PCCode: 138831 FINAL

UNITED STATES ENVIRONMENTAL PROTECTION AGENCY
WASHINGTON, D.C. 20460

MEMORANDUM

Date: December 30, 2013 OFFICE OF CHEMICAL SAFETY

AND POLLUTION PREVENTION

SUBJECT: CYFLUMETOFEN: Report ofthe Cancer Assessment Review Committee

PC Code: 138831

Decision No.: N/A

Petition No.: N/A

Risk Assessment Type: Cancer Assessment

TXR No.: 0056862

MRID No.: N/A DP Barcode: N/ A Registration No.: N/A Regulatory Action: N / A

Case No.: N/A

CAS No.:

40 CFR: N/A

FROM: Kristin Rury, Executive Secretary 

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Cancer Assessment Review Committee

Health Effects Division (7509P)

THROUGH: Jess Rowland, Chair**~~** :**~** I

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### n--~**\2-l;lo ·**  Karlyn Middleton, Co-Chair**~** 

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Cancer Assessment Review Committee

Health Effects Division (7509P)

TO: Yung Yang, Toxicologist

RAB VI

, Health Effects Division (7509P)

Richard Gebken

Product Manager, Registration Division

The Cancer Assessment Review Committee met on December 17, 2013 to evaluate the carcinogenic potential of cyflumetofen in accordance with the EPA ' s Final Guidelines for Carcinogen Risk Assessment (March

, 2005)

. Attached please find the final Cancer Assessment

Document.

1PC Code: 138831 FINAL

2PC Code: 138831 FINAL

3 CANCER ASSESSMENT DOCUMENT

EVALUATION OF THE CARCINOGENIC POTENTIAL OF Cyflumetofen

PC 138831

December 30, 2013



### C ANCER 

### A SSESSMENT 

### R EVIEW 

### C OMMITTEE

HEALTH EFFECTS DIVISION

OFFICE OF PESTICIDE PROGRAMSPC Code: 138831 FINAL

4 CONTENTS

Executive Summary ............................................................................................................. ............4

I. Introduction .............................................................................................................. ..................6

II. Background Information ................................................................................................... ........6

III. Evaluation of Carcinogenicity ........................................................................................... ......6

1. Combined Chronic Toxicity & Carci nogenicity Study in Fischer 344 Rats ...................6

2. Carcinogenicity Study in CD-1 Mice ............................................................................12

IV. Toxicology ............................................................................................................... ..............14

1. Metabolism ................................................................................................................. ...14

2. Mutagenicity ............................................................................................................... ...15

3. Structure Activit y Relationship ......................................................................................19

4. Subchronic and Chronic Toxicity ..................................................................................19

V. Committee’s Assessment of the Weight-of-the Evidence ......................................................23

1. Carcinogenicity ............................................................................................................ ..23

2. Mutagenicity ............................................................................................................... ...24

3. Structure Activit y Relationship ......................................................................................24

VI. Classification of Ca rcinogenic Potential ...............................................................................24

VII. Quantification of Carcinogenicity ........................................................................................ ..24

VIII. Bibliography ............................................................................................................ ..............25PC Code: 138831 FINAL

5 EXECUTIVE SUMMARY

The carcinogenic potential of cyflumetofen wa s evaluated following dietary administration to

male and female Fischer 344 rats and male and female CD-1 mice. Two separate studies were conducted in both species. In 2004, the Registrant conducted a carcinogenicity study in Fischer 344 rats (MRID No. 48542697) an d a carcinogenicity study in CD-1 mice (MRID No. 48542698).

Since there was no evidence of carcinogenicity in e ither species, and to a ddress concern regarding the adequacy of the doses tested in the above studies, the Registrant elected to conduct supplemental studies to evaluate higher doses in mice (MRID No 49209101) and rats (MRID No. 49226601).

Cyflumetofen was administered in the diet to gr oups of 50 rats/sex/dose for 104 weeks. One study was conducted at doses of 0, 150, 500 and 1500 ppm (0, 4.9, 16.5, and 49.5 mg/kg/day in males and 6.14, 20.3, and 61.9 mg/kg/day in females) while a second high dose study was conducted at 0 and 6000 ppm (0 and 220 mg/kg/day in males and 0 and 287 mg/kg/day in females). There was limited evidence of carcinogenicity characterized by the presence of thyroid c-cell tumors and

interstitial cell tumors of the testes in male rats only at the highest dose tested (HDT). There was no evidence of carcinogenicity in female rats at an y dose level. The doses tested in the rat study were considered to be adequate to asse ss the carcinogenic potent ial of cyflumetofen.

The thyroid c-cell tumors were determined to be treatment-related based on the following considerations: a statistically significant trend and a pair-wise significance (both at p < 0.05) for thyroid c-cell adenocarci nomas when compared to controls; a statistically significant trend (p < 0.05) and a pair-wise significance (p<0.01) for thyr oid c-cell adenomas and/or adenocarcinomas combined; and the incidences of the adenocar cinomas (31%) and the combined tumors (57%)

exceeded the historical control ra nge from the testing laboratory (c-cell adenocarcinoma, 6-18%, and combined c-cell adenoma/carcinomas, 26-50%).

The testicular interstitial cell tumors, although exacerbated by treatme nt were not considered to be biologically relevant since this tumor (Leydig ce ll, LCT) is extremely common in the F344 rat, with a reported background incidence of 56-100% in 2-year cancer studies. The reasons for the different background incidence rate for LCT am ong species stem from both quantitative and qualitative differences in the Leydig cell respons e to hormonal stimuli. The rat Leydig cell is extremely sensitive to slight changes in circul ating luteinizing hormone (LH) levels. Among rat strains, the F344 rat is particular ly susceptible to this process a nd is not considered a relevant model for studying LCT. By contrast, the inciden ce rate in humans of LCT is very low (i.e., 0.00004%). Similarly, rats are known to have approxima tely 13-times more LH receptors per cell than humans (Hutaniemi, 1983). Because of the diffe rences present between the human and the rat Leydig cells, non-genotoxic compounds that cause LCTs in rats ha ve little relevance to humans. The data with cyflumetofen supports this species and strain-specific effect on the Leydig cells.PC Code: 138831 FINAL

6 The carcinogenic potential of cyflumetofen wa s evaluated following dietary administration to

male and female CD-1 mice. Two studies we re conducted in the mice. Cyflumetofen was administered to groups of 52 mi ce/sex/dose for 78 weeks. One study was conducted at doses of 0, 150, 500, 1500 and 5000 ppm (0, 15.5, 54.3, 156 and 537mg/kg/day in males and 14.3, 48.1, 144 and 483 mg/kg/day in females) while a second high dose study was conducted at 0 and 10,000 ppm (0 and 1143 mg/kg/day in males and 1132 mg/kg/day in females. There was no evidence of carcinogenicity in male or female mice at any dose level. The doses tested in the mouse were considered adequate to assess the car cinogenic potential of cyflumetofen.

