Day 56

	M	F	M	F

0	4.66±0.24	4.58±0.21	3.65±0.08	3.81±0.23

1	ND	ND	ND	ND

10	ND	ND	ND	ND

100	5.02±0.21	4.84±0.26	4.58±0.19	3.87±0.35

1000	6.98±0.57***	5.12±0.23**	5.82±0.30**	4.16±0.16

3000	6.36±0.75***	5.30±0.20***	6.22±1.49***	4.80±0.33***

ND – no data

1Equivalent doses (mg/kg bodyweight/day) not provided

*p<0.05

** p<0.01

*** p<0.001

Table 6b. Key events measured in the 56-day study in AP rats (Moffat
1998)

Liver Histology

Concentration (ppm)	Day 56

	Centrilobular eosinophilia	Panlobular eosinophilia	Hepatocyte
hypertrophy	Panlobular vacuolation	Centrilobular atrophy

	M	F	M	F	M	F	M	F	M	F

0	-	-	-	-	-	-	-	-	-	-

1	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND

10	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND

100	+	+	-	-	-	-	-	-	-	-

1000	+++	++	-	-	++	-	-	-	-	-

3000	+++	+++	-	-	+++	++	-	-	-	-

ND – no data

- not observed

+ minimal

++ slight

+++ moderate

Table 6c. Key events measured in the 56-day study in AP rats (Moffat
1998)

Concentration (ppm)	Peroxisomal Volume

(% cytoplasm)

	Palmitoyl CoA Oxidation

(nmol NAD+ reduced/mg protein)	Cell Proliferation

(% BrdU labeling)

	Day 56	Day 7	Day 56	Day 7	Day 56

	M	F	M	F	M	F	M	F	M	F

0	ND	ND	19.24±4.45	21.04±4.29	17.26±4.52	20.52±3.63	5.93±3.85
5.00±1.13	2.01±1.39	4.15±1.62

1	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND

10	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND

100	ND	ND	58.56±16.55**	22.46±4.34	75.77±46.77*	25.55±1.58
11.16±2.34	20.69±11.45*	1.43±1.02	5.56±2.23

1000	ND	ND	105.63±31.58***	48.43±10.32**	100.10±21.96**
57.62±8.35***	25.62±8.57***	28.77±8.66***	2.04±0.74	7.00±4.74

3000	ND	ND	127.25±10.36***	73.73±15.64***	117.65±37.16***
96.88±8.14***	11.33±9.83	22.31±11.03**	17.72±18.73*	8.38±1.91*

ND – no data

*p<0.05

** p<0.01

ed in the 2-year rat feeding study, a dose-dependent increase in
palmitoyl CoA oxidation (PCO) was observed, with significant increases
(p<0.05) occurring in males at ≥100 ppm fomesafen after 7 and 56 days;
significant increases (p<0.01) in females occurred at ≥1000 ppm after
7 and 56 days of treatment with fomesafen.  These results were similar
to those observed in male CD-1 mice.  A dose-dependent increase in cell
proliferation, measured as % BrdU labeling, was also measured in AP rats
after 56 days of treatment with fomesafen, with significant increases
(p<0.05) observed only at 3000 ppm.  After 7 days of treatment, cell
proliferation was significantly increased (p<0.01) at concentrations
≥1000 ppm; however, mean values decreased in both sexes at 3000 ppm,
but remained above those of controls.  Similar to results in CD-1 mice,
apoptosis was tested at 0 and 3,000 ppm fomesafen.  The number of
apoptotic bodies in males after 56 days at 3000 ppm seemed to be
slightly larger than control values (Figure 3), while results for
females indicated no difference from controls; however, summary data
were not provided for this endpoint.

Figure 3. Key event (apoptosis) measured in the 56-day study in AP rats
(n=5) (Moffat 1998). AB=apoptotic bodies; 0.1 on x-axis (log scale) = 0
ppm fomesafen

	Overall, dose-response effects for the key precursor events measured
in AP rats were not strongly concordant.  The doses at which relative
liver weights and liver hypertrophy occurred were similar, i.e., at
≥1000 ppm fomesafen; however, PCO was measured at concentrations as
low as 100 ppm in males, while cell proliferation occurred
dose-dependently only at 3000 ppm after 56 days of treatment.  Because
no tumors were measured in AP rats at any dose, concordance between key
precursors and tumors is lacking.  The lack of dose-response concordance
among the precursor events and the lack of a tumor response in rats
strengthen the argument that the proposed mode of action is specific to
CD-1 mice only.

Several of the key precursor events in the proposed mode of action for
CD-1 mice were also observed in other species treated with fomesafen at
oral concentrations similar to those given to AP rats and Charles River
CD-1 mice.  After 28 days of exposure of hamsters to fomesafen,
increased adjusted liver weights, liver hypertrophy, peroxisome
proliferation, and peroxisomal enzyme activity (PCO and catalase) were
observed in Syrian hamsters (Hart et al. 1985; Appendix 1).  All effects
occurred at the highest concentration tested (3000 ppm) and were
generally not dose-dependent, however.  Increased adjusted liver weights
and peroxisomal number were observed in Beagle dogs after 26 weeks of
oral administration to 25 mg/kg fomesafen (Kalinowski et al. 1981;
Appendix 2).  Last, in addition to hepatic inflammation, increased
peroxisomal density was the only observed effect after 2 weeks of
exposure of marmosets to 50 mg/kg fomesafen (Henderson and Jackson 1982;
Appendix 3).  A dose-response concordance analysis for each of these
studies cannot be performed, however, because although hepatic tumor
responses were not observed in either study, both studies were not of
sufficient duration for a potential tumor response to be measured.