The mutagenic potential of cyflumetofen, its metabolites, and an impurity was tested in in vitro

and in vivo studies. There was no concern for mutagenicity for the parent, the metabolites or the impurity. Consequently, mutagenicity can be rule d out as a possible mode of action for tumor

formation.

In accordance with the EPA's Final Guidelines for Carcinogen Risk Assessment (March, 2005), the Cancer Assessment Review Committee (CARC) classified cyflumetofen as “Suggestive Evidence of Carcinogenic Potential”. This classifi cation is based on the presence of a single tumor type (thyroid c-cell) in one sex (male) and one species (rat), and the lack of concern for mutagenicity. When there is suggestive evidence of carcinogenicity, the Agency does not attempt a dose-response assessment as the nature of th e data generally would not support one. Therefore, the Agency has determined that qu antification of risk using a non-li near approach (i.e. the chronic reference dose) will adequately protect for all chr onic toxicity, incl uding carcinogenic ity, likely to result from exposure to cyflumetofen.PC Code: 138831 FINAL

7 I. INTRODUCTION

On December 17, 2013, the Cancer Assessment Revi ew Committee (CARC) of the Health Effects Division of the Office of Pesticides Program met to evaluate the carcinogenic potential of cyflumetofen. Dr. Yung Yang, RAB VI, presented the chronic toxicity/carcinogenicity study in Fischer 344 rats and the carcinoge nicity study in CD-1 mice.

II. BACKGROUND INFORMATION

Cyflumetofen (PC 138831; CAS# 40 0882-07-7) is a new active ingred ient that belongs to the acrylonitrile class of chemicals with a novel insecticidal mode of action – mitochondria complex II electron transport inhibition. It is a selective contac t miticide that is effective on all mite life stages. It is currently proposed only for foliar a pplications via ground or aerial equipment.

The toxicology database is complete for cyflumet ofen. The major target organ in rats, mice, and dogs following short- and long-term oral administration of cyflum etofen is the adrenal glands characterized by increased organ weight and histopathology (vacuolati on and hypertrophy of the adrenal cortical cells). Cyflumetof en has low acute toxicity (acute lethality studies) by oral, dermal, and inhalation routes of exposure. It is minimally irritating to the ey es but not to th e skin. It is a

skin sensitizer.

The toxicity database contains two sets of carcinogenicity studi es in mice and rats. In 2004, the Registrant conducted a carcinogenicity st udy in Fischer 344 rats (IET 01-0125) and a carcinogenicity study in CD-1 mice (IET 02-0039). Since there was no evidence of carcinogenicity in either species, and to address th e adequacy of doses tested in the above studies, the Registrant elected to conduct supplemental st udies at higher doses in mice and rats. This

CARC evaluation includes disc ussion of these four studies.

III. EVALUATION OF CARCINOGENICITY STUDIES

1. Combined chronic toxicity/car cinogenicity Study in F-344 Rats

References : 1. 2004: Carcinogenicity Study in Rats. The Institute of Environmental Toxicology, Ibaraki, Japan. Laboratory report number: IET 01-0125. Study report date: 29-July-2004. Applicant Report Number: OSTA-0046(EN)-FR. DACO 4.4.2. MRID 48542697.

2. 2013: Carcinogenicity Study in Rats. Institut e of Environmental Toxicology; Mitsukaido Ibaraki, Japan. Laboratory report number: IET; Study report Numb er: 11-0002; Report

date: 07-Oct-2001. MRID 49226601PC Code: 138831 FINAL

8 A. Experimental Design

In the 2004 carcinogenicity study (MRID 48542697) , cyflumetofen (OK-5101, 97.67 % a.i.) was administered to Fischer 344 rats (50/sex/dos e) in the diet at dose levels of 0, 150, 500, or 1500 ppm (0, 4.92/6.14, 16.5/20.3, and 49.5/61.9 mg/kg bw/day in males/females) for 104 weeks.

In the 2013 supplementary carcinogenicity study (MRID 49226601), cyflumetofen was administered in the feed to F344 rats (50/se x/dose) at doses of 0 or 6000 ppm (0, 220/287 mg/kg bw/d in males/females) for 104 weeks.

B. Survival Analysis

There were no statistically signifi cant survival dispari ties among the dose groups for male rats in either the 2004 or the 2013 study or in the two studi es combined (Table 1). Because of the lack of survival disparities, the two studies have been co mbined for more robust st atistical tumor analyses.

Table 1. Mortality Rates in F344 Rats Fed Cyflumetofen for 2 years

Cox or Generalized K/W Test Results for the Combined Studies (2004 and 2013)

Dose (ppm) Weeks: 1-26 Weeks: 27-52 Weeks: 53-78 Weeks

79-105 f Total

0 0/100 1/100 2/99 14/97 17/100

(17)

150 0/50 0/50 2/50 8/48 10/50

(20)

500 0/50 0/50 2/50 6/48 8/50

(16)

1500 0/50 0/50 2/50 7/48 9/50

(18)

6000 0/50 0/50 0/50 7/50 7/50

(14)

+Number of animals that died during the interval/Numbe r of animals alive at the beginning of the interval.

f Final sacrifice at week 105. ( ) Percent.PC Code: 138831 FINAL

9 C. Discussion of Tumor Data

Tumors of the thyroid glands and the testes we re seen in male rats only. There were no

treatment-related tumors in female rats. The statis tical analyses of these tumors in the male rats were based upon the Exact test for tr end and the Fisher’s Exact Test.

Thyroid tumors : As shown in Table 2, there was a statis tically significant tr end, and a pair-wise significance at the high dose (bot h at p < 0.05) for thyroid C-cell adenocarcinomas when compared

to controls. There was also a st atistically significant trend (p < 0.05) and a pair-wise significance (p<0.01) for thyroid c-cell adenom as and/or adenocarcinomas combined at the low (150 ppm) and high (6000 ppm) dose groups. It is to be noted that animals in the 150 and 500 ppm dose groups only received microscopic evaluations of the thyr oid if gross lesions were palpated. At the high dose, the incidences of adenomas (31%) were with in the historical contro l range (20-44%) of the testing laboratory; whereas, the incidences of adenocarcinomas (31%) and the combined tumors (57%) exceeded the historical control range (c-c ell adenocarcinoma, 6-18% ), and combined c-cell adenoma/carcinomas, 26-50%).