Strength, Consistency, and Specificity of Association of Tumor Response
with Key

Events

 agonists is limited.  As Table 1 indicates, there are three key
events that are causal in the peroxisome proliferation mode of action:
1) activation of the PPAR; 2) disruption of cell
proliferation/apoptosis; and 3) clonal expansion of initiated
hepatocytes.  These are the required causal steps in a PPAR-mediated
mode of action (Klaunig et al. 2003; Cattley et al. 1998).  In the
proposed mode of action for fomesafen-induced hepatocarcinogenesis,
increased cell proliferation (as measured by DNA synthesis) and clonal
expansion (as evidenced by tumor formation) were observed in short-term
and long-term studies, respectively.  This fact adds to the strength of
the proposed mode of action.

PPAR activation was not directly demonstrated in any study.  This
only slightly weakens the proposed mode of action, however.  Activation
of the PPAR is usually demonstrated through the use of an in vitro
reporter assay (Klaunig et al. 2003).  Although an in vitro reporter
assay was not performed to demonstrate either a change in the
3-dimensional binding of PPAR after fomesafen treatment or binding of
PPAR to a peroxisome proliferator response element (PPRE), one in
vitro assay (Roberts 2001) measured the effects of fomesafen on DNA
synthesis, apoptosis, and palmitoyl CoA oxidase (PCO) activity in both
PPAR wild-type (WT) and PPARnull transgenic mouse hepatocytes.
DNA synthesis and PCO were increased and apoptosis was decreased,
relative to untreated cells, in WT hepatocytes at increasing in vitro
concentrations of fomesafen; however, there was no expected change in
any of these endpoints in hepatocytes extracted from PPAR-null mice.
This result supports the conclusion that PPAR activation was
responsible for the observed increase in DNA synthesis and PCO and
decrease in apoptosis after exposure of mouse hepatocytes to fomesafen.
Although these experiments were not carried out in vivo, they do
strengthen the proposed mode of action, because they implicate PPAR
in fomesafen-induced effects in hepatocytes.

Stop/recovery studies generally add strength to a proposed mode of
action.  The fomesafen database contains only one such study.  Sotheran
et al. (1980) performed a short-term study, in which rats were
administered 0 or 1000 ppm fomesafen for 4 weeks.  A significant
increase (P<0.01) in peroxisome proliferation and adjusted liver weights
was observed in males at 1000 ppm (Appendix 4) after 28 days.  Three
weeks following the stoppage of treatment in males administered 1000 ppm
fomesafen, the number of peroxisomes returned to control levels, while a
time-dependent decrease in adjusted liver weights occurred after 3 and 6
weeks of recovery.  Although these results do not strengthen the
relationship between peroxisome proliferation and tumor formation, they
do strengthen the association between exposure to fomesafen, peroxisome
proliferation, and adjusted (relative) liver weights, which are three
key events in the proposed mode of action.

	Consistency of the association of the key events and tumor formation
refers to the repeatability of the key events in different studies,
especially if the studies have different experimental designs
(Sonich-Mullin et al. 2001).  Several of the key precursor events
observed following oral administration of fomesafen to mice were also
observed in rats, hamsters, dogs, and marmosets, although at different
concentrations and after varying lengths of exposure.  In the 2-year
bioassay in rats (Milburn et al. 1984), increased adjusted liver weights
and peroxisome proliferation were observed, however without any tumor
response.  In a short-term feeding study in rats (Moffat 1998),
increased relative liver weights, liver hypertrophy, and increased cell
proliferation and PCO activity were observed after 56 days of treatment
with fomesafen.  After 28 days of exposure of hamsters to fomesafen,
increased adjusted liver weights, peroxisome proliferation (volume
density), and peroxisomal enzyme activity (PCO and catalase) were
observed in Syrian hamsters (Hart et al. 1985).  PCO activity was
similar to controls, however, after 10 or 14 days of exposure of guinea
pigs to 50 mg/kg fomesafen (Elcombe 1988).  Increased adjusted liver
weights and peroxisomal number were also observed in Beagle dogs
following 26 weeks of oral administration of fomesafen (Kalinowski et
al. 1981), while increased peroxisomal density was observed after 2
weeks of exposure of marmosets to fomesafen (Henderson and Jackson
1982).  Observations of many of the key events was, therefore,
relatively consistent across a number of studies in several species.