Table 2. Incidences of Thyroid C-Cell Tu mor Rats in F344 Rats Fed Cyflumetofen

Fisher’s Exact and Exact Tr end Test Results for the Co mbined Studies (2004 and 2013)

0 ppm 150 ppm 500 ppm 1500 ppm 6000 ppm

Adenomas

(%)

p = 20/99 (20)

0.1457 8 a /23 (35)

0.1128 4/15 (27)

0.3903 11/50 (22)

0.4779 15/49 (31)

0.1164

Adenocarcinomas

(%)

p = 15/99 (15)

0.0214* 7/23 (30)

0.0826 2/15 (13)

0.6977 4/50 (8)

0.9375 15 b /49 (31)

0.0253*

Combined

(%)

p = 33 c /99 (33)

0.0123* 15/23 (65)

0.0052** 6/15 (40)

0.4066 14 d /50 (28)

0.8011 28 e /49 (57)

0.0049**

+ Number of tumor bearing animals/Number of animals ex amined, excluding those that died before week 52.

#Animals in the 150 and 500 ppm dose gr oups only received microscopic evaluatio ns of the thyroid if gross lesions were palpated.

a First Adenoma observed at week 98, dose 150 ppm

b First Adenocarcinoma observed at week 94, dose 6000 ppm

c Two animals in the control group had both an adenoma and an adenocarcinoma.

d One animal in the 1500 ppm dose group had both an adenoma and an adenocarcinoma.

e Two animals in the 6000 ppm dose group had both an adenoma and an adenocarcinoma. Note: Significance of trend denoted at control

. Significance of pair-wise comparison with control denoted at dose level.

If * , then p < 0.05. If ** , then p < 0.01.PC Code: 138831 FINAL

10 Table 3. Historical Control Data for Thyroid C-cell Tumors at Testing (IET) Laboratory a

Organs and lesions Study ID A B C D E F G H I b

Year 1997 1997 1998 1998 1998 1998 1999 2003 2003

N 50 50 50 50 50 50 50 50 50 Thyroid: Adenoma, C-cell 8 8 20 13 8 14 12 9 9 (%) (16) (16) (40) (26) (16) (28) (24) (18) (18) Carcinoma, C-cell 0 0 2 4 0 4 2 5 6 (%) (0) (0) (4) (8) (0) (8) (4) (10) (12)

Organs and lesions Study ID J K L M N O P

Year 2005 2005 2008 2009 2009 2011 2012

N 50 50 50 50 50 50 50 Thyroid: Hyperplasia, C-cell (%) --

-- 15 (30) 20 (40) 9 (18) 19 (38) 23 (46) 35 (70) Adenoma, C-cell 16 20 21 22 10 14 18 (%) (32) (40) (42) (44) (20) (28) (36) Carcinoma, C-cell 3 2 7 3 5 9 7
(%) (6) (4) (14) (6) (10) (18) (14)

C-cell Adenoma/carcinoma (combined) (%) 19 (38) 22 (44) 25 (50) 25 (50) 13 (26) 20 (40) 21 (42)

a Taken from page 45 of the study report and additional data submitted by the applicant

b Control data from the original 2-year rat carcinogenicity study (PMRA 2146822, MRID 48542697)

Testicular tumors : As shown in Table 4, for the interst itial cell tumors, there was a significant trend (p < 0.01), and a pair-wise significan ce (p < 0.05) at the hi gh dose (6000 ppm) when compared to controls. However, the incidences of interstitial cell tumors in the current study (82 to 96%) were within the testing laboratories historical control range (56 – 100%). It should be noted that there is a high background rate for this tumor type in the partic ular strain of rat (Fischer344) tested. By contrast,, the incidence rate in human s of LCT is very low (i.e., 0.00004%). Similarly, rats are known to have approximately 13-tim es more LH receptors per cell than humans (Hutaniemi, 1983). Also, there was no corroborativ e increase in non-neoplastic lesions of the testes at the high dose. In fact, there was a decrease in the incidence of interstitial cell hyperplasia at the high dose (3/50; 6%) when compared to controls (19/50; 28%). Consequently, the observance of this tumor type is of que stionable biological significance/relevancePC Code: 138831 FINAL

11 Table 4. Incidence of Testicular Tumo rs in F344 Rats Fed Cyflumetofen

Fisher’s Exact and Exact Tr end Test Results for the Co mbined (2004 & 2013) Studies

0 ppm 150 ppm 500 ppm 1500 ppm 6000 ppm

Interstitial Cell Tumors

(%)

p = 81/99 (82)

0.00565** 42/48 (88)

0.38475 43 a /48 (90)

0.16556 46/50 (92)

0.07560 48/50 (96)

0.01163*

+Number of tumor bearing animals/Numb er of animals examined , excluding those that died before week 52.

a First Interstitial Cell Tumor observed at week 77, dose 500 ppm. Note: Significance of trend denoted at control

. Significance of pair-wise comparison with control denoted at dose level.

If * , then p < 0.05. If ** , then p < 0.01.

Table 5. Historical Control Data for Interstitial Cell Tumors at Testing (IET) Laboratory (1993-2012)

Study Year

1993 1994 1995 1997 1998 1999 2003 2005 2008 2009 2011 2012

Study 1 43/50

86% 42/50

84% 43/50

86% 49/50

98% 39/50

78% 35/50

70% 43/50

86% 42/50

84% 36/50

72% 31/50

62% 28/50

56% 36/50

72%

Study 2 42/50

84% 48/50

96% 42/50

84% 48/50

96% 40/50

80% 43/50

86% 41/50

82%

Study 3 44/50

88% 46/50

92% 39/50

78%

Study 4 50/50

100% 34/50

68%

Year: Year at termination of treatment.

C. Non-Neoplastic Lesions

In the 2004 study (MRID48542697), An increased incidence of diffu se hypertrophy and hyperplasia of adrenal cortical cells was noted in high dose males and females after histopathological examination (T able 6). This finding correlates with the increased adrenal weights at this dose level. In addition, an increas ed incidence of luminal d ilatation of the gland in the uterine horn was observed in mid and high dose females. Both of these findings were considered to be treatment related. High dose male s exhibited a decreased incidence of atrophy of the seminiferous tubules and interstitial cell hype rplasia in the testis. Since the change was a decrease, these observations were not consider ed toxicologically rele vant. No biologically relevant non-neoplastic lesions in the thyroid were observed.PC Code: 138831 FINAL

12 Table 6. Incidence of Selected Non-Neoplastic Lesions (all animals) in Rats Fed Cyflumetofen (2004 study)

Males (ppm)

Organ and lesions 0 150 500 1500

Adrenal

cortical cell hypertrophy (diffuse)

cortical cell hyperplasia N=50

4

13 N=50

2

7 N=50

4

10 N=50

16**

17

Thyroid

Hyperplasia C-cell N=42

19 N=13 a

1 N=7 a

1 N=41

15

Testis

Atrophy, seminiferous tubule

Hyperplasia, interstitial cell N=42

0

18 N=39 a

6

9 N=41 a

4

13 N=41

2

7

Females (ppm)

Organ and lesions 0 150 500 1500

Adrenal

cortical cell hypertrophy (diffuse)

cortical cell hyperplasia

Uterine horn gland luminal dilatation N=50

3

16

6 N=50

3

15

7 N=50

1

14

12 N=50

23**

23

14**

Thyroid

Hyperplasia C-cell N=43

12 N=10 a

2 N=4 a

1 N=45

8

** Significantly different from control, p<0.01

a Examined on the animals that showed macroscopic lesions.

In the 2013 supplementary carcinogenicity study (MRID 49226601), histopathological

examination revealed an increased incidence of diffuse hypertrophy and focal vacuolation of

adrenal cortical cells in treated males and fema les (Table 7). In addition, treated females also

exhibited an increased incidence of diffuse ad renal cortical cell vacuolation. These findings correlate with the increased adrenal weights obser ved in treated animals of both sexes. Treated

males also exhibited an increased incidence of focal atrophy of the acinar cells in the pancreas. Although the incidence was within the range of re ported historical control data values (7/50 – 36/50 for years 2008-2012), th is lesion was also observed in treated males in the 1-year rat chronic toxicity study (MRID 49029701) at the same dose le vel (6000 ppm) and was therefore determined

to be related to treatment. While the inciden ce of atrophy of the semi niferous tubules and hyperplasia of the interst itial cells of the testis were statis tically significantly decreased, these changes were considered to be secondary effects to the development of testicular interstitial cell

tumors.PC Code: 138831 FINAL

13 Table 7: Incidence of Selected Non-Neoplastic L esions (all animals) in Rats Fed Cyflumetofen
(2013 Study)

Organ / Lesion Dose Group (ppm)

Male Female

0 6000 0 6000

N=50 N=50 N=50 N=50

Testis:
Atrophy, seminiferous tubule 27/50 5/50**

- - Hyperplasia, interstitial cell 19/50 3/50**

- - Pancreas: Atrophy, acinar cell, focal 15/50 27/50* 5/50 2/50 Thyroid:
Hyperplasia, C-Cell 17/50 18/50 16/49 14/50

Adrenal: Hypertrophy, cortical cell, diffuse 7/50 44/50** 5/50 39/50** Vacuolation, cortical cell, diffuse 12/50 7/50 3/50 23/50**

Vacuolation, cortical cell, focal 6/50 15/50* 12/50 24/50**
Fisher's exact probability test: **P<0.01; * P<0.05.

D. Adequacy of the Dosing for Assessment of Carcinogenicity

The doses tested were considered to be ade quate to assess the car cinogenic potential of cyflumetofen based on effects on cl inical signs of toxicity (femal es), decreased body weight (both sexes) and body weight gain (males), changes in hematological parameters (males), organ weights (both sexes), histopathology findings (both sexes), and tumor formations at the high dose (6000 ppm) in the supplemental study.

2. Carcinogenicity Study in CD-1 Mice

References : 1. 2004: Carcinogenicity Study in Mice. The Institu te of Environmental Toxicology, Ibaraki, Japan. Laboratory report number: IET 02-0039. Study report date: 27-July-2004. Report Number: OTSA-0061 (EN) -FR. DACO 4.4.3. (MRID 48542698)

2. 2013: BAS 9210 I (Cyflumetofen): Carcinogeni city Study in Mice. The Institute of Environmental Toxicology, Ib araki, Japan. Laboratory report number: IET 11-0016. Study report date: 08-August-2013. Re port Number: 2013/1251154. DACO 4.4.3. (MRID 49209101)PC Code: 138831 FINAL

14 A. Experimental Design

In the 2004 carcinogenicity study (MRID 48542698) , cyflumetofen (OK-5101, 97.67 % a.i.) was administered to CD-1 mice (52/sex/dose) in the diet at dose levels of 0, 150, 500, 1500 or 5000 ppm (0, 15.5/14.3, 54.3/48.1, 156/144, and 537/483 mg/kg/day in males/females) for 78 weeks.

In the 2013 supplementary carcinogenicity study (MRID 4920910), cyflumetofen (OK-5101, 97.82 % a.i.) was administered to SPF ICR (Crj:CD1) mice (52/sex/dose) in the diet at dose levels of 0 and 10,000 ppm (0 and1143/1132 mg/kg bw/day in males/females) for 78 weeks.

B. Discussion of Tumor Data

There was no treatment-related increase in any tumo r type in either sex of mice when compared to

controls.

C. Non-Neoplastic Lesions

In the 2004 carcinogenicity study (MRID 48542698), histopathological examination noted an

increased incidence of diffuse adrenal cortical cell vacuolation in males and females of the 5000 ppm dose group (Table 8). This finding was cons idered to be treatment related due to a corresponding increase in adrenal weight. In addition, an increased incidence of mucosal epithelial cell hyperplasia of the glandular stomach in 150 ppm group males was observed. Since this

increase did not occur in a dos e-related manner and there was no corresponding hist opathological finding at this dose level, this finding wa s considered not toxi cologically relevant.

Table 8. Incidence of Selected Non-Neoplastic Lesions (all animals) in Mice Fed Cyflumetofen

Males (ppm)

Organ and lesions 0

n = 52 150

n = 52 500

n = 52 1500

n = 52 5000

n = 52

Adrenal cortical cell vacuolation (diffuse) 1 3 2 3 8*

Females (ppm)

Organ and lesions 0

n = 52 150

n = 52 500

n = 52 1500

n = 52 5000

n = 52

Adrenal cortical cell vacuolation (diffuse) 7 4 4 9 26**

* Significantly different from control, p<0.05 ** Significantly different from control, p<0.01PC Code: 138831 FINAL

15 In the 2013 supplementary carcinogenicity study (MRID 49209101), histopathological

examination revealed an increased incidence of diffuse adrenal cortical cell vacuolation and deposition of brown pigment in the cortico-medulla ry junction in treated males and females (Table 9). These findings were considered to be treatm ent-related due the magnit ude of the increase in both sexes and the corresponding increase in adrenal weight in females. In addition, an increased incidence of extramedullary hematopoiesis was observed in the spleen of treated females.

Table 9. Incidence of Selected Non-Neoplastic Lesions (all animals) in Mice Fed Cyflumetofen

0 mg/kg/day 1143 mg/kg/day

Males (52/dose)

Adrenal cortical cell vacuolation (diffuse) 0 21**

Brown pigment deposition, co rtico-medullary junction 6 14**

0 mg/kg/day 1132 mg/kg/day

Females (52/dose)

Adrenal cortical cell vacuolation (diffuse) 2 41**

Brown pigment deposition, co rtico-medullary junction 27 44**

a Data obtained from page 140 in the study report * Significantly different from control, p<0.05 ** Significantly different from control, p<0.01

D. Adequacy of Dosing for Assessment of Carcinogenicity

The dose tested in the supplementary study wa s the Limit Dose (10,000 ppm or 1000 mg/kg/day) and was considered to be adequate to asse ss carcinogenicity of cyflumetofen in mice. Treatment-related findings at this dose included: clinical signs of toxi city, increased adrenal weights, enlarged spleen, increased incidence of extramedularry hematopoies is in the spleen of treated females, increased incidence of diffuse ad renal cortical cell vacuolation, and deposition of brown pigment in the cortico-medullary junc tion of the adrenals in both sexes.