Specificity of the association of the key events and tumor formation in
a proposed mode of action refers to the fact that the proposed mode of
action is specific to a given site of action (e.g., organ), cell type,
species, or strain of animal, as long as evidence of other modes of
action is lacking.  In the case of fomesafen, the proposed mode of
action is specific to mice only.  Although some or several key events
were observed in studies performed in other species, especially AP rats,
liver tumors developed in Charles River CD-1 mice only.  The lack of
tumor formation in the rat is not surprising, given the fact that
peroxisome proliferation was not increased more than 5-fold after 1 year
of treatment with fomesafen.

 agonists.  Specifically, there was no evidence of pancreatic acinar
cell tumors (PACTs) in any of the treated groups in either sex, and the
incidence of interstitial cell tumors in the testes after 2 years was
1/127 (1%), 0/63 (0%), 2/64 (3%), 0/64 (0%), and 0/64 (0%) at 0, 1, 10,
100, and 1000 ppm fomesafen (Colley et al. 1983).  The lack of
dose-response supports the conclusion that the development of
interstitial cell tumors was not substance-related.  Therefore, because
fomesafen caused the development of hepatic tumors only and none of the
other members of the so-called “tumor triad” (PACTs and Leydig cell
tumors) was observed in CD-1 mice, the specificity of the proposed mode
of action is strongly supported.

Biological Plausibility and Coherence

	The mode of action proposed in this paper is consistent with what is
currently known about the process of liver carcinogenesis.  Table 2
demonstrates that although there are many steps in the general mode of
action of a peroxisome proliferator, knowledge of the causal steps
leading to liver tumors is limited.  There are only three causal steps
in the process as it is currently understood, and two of these steps
(disruption of cell proliferation/apoptosis and clonal expansion of
initiated cells) are common to many, if not all, other carcinogens.
Activation of PPAR has both a causal and highly specific association
to hepatocarcinogenesis mediated by peroxisome proliferators (Table 2).

The two other key events that are highly specific to the proposed mode
of action are the expression of peroxisomal genes and peroxisome
proliferation.  Each of these events is correlated with and not
causative of liver cancer; therefore, liver tumor formation may be
occurring independently of peroxisome proliferation (Bosgra et al.
2005).  However, based on the currently accepted understanding of
hepatocarcinogenesis mediated by peroxisome proliferators, the proposed
mode of action for fomesafen is biologically plausible in animals.  For
example, two of the three currently recognized causal key events,
disruption of cell proliferation and clonal expansion, have been
demonstrated in at least one animal species.  In addition, two of the
three highly specific key events, namely, peroxisomal gene expression
and peroxisome proliferation, have been observed in multiple species of
animals.  Table 7, which is an adaptation of Table 2 (Klaunig et al.
2003), summarizes the key events and animal evidence for
fomesafen-induced hepatocarcinogenesis in mice.

Table 7. Key events associated with the hepatocarcinogenesis of
fomesafen.

Key event	Evidence in animals

	Activation of PPAR	No data (indirect in vitro evidence in
PPAR-null mouse hepatocytes; Roberts 2001)

Expression of peroxisomal genes	Dose-dependent increase in palmitoyl CoA
oxidase activity in mice, rats, and hamsters (Moffat 1998; Hart et al.
1985)

Increase in hepatic catalase activity in hamsters (Hart et al. 1985)

PPAR-mediated expression of cell cycle, growth, and apoptosis	No in
vivo data

Non-peroxisome lipid gene expression	Decreased serum triglycerides and
cholesterol in mice and rats (Milburn et al. 1984; Moffat 1998)

Peroxisome proliferation	Dose-related increase in peroxisomal volume
density or number in mice, rats, hamsters, dogs, and marmosets (Moffat
1998; Hart et al. 1985; Kalinowski et al. 1981; Henderson and Jackson
1982)

Disruption of cell proliferation/apoptosis	Dose-dependent increase in
cell proliferation in mice and rats

Dose-related increase in relative liver weights2 in mice, rats, and
hamsters (Colley et al. 1983; Milburn et al. 1984; Moffat 1998; Hart et
al. 1985)

No data for decreased apoptosis

Inhibition of gap junction intercellular communication (GJIC)	No data

Hepatocyte oxidative stress	No data

Kupffer cell-mediated events	No data

Selective clonal expansion	Hepatocellular tumors in mice only (Colley et
al. 1983)

1Required causal or associative key events with high specificity and a
large weight of evidence in bold.  A lack of data for any of the other
key events does not weaken the proposed mode of action (see p. 6).

2It should be noted that in the absence of evidence of hyperplasia,
increased relative liver weight is indicative of liver hypertrophy
only.

Fomesafen is a member of the diphenyl ether herbicide family, which
includes other compounds such as lactofen, acifluor(f)en, and
oxyfluor(f)en.  Both lactofen and acifluorfen have been shown to induce
liver tumors in rodents whereas oxyfluorfen is only marginally active.
Representative structures are shown here:

The mode of action of rodent hepatocarcinogenesis for both lactofen and
acifluorfen (HED MTARC; TXR #s 0051907 and 0052006, respectively) has
been studied and shown to involve peroxisome proliferation.  Lactofen
can be readily hydrolyzed by esterases to yield acifluorfen as its
primary metabolite.  Metabolism studies by Syngenta Crop Protection,
Inc. (Peffer 2004) showed that up to 10% of fomesafen may be hydrolyzed
to yield a carboxylic acid metabolite as the most significant
metabolite.  Thus, fomesafen, acifluorfen, and lactofen may actually
have common carboxylic acid metabolite(s).  Structure-activity
relationships studies have shown that one of the major structural
requirements of most peroxisome proliferators is the presence of an
acidic functional group (e.g., carboxylic) either in the parent compound
or a metabolite (Woo and Lai 2003).  It is interesting to note that,
despite structural similarity, oxyfluorfen, which cannot be metabolized
to a carboxylic acid metabolite, is only marginally active as a
hepatocarcinogen.  Overall, these findings strengthen the biological
plausibility of the proposed mode of action for fomesafen-induced liver
tumor formation in animals.

Note:  Both Lactofen and Acifluorfen were classified as “  SEQ CHAPTER
\h \r 1 likely to be carcinogenic to humans at high enough doses to
cause these biochemical and histopathological effects in livers of
rodents but unlikely to be carcinogenic at doses below those causing
these changes.”  (Aciflurofen, like Fomesafen had only liver tumors in
mice, but Lactofen had liver tumors in mice and  rats.)    Peroxisome
Proliferation was supported as the mode of action and CARC recommended
using the MOE approach for estimating human cancer risk for both
chemicals.  Since that time, PPAR( agonist-mediated hepatocarcinogenesis
in rodents and relevance to human health risk assessment have been more
definitively presented in publications, such as Klaunig et al (2003) and
the FIFRA SAP Memo (March 5, 2004).

	Coherence refers to the overall consistency of the database relative
to the proposed mode of action.  Fomesafen has been shown to have
hypolipidemic effects in short-term studies in mice and rats (Moffat
1998) and in the 2-year bioassay in rats (Milburn et al. 1984).
Specifically, decreases in plasma triglycerides, and to a lesser extent
cholesterol, were observed in each of these studies.  Decreases were
usually at the highest dose tested and often dose-dependent.  Decrements
in serum triglyceride and cholesterol concentrations are consistent with
the proposed mode of action, because peroxisome proliferators also
induce hypolipidemic effects in animals through non-peroxisome lipid
gene expression (Klaunig et al. 2003).  The induction of this non-cancer
endpoint is, therefore, consistent with the proposed mode of action and
thereby strengthens the coherence of the database.  Because fomesafen is
neither a reproductive nor a developmental toxicant, the proposed mode
of action for hepatocarcinogenesis is not relevant for these non-cancer
endpoints.

 in hepatocytes is proposed as the mode of action for liver tumors.
This internal consistency in the database (coherence) strengthens the
proposed mode of action.

Other Modes of Action

Based on the database for fomesafen, two additional modes of action have
been hypothesized.  These are mutagenicity and cytotoxicity (including
hepatic porphyria) followed by regenerative proliferation.  The weight
of the evidence supports the conclusion that fomesafen is not mutagenic.
 In addition, there are no consistent data to support a
cytotoxic/regenerative mode of action.

Mutagenicity

Eleven mutagenicity studies were submitted for review.  Nine studies
[Salmonella typhimuruim reverse mutation test (CTL/P/596, MRID 103018),
gene mutation in L5178Y mouse lymphoma cells (CTL/P/2058, MRID
40910804), in vitro mammalian cytogenetics chromosomal aberration assays
in human lymphocytes (MRID 44569805, 44569806), dominant lethal study in
CD-1 mice (CTL/P/609, 103019), in vivo bone marrow cytogenetic assays in
rats (CTL/P/623/ MRID 00164907, CTL/P/823/ MRID 00135628), in vivo
micronucleus assay in mice (CTL/P/2156, MRID 40910805)] and mammalian
cell transformation in baby hamster kidney fibroblasts (CTL/P/596, MRID
103018) were acceptable and used to evaluate the mutagenic potential of
fomesafen.

ed at concentrations ≤100 ug/mL.  In the second study (MRID 44569805),
fomesafen was tested up to a cytotoxic concentration of 1000 ug/mL.  A
significant increase in chromosome breaks was observed in one donor at
1000 ug/mL; however, this was most likely secondary to cytotoxicity as
demonstrated by the decrease in the mitotic index (MI, 57%).  Fomesafen
was not clastogenic at concentrations ≤500 ug/mL (-S9) and 250 ug/mL
(+S9).  In the dominant lethal assay, there was no effect on fertility,
and dominant lethality was not demonstrated.  In this study, adequate
toxicity was demonstrated by the death of three animals during the first
3 days of treatment and a fourth by week 4.  Neither clastogenicity nor
aneugenicity was observed in male and female C57B1/6JfCD-1/Alpk mice
after a single dose of fomesafen at 279 and 344 mg/kg (80% of LD50) for
males and females, respectively, in the mouse micronucleus assay
(CTL/P/2156, MRID 40910805).  In addition, fomesafen did not cause cell
transformation in baby hamster kidney fibroblasts at 240 ug/mL (LD50).
No genetic toxicology studies using fomesafen were found in the open
literature.

Cytotoxicity/regenerative hyperplasia

            Hepatic porphyria (porphyrin accumulation) was hypothesized
to occur after in vivo exposure of mice to fomesafen, due to its mode of
action in plants.  Diphenyl ether herbicides are known to induce the
accumulation of a toxic intermediate, protoporphyrin IX, in plant cells
through the inhibition of protoporphyrinogen oxidase.  Cell membrane
peroxidation is believed to ensue, ultimately leading to cell lysis
(Matringe et al. 1989).