IV. TOXICOLOGY

1. Metabolism

There were a series of metabolism st udies conducted for cyflumetofen. 14 C-cyflumetofen was rapidly absorbed following administration of the lo w dose, with plasma levels of radioactivity peaking at 1 hour post-dosing. Absorption was s lightly slower following administration of the high dose, with peak plasma levels attained at 2-4 hours post-dosing i ndicating saturation of absorption. At the high dose, females demonstrated higher C

max and area under the curve (AUC)

values than males. At 72 hours post-dosing, the gastroin testinal tract, carcass, liver a nd kidney contained the highestPC Code: 138831 FINAL

16 proportions of radioactivity. Most of the radioactivity excreted in urine from low dose rats (90-96%) was eliminated within 24 hours after do sing, whereas at the high dose, excretion was slower with only 74-88% elimin ated within 24 hours of dosing. At both low and high doses, 60-89% of the radioactivity excreted in f eces was eliminated within 24 hours of dosing.

The unchanged parent was not detected in th e urine of any test group. Metabolite AB-3 was identified as a major metabolite in the urine of female rats but was a minor urinary metabolite in

males. Metabolite AB-2 was a minor metabolite in urine of all test groups except females from the low dose group, in which residues for this metabo lite reached 4% of the administered dose. The major metabolites detected in the bile from all test groups we re the glucuronic acid conjugates

of metabolites AB-1 and AB-3. Metabolite AB-2 wa s also detected at lower levels in the bile from all test groups. Unique metabolites identified in the bile of rats administered the A-ring or B-ring label included the glucuron ic acid conjugate of A-6 and th e glutathione conjugate of B-1, respectively.

14 C-cyflumetofen was metabolized in the rat primarily by hydrolytic cleavage of the trifluoromethylbenzoyl moiety resulting in metabo lite B-1 (trifluoromethylbenz oic acid) and A-18. Another minor pathway included successive hyd roxylation of the tert -butyl side chain.

2. Mutagenicity :

Cyflumetofen, two metabolites (B-1 and B-3) and one impurity (AB-13) were evaluated in a comprehensive battery of Genetic Toxicology assays . The data indicate that neither the parent compound nor metabolite B-1 were mutagenic in b acteria. However, metabolite B-3 induced a weak but reproducible and concen tration-related mutagenic respons e (1.7- 2.2 fold increases at 3330 and 5000 μg/plate) in Salmonella typhimurium TA100; the effect was, nevertheless, abolished in the presence of exogenous metabo lic activation. There was also evidence of a mutagenic response in the L5178Y mouse lymphoma assay that was induced by the parent and the two metabolites. The response was the strongest in the parent compound (90 μg/mL –S9 vs. the limit concentration for the two metabolites). Ho wever, for both the parent and metabolite B-1, mutagenic activity was only seen at se verely cytotoxic co ncentration (i.e., ≈ 10% survival); consequently, these results are not considered sufficient evidence of a mutagenic response in this test system. For metabolite B-3, the 4.3-fold in crease in the mutation fr equency at 1891 μg/mL (equivalent to the limit concentration of 10 mM) was confined to this level and not observed in the S9-activated portion of the assay.

For the remaining genetic endpoints, the parent and the two metabolites were not clastogenic in Chinese hamster lung cells (parent in two studi es performed by independ ent laboratories) or peripheral human lymphocytes (metabolites B-1 and B-3). Similarly, cyflumetofen was not clastogenic or aneugenic in mouse bone marrow cel ls up to the limit dose. Finally, neither the parent nor the two metabolites i nduced a genotoxic response in hepa tocytes harvest from male rats orally exposed up to the li mit dose (parent and metabolite B-1) or an overtly toxic level for the test animals (metabolite B-3).PC Code: 138831 FINAL

17 Studies performed on an impurity of cyflumetofen (AB-13) showed negati ve results for reverse gene mutations in bacteria ( S. typhimurium TA 1535, TA 1537, TA 98 or TA 100 or Escherichia

coli WP2 uvr A), forward gene mutations in mamma lian cells (L5178Y mous e lymphoma cells),

and chromosome aberrations in mammalian cells (human periphe ral lymphocytes).

Overall, the data indicate th at while there was some evidence of a mutagenic response from in vitro assays, the responses were confined to severe ly cytotoxic concentrations, abolished in the presence of exogenous metabolic activation, or were not expresse d in whole animal studies. Based on these considerations, there is no co ncern for mutagenicity for the parent or the

metabolites . Similarly, the impurity was uniformly nega tive; hence, there is also no concern for mutagenicity. All assays were well-conducted, ac ceptable and satisfy FIFRA test guidelines for mutagenicity testing. Consequently, mutagenicity can be ruled out as a possible mode of action

for tumor formation. Results from thes e assays are summarized below:

Parent

Cyflumetofen was negative in bacteria in two independent trials using the pre-incubation method

at concentrations of 20.6 to 5000 μg/plate +/-S9 (MRID 48542687).

In the mouse lymphoma forward gene mutation as say, concentrations of 20 to 90 μg/mL –S9 or 10 to 140 μg/mL +S9 for 3 hours. Although increase s in the total mutation frequencies (MF) occurred at 90 μg/mL –S9 (3.2-fold) and at 140 μg /mL +S9 (6.1-fold), these concentrations were insoluble and severely cytotoxic (relative suspen sion growth (RSG) was 10% at 90 μg/mL –S9 and 12% at 140 μg/mL +S9). These results are, ther efore, not considered sufficient evidence of a mutagenic response in this test system (MRID 48542692).

In an in vitro chromosome aberration study, Chinese hams ter lung fibroblasts (V79) cells were exposed to 6.25 to 50 μg/mL –S9 and 25 to 200 μg/mL +S9 for 6 hours and allowed an 18-hour recovery period. Cells were also continuously treated with 3.75 to 30 μg/mL –S9 for 24 or 48

hours and harvested at the end of treat ment. Compound precipitation was seen at ≥ 200 μg/mL

+S9. Less than 50% of the cells were available for analysis at the highest dose tested with or without S9 activation. There were however, and no significant increases in cells with structural or numerical chromosome aberrations (MRID 48542689).