Mammalian cells also contain protoporphyrinogen oxidase, which catalyzes
the second-to-last-step of heme biosynthesis.  In order to test the
hypothesis that fomesafen inhibits protoporphyrinogen oxidase, thus
leading to an accumulation of porphyrin in the liver (a marker for
cytotoxicity), Moffat and Townsley (2004) treated male and female CD-1
mice with 0, 1, 10, 100, or 1000 ppm fomesafen to CD-1 mice for 7 or 28
days.  A dose-dependent increase in relative liver weights was observed
in both sexes at both time periods, with significant increases (P<0.05)
at 100 and 1000 ppm fomesafen, while liver hypertrophy was observed only
at 100 and 1000 ppm.  Although hepatic porphyrin accumulation was not
observed by light microscopy, dose-dependent and significant increases
(P<0.05) in hepatic porphyrin content were measured biochemically at 100
and 1000 ppm.

The increase in hepatic porphyrin content at 100 and 1000 ppm after 7 or
28 days is consistent with hepatoxicity observed in mice in both long-
and short-term studies.  Table 8a summarizes the concentrations of
alanine aminotransferase (ALT) and alkaline phosphatase (ALP) in the
blood after one year of exposure to fomesafen.  Both ALT and ALP, which
are hepatic enzymes and indicators of hepatocellular injury when
elevated, were significantly increased (P<0.05) in a dose-dependent
manner after one year of exposure to 100 or 1000 ppm fomesafen.  At 1000
ppm fomesafen, ALT and ALP levels were at least 3-fold higher than
concurrent control levels.  Because these are the same concentrations at
which significant increases in mean absolute liver weight and
hepatomegaly occurred, the clinical chemistry results argue that
hepatocellular toxicity is also taking place after administration of
fomesafen.  Similar clinical chemistry effects were observed in rats.
While hyperplasia was not observed in any study, increased DNA synthesis
was measured in both short-term assays.  In addition, a dose-dependent
increase in single cell necrosis and brown macrophages and Kupffer cells
was observed in male mice at 79 weeks of treatment (termination) with
fomesafen (Table 8b).

Table 8a. ALT and ALP measurements taken in the 2-year bioassay in CD-1
mice (Colley et al. 1983)

Clinical Chemistry (mU/ml)

Concentration (ppm)	Week 51	Week 52

	ALT	ALP	ALT	ALP

	Combined sexes	Combined sexes	Combined sexes	Combined sexes

0	41±23	9±3	36±18	13±5

1	70±30	9±2	39±21	14±5

10	51±38	11±6	43±17	12±4

100	100±110**	22±6**	56±20*	23±12

1000	266±193**	891±1154**	118±85**	186±219**

* p<0.05

** p<0.01

Table 8b. Liver histology in the 2-year bioassay in CD-1 mice (Colley et
al. 1983)

Liver Histology

Concentration (ppm)	52-wk. interim sacrifice	Intercurrent Deaths
Termination [Week 79 (M) or 89 (F)]

	Brown pigmented macrophages1	Single cell necrosis	Brown pigmented
macrophages	Single cell necrosis	Brown macrophages & Kuppfer cells

	M	F	M

	F	M	F	M

	F	M

	F

0	0/23 	5/24 (21)	0/58 (0)

	0/54	1/58 (2)	1/54 (19)	0/46

	0/50	0/46	0/50

1	0/12	1/10 (10)	1/24 (4)	0/35	1/24 (4)	4/35 (11)	0/27	0/19	0/27	0/19

10	2/11 (18)	1/12

(8)	0/34 (0)

	0/34	8/34 (24)	7/34 (21)	0/19	0/18	3/19 (16)	0/18

100	5/9 (56)	7/10 (70)	3/40 (8)

	0/28	5/40 (13)	3/28 (11)	1/15 (7)

	0/24	9/15 (60)	4/24 (17)

1000	5/9 (56)	10/11 (91)	7/44 (16)	3/38 (8)	23/44 (52)	19/38 (50)	5/11
(45)	0/15	9/11 (82)	7/15 (47)

1 No increase in single cell necrosis at any concentration

2 incidences in parentheses

While the histology and clinical chemistry results indicate that
hepatocellular toxicity occurred in both long and short-term studies
(Tables 9a and 9b), several observations should be noted that challenge
a cytotoxic mode of action.  For example, the increases in ALT, AST, and
ALP were highly variable and suggest that very high doses of fomesafen
are needed to induce cytotoxicity at early time periods.  More
importantly, the cell proliferation data from the 28- and 56-day studies
are not consistent with a cytotoxic mode of action, because cell
proliferation decreased after 56 days.  A sustained cell proliferative
response would be expected in a cytotoxic mode of action.  Third, -GT
and bilirubin levels did not differ from controls in any short-term
assay, thereby suggesting that hepatobiliary obstruction did not occur.
In addition, single cell necrosis was not observed either at 52 weeks in
either sex or at 89 weeks (termination) in females in the 2-year
bioassay in the mouse.  Fourth, the incidence of single cell necrosis in
intercurrent deaths in males and of brown pigmented macrophages at 52
weeks (females) and for intercurrent deaths (both sexes) was not
dose-dependent.  Fifth, hepatic vacuolation and atrophy (indicators of
hepatotoxicity) were not observed in mice at 100 ppm fomesafen after 56
days of exposure.  This observation contrasts with the finding that all
of the key events in the proposed mode of action in mice were observed
at ≥100 ppm, while significant increases in PCO were observed in both
sexes (P<0.05) at concentrations as low as 10 ppm.

Table 9a. ALT, AST, and ALP measurements taken in the 28-day study in
CD-1 mice (Moffat 1998)

Clinical Chemistry (Experiment 1)

Concentration (ppm)	Day 7	Day 28

	ALT	AST	ALP	ALT	AST	ALP

	M	F	M	F	M	F	M	F	M	F	M	F

0	20.0±5.1	25.4±17.3	35.0±5.7	47.0±24.5	224.5±57.9	197.0±40.0
24.8±9.1	23.4±5.1	36.0±9.3	58.0±5.9	158.4±36.3	188.4±46.6

1	23.4±4.4	18.4±3.2	33.6±5.6	39.2±5.8	255.6±48.8	240.6±62.0
21.2±3.9	21.2±3.4	34.2±3.6	55.6±12.1	150.2±51.9	211.0±74.8

10	22.0±1.6	20.2±6.5	30.5±3.1	40.8±5.9	225.5±19.4	247.0±64.1
20.0±4.5	24.2±3.0	38.6±7.5	65.4±22.2	210.0±62.5	175.8±33.3

100	23.4±7.1	24.6±11.9	35.4±4.2	49.4±5.9	394.2±121.2*	257.0±70.3
34.4±18.1	26.0±5.2	44.4±9.5	60.8±8.2	184.2±20.9	190.6±43.9

1000	149.4±68.3***	24.2±6.6	101.8±42.1**	41.0±4.4	521.0±119.1***
686.6±546.0	67.6±28.7***	55.2±29.4**	63.8±11.5***	78.6±11.1*
364.0±132.4***	385.8±228.6*

3000	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND

ND – no data

*p<0.05

** p<0.01

*** p<0.001

Table 9b. ALT, AST, and ALP measurements taken in the 56-day study in
CD-1 mice (Moffat 1998)

Clinical Chemistry (Experiment 2)

Concentration (ppm)	Day 7	Day 56

	ALT	AST	ALP	ALT	AST	ALP

	M	F	M	F	M	F	M	F	M	F	M	F

0	21±5	23±7	31±2	48±12	176±47	199±41	26±5	21±4	37±4	46±6
107±17	135±48

1	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND

10	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND	ND

100	27±9	21±4	40±8	37±6	168±105	240±67	28±7	39±26	41±4	58±21
151±32	207±50

1000	32±4	28±14	37±2	51±20	316±147	192±99	90±42	34±14	67±36
55±14	961±660	257±180

3000	105±56***	65±53*	82±30***	66±21	466±165**	951±617**
191±125**	162±110**	160±106**	137±63**	3654±2075***	3062±1758**

ND – no data

*p<0.05

** p<0.01

*** p<0.001

Uncertainties, Inconsistencies, and Data gaps

Several inconsistencies were observed in the various datasets presented
here.  The first is the increased incidence of adenomas in male mice at
1 ppm, but not 10 ppm fomesafen (Table 1).  The result is inexplicable,
given the fact that the incidence is not only significantly increased
above controls (p<0.001), but is also outside the laboratory’s
historical control range.  In Table 4c, cell proliferation (DNA
synthesis) at 28 days in female mice decreased to baseline, rather than
increasing, at 1000 ppm as expected.  The significant increase at 100
ppm (p<0.05) is questionable, however, given the large variability
around the mean.  Similarly, cell proliferation in male rats on day 7
decreased to baseline at 3000 ppm fomesafen (Table 6c).  This decrease
is inconsistent with a dose-dependent increase in DNA synthesis in males
on day 7.  Fourth, the significantly increased incidence (p<0.05) of
peroxisome proliferation in male rats at termination (Table 5) was not
dose-dependent.  However, this discrepancy is likely due to the
differential number of rat livers examined (3, 1, and 6 at 5, 100, and
1000 ppm, respectively).

Conclusions

Based on the data presented here and the current state of the science
with regard to peroxisome proliferators, it is reasonable to conclude
with a high level of confidence that fomesafen-induced
hepatocarcinogenesis in CD-1 mice is mediated through activation of
PPAR.  Two of the three known causal events in the proposed mode of
action, i.e., increased cell proliferation and tumor formation, have
been shown to occur in short- and long-term bioassays, respectively.
The third known causal event, activation of PPAR, has also been
indirectly demonstrated in one in vitro assay.  In addition, two key
associative events, namely increased expression of peroxisomal genes and
proliferation of peroxisomes, have been demonstrated in several
short-term studies of fomesafen across species.