In a second in vitro chromosome aberration study, Chinese hamster lung fibroblasts (V79) cells were exposed to 2.5 to 80 μg/mL –S9 and 10 to 160 μg/mL +S9 for 4 hours and allowed a 14-hour recovery period. Due to severe cytotoxicity, th e assay was repeated with 3.1 to 10 μg/mL –S9. Cells were also continuously treated with 3.1 to 40 μg/mL –S 9 for 18 hours and harvested or allowed an additional 10 hour recovery. With S9, cells were dosed with 20-320 μg/mL for 4 hours and harvested after a 24-hour recovery period. Compound precip itation was seen at ≥ 120 μg/mL. Less than 50% of the cells were available for analysis at ≥ 160 μg/mL +S9, ≥ 20 μg/mL-S9 (18-hour exposure), or 40 μg/mL-S9 (18-hour exposure with 10-hour recovery). There were however, and no signi ficant increases in cells w ith structural or numerical chromosome aberrations (MRID 48542691).PC Code: 138831 FINAL

18 In the mouse bone marrow micronucleus assay, levels up to the limit dose were not overtly toxic to the test animals, cytot oxic to the target cells or induce an aneugenic or a clastogenic response (MRID 48542694).

In an in vivo unscheduled DNA synthesis ( UDS) assay in male Wistar rats, metabolite B-1 was not overtly toxic to the test animals or genotoxic in the hepatocytes harvested 2-4 or 12-16 hours after oral administration of levels up to the limit dose of 2000 mg/kg (MRID 48542695).

Metabolite B-1

Metabolite B-1 was negative in bacteria in two i ndependent trials using the plate incorporation method at concentrations of 3 to 5000 μg/plate +/-S9 (MRID 48542710).

In the mouse lymphoma forward gene mutation assay, concentrations of 1 to 1901 μg/mL +/–S9 (1901 μg/mL is equivalent to the limit concentr ation of 10 mM) were tested using a 3-hour exposure. A second assay was performed with comparable nonactivated concentrations and a 24-hour exposure and S9-activated levels of 3-1910 μg/mL with a 3-hour exposure. The nonactivated assay was repeated for the 24-hour exposure with 3- 1910 μg/mL. The 3-hour exposure with or without S9 activation was nega tive up to the highest dose tested (HDT). Although marked increases in the total MF occurred at the HDT without S9 activation after the

24-hour treatment in both the initial (8.2-fold) and repeat trials (7-f old), these concentrations were severely cytotoxic (RSG was 13 or 9%, respecti vely). Consequently, these results are not considered sufficient evidence of a mutagenic response in this test system (MRID 48542712).

In an in vitro chromosome aberration study, peripheral human lymphocytes were exposed to 333 to 1901 μg/mL +/–S9 (1901 μg/mL is equivalent to the limit concentration of 10 mM) for 3 hours and allowed a 21-hour recovery period. Cells we re also continuously tr eated with 100 to 1000 μg/mL –S9 for 24 or 48 hours and ha rvested at the end of exposure. With S9, cells were dosed with 333-1901 μg/mL for 3 hours and harvested afte r 48 hours. There were no significant increases

in cells with structural chromosome aberrations without S9 activation up to concentrations that

caused a ≥ 50% decrease in relative suspension growth ( ≥ 750 μg/mL, prolonged 24-hour exposure or ( ≥ 560 μg/mL, prolonged 48-hour exposure). Sim ilarly, no evidence of a clastogenic response was seen after exposure for 3 hours with or with S9 up to the limit dose (MRID 48542711).

In an in vivo unscheduled DNA synthesis ( UDS) assay in male Wistar rats, metabolite B-1 was not overtly toxic to the test animals or genotoxic in the hepatocytes harvested 2-4 or 12-16 hours after oral administration of levels up to the limit dose of 2000 mg/kg (MRID 48542713).

Metabolite B-3 [2-(trifluoromethyl)benzamide]

Metabolite-3 induced a reproduc ible and dose-related increas e in mutant colonies of Salmonella typhimurium TA100. Fold increases were 1.7 and 1.9-fold at 3330 and 5000 μg/plate –S9 in Trial 1 and 1.7 and 2.2-fold at 3330 and 5000 μg/plate –S9 in Trial 2. The response was, however, abolished in the presence of S9 activ ation and was not seen in any other Salmonella strain or in

Escherichia coli WP2 uvrA (MRID 48542715).PC Code: 138831 FINAL

19 In the mouse lymphoma forward gene mutation assay, concentrations of 1 to 1891 μg/mL +/–S9 (1891 μg/mL is equivalent to the limit concentr ation of 10 mM) were tested using a 3-hour exposure. A second assay was performed with comparable nonactivated concentrations and a 24-hour exposure and comparable S9-activated c oncentrations with a 3-hour exposure. The 3-hour exposure with or without S9 activation was negative up to the highest dose tested (HDT).

A marked increase in the total MF occurred at the HDT without S9 activation (4.3-fold) following the prolonged and in the absence of severe cytoto xicity. However, the response was confined to

this concentration and not observed when S9 activ ation was incorporated in to the reaction mixture (MRID 48542717).

In an in vitro chromosome aberration study, peripheral human lymphocytes were exposed to 333 to 1891 μg/mL +/–S9 (1891 μg/mL is equivalent to the limit concentration of 10 mM) for 3 hours and allowed a 21-hour recovery period. Cells we re also continuously tr eated with 100 to 1891 μg/mL –S9 for 24 hours or 33 to 1891 μg/mL –S9 for 48 hours. Cells were harvested at the end of the prolonged exposures. With S9, cells were dosed with 333-1891 μg/mL for 3 hours and

harvested after 48 hours. There were no significant increases in cells with structural chromosome aberrations with or without S9 up to the limit concentrations (MRID 48542716).

In an in vivo unscheduled DNA synthesis (UDS) assay in male Wistar rats, metabolite B-3 was

toxic to the test animals at 350 mg/kg but was not genotoxic in th e hepatocytes harvested 2-4 or

12-16 hours after oral administrati on of 0, 175 or 350 mg/kg (MRID 48542718).

Impurity AB-13

Impurity AB-13 was negative in bacteria in two i ndependent trials using the plate incorporation method up to insoluble concentrations ( ≥ 333 μg/plate +/-S9) (MRID 48542688).

In the mouse lymphoma forward gene mutation a ssay, concentrations of 0.03 to 100 μg/mL +/–S9 were tested using a 3-hour expos ure. A second assay was performe d with similar concentrations and a 24-hour exposure for the nonactivated phase of testing and with a 3-hour exposure for the S9-activated concentrations. The 3-hour exposur e with or without S9 activation was negative up to the highest dose tested. Compound precipita tion was seen at 100 μg/mL +/–S9. RSG was generally similar to the control at all concentr ations and exposure intervals. There was no appreciable increase in the MF at any non activated or S9-activated concentration (MRID 48849413).

In an in vitro chromosome aberration study, peripheral human lymphocytes were exposed continuously to 10 to 333 μg/mL –S9 for 24 or 48 hours and harvested at the end of the prolonged exposures. With S9, cells were dosed with 10 to 333 μg/mL for 3 hours and harvested after 48 hours. Compound precipitation was noted at ≥ 100 μg/mL. There were no adverse effects on the mitotic indices or significant increases in cells with structural chromosome aberrations with or without S9 up to insoluble levels (MRID 48542716690).PC Code: 138831 FINAL

20 3. Structure-Activity Relationship

No structure-activity relationship information is available for cyflumetofen in the OncoLogic

expert system.

4. Subchronic and Chronic Toxicity

a) Subchronic Toxicity in Rats

In a subchronic toxicity study (MRID 48542682), cyflumetofen (OK-5101, 97.6% a.i.) was

administered to 10 Fischer rats/sex/dose in the diet at dose levels of 0, 100, 300, 1000 and 3000 ppm (0, 5.4, 16.5, 54.5 and 167 mg/kg//day in ma les, 0, 6.28, 19.0, 62.8 and 193 mg/kg bw/day in females) for 90 days.

Indications of liver effects we re seen. OK-5101 caused an increase in prothrombin time in high dose males. In females, increases observed in albumin/globulin ratio in the 1000 and 3000 ppm groups were correlated to the decreases in obser ved globulin levels. Organ examination revealed that liver weights were increased in both males and females at the high dose and in the 1000 ppm males. At necropsy, all females in the 3000 ppm group had white and enlarged adrenals. In addition, adrenal weights were increased in the 1000 and 3000 ppm females. Histopathology

examinations found an increase of diffuse vacuola tion and diffuse hypertrophy of cortical cells in the adrenals of all 1000 and 3000 ppm animals.Rela tive kidney weights we re increased in high dose males and females. Eight of ten high dose fe males were observed to have vacuolation of interstitial gland cells in their ovaries.

The LOAEL is 1000 ppm (54.5 mg/kg/day in male s, 62.8 mg/kg/day in females) based on the hematology and organ weight change s in the liver, organ weight and histopathology effects in the adrenals, organ weight increase in the kidneys and histopathological effects observed in the ovaries. The NOAEL is 300 ppm (16.5 mg/kg/da y in males, 19.0 mg/kg/day in females).

b) Subchronic Toxicity Study in Mice

In a subchronic toxicity study (MRID 48542683), cyflumetofen (OK-5101, 97.7% a.i.) was

administered to 10 SPR ICR mice/sex/dose in diet at dose levels of 0, 300, 1000, 3000 and 10000 ppm (0, 35.4, 117, 348 and 1200 mg/kg/day in ma les; 0, 45.0, 150, 447 and 1509 mg/kg/day in females) for 90 days.PC Code: 138831 FINAL

21 Body weight and food consumption were decreased at ≥ 3000 ppm in females. Organ weight examinations revealed increased adrenal weight s in high dose males. During necropsy, 9 of 10 male mice in the 10000 ppm exhibited enlarged ad renals. Histopathological findings showed an increased incidence of diffuse hypertrophy of co rtical cells in male s at the 3000 and 10000 ppm

doses. Two and seven females exhibited diffuse vac uolation of cortical cells in the adrenals at 3000 and 10000 ppm, respectively. The degree of sever ity and incidence of the lesion appeared in a dose-related manner, indicati ng a treatment-related effect.

The LOAEL is 3000 ppm (348 mg/kg/day in ma les; 447 mg/kg/day in females), based on decreased body weights in females, and diffuse hypert rophy and vacuolation of cortical cells in the adrenals in both sexes. The NOAEL is 1000 ppm (117 mg/kg/day in males; 150 mg/kg/day in females).

c) Chronic Toxicity in Rats

In a chronic toxicity study (MRI D 48542696), cyflumetofen (OK-5101, 97.67%) was administered to Fischer (F344/DuCrj) rats (50/sex/ dose) in the diet at dose levels of 0, 50, 150, 500 and 1500 ppm (equivalent to 0/0, 1.90/2.31, 5.63/6.92, 18.8/23.3 and 56.8/69.2 mg/kg bw/day in males/females, respectively) for one year. Interim sacrifices (10/sex/dose) were performed at 4, 13 and 26 weeks. All remaining animals were sacrificed at 52 weeks.

There were no adverse treatment-re lated effects on mortality, c linical signs, body weight, food consumption, haematology, clinical chemistry and ur inalysis parameters. Treatment related effects

were noted in the adrenals of both sexes at the hi gh dose. In females, absolute and relative adrenal weights were increased at all scheduled sacr ifices, along with diffuse hypertrophy of adrenal cortical cells. Effects in males were slightly differ ent with diffuse vacuolation of the cortical cells

recorded at all interim and termin al sacrifices. An increased inci dence of vacuolation of ovarian interstitial gland cells was obs erved in high dose females from week 13 on, along with increased ovarian weights at terminal sacrif ice. An increased incidence of atrophy of the seminal vesicles and coagulating gland were recorded in high dose males at terminal sacrifice only. There were no corresponding effects observed during histopathological examination.

The LOAEL for chronic administration of cyflum etofen in rats is 1500 ppm (equivalent to 56.8/69.2 mg/kg/day in males/female s, respectively) based on diffuse vacuolation of adrenal

cortical cells and increased incide nce of atrophy of the seminal vesi cles and coagula ting glands in

males and increased adrenal wei ghts and ovarian weights, diffuse hypertrophy of adrenal cortical

cells and vacuolation of the ovarian interstitia l gland cells in females. The NOAEL is 500 ppm (equivalent to 18.8/23.3 mg/kg/day in males/females, respectively).PC Code: 138831 FINAL

22 d) Carcinogenicity in Rats (2004 study)

In a carcinogenicity study (MRID 48542697), OK-5101(97.67 % a.i.) was administered to 50

Fischer rats/sex/dose in the diet at dose levels of 0, 150, 500, and 1500 ppm (0, 4.92/6.14, 16.5/20.3, and 49.5/61.9 mg/kg/day in males/female s) for 104 weeks.There were no compound related effects on mortality, clinical signs, body weight, food consumption, hematology or gross necropsy observations. Diffuse hypertrophy of the adrenal cortex was noted in high dose males and females in conjunction with increased adrenal we ights. In addition, luminal dilatation of the gland of the uterine horn was observed in high do se females. At the doses tested, there was no

treatment related increase in tumor incidence when compared to controls. The LOAEL is 1500 ppm (49.5/61.9 mg/kg/day), based on increased adrenal weight s, hyperplasia and hypertrophy of the adrenal cortex in both sexes, and luminal dilatation of the gland in the uterine horn in females. The NOAEL is 500ppm (16.5/20.3 mg/kg/da y). There was no treatment related increase in tumor incidence when compared to controls.

e) Carcinogenicity in Rats (2013 study)