The available data do not support the conclusion that fomesafen is
mutagenic.  While there is evidence of hepatotoxicity and hepatic
porphyria following administration of fomesafen, the data suggest that
high concentrations of fomesafen (>1000 ppm) are needed for
hepatotoxicity to take place.  In addition, although sometimes difficult
to detect, compensatory hyperplasia was not observed following oral
exposure to fomesafen.  While increased DNA synthesis was observed in
short-term studies in the mouse and rat, the results after 56 days of
treatment indicate that this effect was not sustained.  Therefore, the
data do not support cytotoxicity followed by regenerative proliferation
as an alternative mode of action.

Human Relevance (including lifestages)

as the mode

 and human hypolipidemic drugs work through this receptor, the mode
of action of fomesafen-induced hepatocarcinogenesis is qualitatively
plausible in humans.

The proposed mode of action in adults, however, is not quantitatively
plausible,

 activation by fomesafen has not been tested in an in vitro gene
reporter assay, in vitro reporter assay data are lacking for mouse
PPAR activation following administration of fomesafen as well.

M fomesafen for 72 hrs. (Brady 1998).  PCO was increased 2.4-fold at
1,000 M.  However, there was no statistically significant increase in
hepatocyte PCO across human donors, including a 9- and 10-year-old male
and female, respectively, after treatment with fomesafen.  The
non-significant change in PCO was confirmed in a follow-up study in
which hepatocytes of a 4-year-old male were exposed to 0-500 M
fomesafen for 72 hrs. (Roberts 2001).  Despite deficiencies in each
study, no population or lifestage that is particularly susceptible to
the proposed mode of action has been identified in the existing database
for fomesafen.

Table 10. Key events and plausibility in humans (Peffer 2004)

Overall MOA Summary

 activation and toxicokinetic and toxicodynamic factors are taken
into account (Klaunig et al. 2003).

IV.	WEIGHT-OF-THE-EVIDENCE CONSIDERATIONS

Carcinogenicity

Mouse

Dietary administration of fomesafen was associated with increased
incidences of liver tumors in male and female mice.

In male mice, the incidence of liver adenomas, carcinomas, and adenomas
and/or carcinomas combined for the control , 1, 5, 100, and 1000 ppm
dose groups, respectively, were as follows:

Adenomas: 13/110 (12%), 19/56 (34%), 5/52 (10%), 17/54 (31%), 14/36(39%)

Carcinomas: 17/127 (13%), 7/63 (11%), 12/64 (19%), 10/64 (16%), 28/64
(44%)

Combined: 30/127 (24%), 26/63 (41%), 17/64 (26%), 27/64 (42%), 42/44
(66%)

In female mice, the incidence of liver adenomas, carcinomas, and
adenomas and/or carcinomas combined for the control , 1, 5, 100, and
1000 ppm dose groups, respectively, were as follows:

Adenomas: 3/128 (2%), 1/63 (1%), 1/62 (1%), 8/62 (13%), 12/48 (25%)

Carcinomas: 0/128 (0%), 1/64 (1%), 2/64 (3%), 2/64 (3%), 16/64 (25%)

Combined: 3/128 (2%), 2/64 (3%), 3/64 (5%), 10/64 (16%), 28/64 (44%)

Fomesafen was associated with significantly elevated incidences of liver
tumors (adenomas, carcinomas, and adenomas/carcinomas combined) in male
and female CD-1 mice.  In males, adenomas were elevated at 1, 100, and
1000 ppm and carcinomas were elevated at 1000 ppm. In addition, the
combined adenomas and/or carcinomas were increased at 1000 ppm.  In
females, adenomas were elevated at 100 and 1000 ppm and carcinomas were
elevated at 1000 ppm. In addition, adenomas and/or carcinomas combined
were increased at 100 and 1000 ppm.  There was evidence for a
progression of benign tumors to malignancy, and for a reduction in the
latency period for the time-to-tumor appearance.  There was no evidence
for the occurrence of hyperplastic changes in the livers of treated
mice. These tumors were considered to be treatment-related.

Liver tumors were seen in male mice at dose levels (i.e., 100 and 1000
ppm) that seemed to exceed a MTD level, and also at a dose level (i.e.,
1 ppm) that was below a MTD level.  In addition, liver tumors were seen
in female mice at a dose level (i.e., 1000 ppm) that seemed to exceed a
MTD level, and also at a dose level (i.e., 100 ppm) that approximated a
MTD level.

Rat

Fomesafen was not carcinogenic when administered in the diet to Wistar
albino rats at doses ranging from 1 to 1000 ppm.  The highest dose
tested in males (1000 ppm) in the chronic rat bioassay exceeded a MTD
level, whereas this dose in females approximated the MTD.

Mutagenicity

Fomesafen is not mutagenic or clastogenic in in vitro and in vivo
studies. The data do not support a mutagenic mode of action.

MOA

 activation and toxicokinetic and toxicodynamic factors are taken
into account (Klaunig et al. 2003). The weight of the evidence supports
the conclusion that fomesafen is not mutagenic and there are no
consistent data to support a cytotoxic/regenerative mode of action.

SAR

Fomesafen is a member of the diphenyl ether herbicide family and is
structurally closely related to lactofen, acifluor(f)en, and
oxyfluor(f)en.  Both lactofen and acifluorfen have been shown to induce
liver tumors in rodents whereas oxyfluorfen is only marginally active.

V.	CLASSIFICATION OF CARCINOGENIC POTENTIAL

 activation and toxicokinetics.

QUANTIFICATION OF CARCINOGENIC POTENTIAL

The quantification of risk is not required.

VII.	REFERENCES  tc "V. 	SUMMARY"

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Appendix 1

Note: Increased centrilobular hypertrophy in males (9/10) and females
(9/10), relative to controls (0/10), was also observed at 3000 ppm (Hart
et al. 1985).

Appendix 2

Appendix 3

Appendix 4

Note: Ref 1.  ICI Americas Inc. Registration Application for "Flex" 2 LC
Herbicide in Soybeans. Section C. Mammalian Toxicology (April 1984).
According to Sotheran et al. (1980), peroxisome quantitation took place
after 3 weeks of recovery, not 14 days.

NOTE

DATE: 	November 3, 2005

SUBJECT: 	Fomesafen: Second Report of the Cancer Assessment Review
Committee

FROM: 	Esther Rinde, Ph.D.

Health Effects Division (7509C)

Although I have signed the Fomesafen Document, I would like to add the
following comment:

I think that the Descriptor for Fomesafen which is (Not Likely( should
have read instead :

(Not Likely to be Carcinogenic to Humans based on Quantitative Species
Differences in PPARα Activation and Toxicokinetics.(

This would clearly distinguish it from chemicals with no tumor response
at all, especially when the classification is quoted or placed on a
list, without benefit of the accompanying paragraph in the document.

 agonist-induced rodent tumors: modes of action and human relevance.
Crit Rev Toxicol 33(6):655-780.

 Increased ALT, AST, and/or ALP were also observed in a dose-dependent
manner at weeks 1, 4, 8, and 52

 Evidence for fomesafen-induced peroxisome proliferation and
perturbations in cell proliferation/apoptosis in human hepatocytes is
based on in vitro data only (Brady 1998; Roberts 2001).

FOMESAFEN	CANCER ASSESSMENT DOCUMENT             	                FINAL

 PAGE

 PAGE   3

FOMESAFEN                       CANCER ASSESSMENT DOCUMENT
                                  FINAL

 PAGE

 PAGE   49


[Federal Register Volume 79, Number 141 (Wednesday, July 23, 2014)]
[Proposed Rules]
[Page 42745]
[FR Doc No: 2014-17322]

=======================================================================
-----------------------------------------------------------------------

ENVIRONMENTAL PROTECTION AGENCY

40 CFR Part 13

[EPA-HQ-OA-2014-0012; FRL-9914-28-OCFO]

Administrative Wage Garnishment

AGENCY: Environmental Protection Agency (EPA).

ACTION: Proposed rule, extension of public comment period.

-----------------------------------------------------------------------

SUMMARY: The EPA is extending the period for providing comments on the
proposed rule entitled, Administrative Wage Garnishment published in
the Federal Register on July 2, 2014 to September 2, 2014.

DATES: Comments. The public comment period for the proposed rule
published July 2, 2014, (79 FR 37704) is being extended to September 2,
2014 in order to provide the public additional time to submit comments
and supporting information.

ADDRESSES: Comments. Written comments on the proposed rule may be
submitted to the EPA electronically, by mail, by facsimile or through
hand delivery/courier. Please refer to the proposal (79 FR 37704) for
the addresses and detailed instructions.
    Docket: Publically available documents relevant to this action are
available for public inspection either electronically at http://www.regulations.gov or in hard copy at the Administrative Wage
Garnishment Docket in the EPA Docket Center, EPA/DC, EPA West, Room
3334, 1301 Constitution Ave. NW., Washington, DC. The Public Reading
Room is open from 8:30 a.m. to 4:30 p.m., Monday through Friday,
excluding legal holidays. The telephone number for the Public Reading
Room is<PHONE_NUMBER>. The EPA has established the official public
docket  EPA-HQ-OA-2014-0012.

FOR FURTHER INFORMATION CONTACT: FPPS c/o Anita Jones, OCFO/OFM/FPPS,
Mailcode 2733R, Environmental Protection Agency, 1200 Pennsylvania Ave.
NW., Washington, DC 20460; telephone number:<PHONE_NUMBER>; fax
number:<PHONE_NUMBER>; email address<EMAIL_ADDRESS>
SUPPLEMENTARY INFORMATION: In response to requests from the public, the
EPA is extending the previously announced public-comment period. The
public-comment period will end September 2, 2014, rather than August 1,
2014. The direct final rule published at 79 FR 37644 on July 2, 2014
was withdrawn. The withdrawal notice of the direct final rule was
published in the Federal Register on Thursday, July 17, 2014 at 79 FR
41646.

List of Subjects in 40 CFR Part 13

    Environmental protection, Administrative practice and procedure,
Claims, Debt Collection, Government employees, Garnishment of wages,
Hearing and appeal procedures, Salaries, Wages.

    Authority: 5 U.S.C. 552a, 5512, and 5514; 31 U.S.C. 3711 et
seq.; 3720A; and 3720D.

    Dated: July 17, 2014.
Jeanne Conklin,
Acting Director Office of Financial Management.
[FR Doc. 2014-17322 Filed 7-22-14; 8:45 am]
BILLING CODE 6560-50-P