In the 2013 study (MRID 49029701), OK-5101 (cyflumeto fen, 97.82 % a.i.) was administered to SPF/VAF Fischer rats (F344/DuCrj) (50/sex/dose) in the diet at dose levels of 0 and 6000 ppm (0 and 250/319 mg/kg/day, ♂ / ♀ ) for 52 weeks. Interim sacrifices (10/sex/dose) were performed at 4, 13, 26 weeks. All remaining animals were sacrificed at 52 weeks. There were no

treatment-related deaths or eff ects on food consumption, food effici ency, or in the ophthalmologic assessment. In treated females, an increased incidence of soiled fur around the external genital

area and a consistent decrease in body weight and body weight gain when compared to the control group were observed throughout most of the st udy. Males of the 6000 ppm group had reduced body weight gain throughout the st udy. In treated animals of both sexes, affected hematological parameters included decreased platelet count and fibrinogen concen tration and elongated prothrombin time and activated partial thrombopla stin time (APTT). Blood urea nitrogen (BUN)

was elevated from 26 and 52 week in males and at all sampling points in females and was slightly outside of the submitted historical control data range for the females. This observation was considered treatment-related in conjunction with in creased kidney weights for the treated animals. In both sexes at 6000 ppm, absolu te and relative liver, kidney and adrenal weights were increased at all sacrifice time points. Gros s necropsy revealed an increase in the incidence of white colour in the adrenal gland in both sexes at 6000 ppm as well as adrenal enlargement in females. Treatment-related histological obse rvations included effects noted in the adrenal gland such as diffuse hypertrophy of the cortical cell in treated females and diffuse cortical cell vacuolation in

treated males and focal vacuolation of the cortical cell in treated females with higher incidence of these lesions by the end of the study. In treated males, gross lesions in the testis such as atrophy, softening, and spots were observed as well as an in creased incidence of inte rstitial cell hyperplasia. Atrophy of the acinar cell of the pancreas was also noted in the treated males. In treated females,

vacuolation of the inte rstitial gland cell of the ovary was observed. The LOAEL was 6000 ppm (250/319 mg/kg/day, ♂ / ♀ ) based on increased adrenal weights accompanied by gross pathology and histopathology lesi ons noted in both sexes, vacuolation of the interstitial gla nd cell of the ovary and hyperp lasia of the inters titial cell of th e testes. A NOAEL was not set in this study.PC Code: 138831 FINAL

23 f) Carcinogenicity Study in Mice (2004 study)

In a carcinogenicity study, OK-5101(97.67 % a.i.) was administered to 52 CD-1 mice/sex/dose in the diet at dose levels of 0, 150, 500, 1500 and 5000 ppm (0, 15.5/14.3, 54.3/48.1, 156/144, and 537/483 mg/kg/day in males/females) for 78 weeks.

There were no compound-related effects on mo rtality, clinical signs, body weight, food consumption, hematology or gross necropsy observ ations at any dose level. Diffuse adrenal

cortical cell vacuolation was not ed in 5000 ppm males and females, with a corresponding increase in adrenal weight in males. At the doses teste d, there was no treatment re lated increase in tumor incidence when compared to controls.

The LOAEL is 5000 ppm (537/483 mg/kg/day), based on an increased incidence of diffuse adrenal

cortical cell vacuolation in male s and females, and an increase in adrenal weight in males. The NOAEL is 1500 ppm (156/144 mg/kg/day). There was no treatment related increase in tumor incidence when compared to controls.

g) Carcinogenicity Study in Mice (2013 study)

In the 2013 carcinogenicity study, BAS 9210 I (cyflu metofen, 97.82 % a.i.) was administered to SPF ICR (Crj:CD1) mice (52/sex/dose) in the diet at dose levels of 0 and 10,000 ppm (0 and

1143/1132 mg/kg/day in males/females) for 78 weeks.

There were no compound-related effects on mortality, body weight, food consumption, or hematology parameters at the dose tested. Diffuse adrenal cortical cell va cuolation and deposition of brown pigment in the cortico-medullary junc tion of the adrenals was observed in treated

animals of both sexes. Treated females exhi bited a significant increase in the incidence of pale-colored skin and pale-colored eye/eyelid, increased adrenal we ights, enlarged spleen and an increased incidence of extramedullary hematopoiesis in the spleen.

The LOAEL for this confirmatory study is 10,000 ppm (1143/1132 mg/kg/day), based on clinical signs of toxicity, increased adre nal weights, enlarged spleen and an increased incidence of extramedullary hematopoiesis in the spleen of treated females and an increased incidence of diffuse adrenal cortical cell vacuolation and depo sition of brown pigment in the cortico-medullary junction of the adrenals in both sexes. A NOAEL was not identified for this study.

At the dose tested, there was no treatment-related increase in tumor incidence when compared to controls. Dosing was considered adequate as animals were dosed above the limit dose of 1000 mg/kg/day.PC Code: 138831 FINAL

24 5. Mode of Action Studies

There is no mode of action (MOA) data availabl e for thyroid c-cell adenoma/adenocarcinoma at this time. However, MOA data were submitted fo r the Leydig cell tumors (LCT). The CARC did not review the MOA for the LCTs since the MOA for development of this lesion depends on transient, rat-specific, hormonally mediated mechan isms that have low relevance to humans due to the qualitative and quantitative differen ces between rat and human Leydig cells( Boorman et al. 1990).

V. COMMITTEE’S ASSESSMENT OF THE WEIGHT-OF-THE-EVIDENCE

1. Carcinogenicity

The carcinogenic potential of cyflumetofen was evaluated following dietary administration

to male and female Fischer 344 rats. Two studies were conducted in the rats. One study was conducted at doses of 0, 150, 500 and 1500 ppm wh ile a second high dose study was conducted at 0 and 6000 ppm. There was no evidence of carcinogenici ty in female rat at any dose level. There was limited evidence of carcinogenic ity characterized by the presence of thyroid c-cell tumors and

interstitial cell tumors of the test ed in male rats only at the hi ghest dose tested (HDT). The doses tested were determined to be adequate to asse ss the carcinogenic potential of cyflumetofen in male

and female rats.

The thyroid c-cell tumors were determined to be treatment-related based on the following considerations: a statistically significant trend, and a pair-wise significance (both at p < 0.05) for thyroid c-cell adenocarci nomas when compared to controls; a statistically significant trend (p < 0.05) and a pair-wise significance (p<0.01) for thyr oid c-cell adenomas and/or adenocarcinomas combined; and the incidences of the adenocar cinomas (31%) and the combined tumors (57%)

exceeded the historical control ra nge of the testing laboratory (c -cell adenocarcinoma, 6-18%, and combined c-cell adenoma/carcinomas, 26-50%).

The testicular interstitial cell tumors, although exacerbated by treatme nt were not considered to be biologically relevant since this tumor (Leydig ce ll, LCT) are extremely common in the F344 rat, with a reported background incidence of 56-100% in 2-year cancer studies. The reasons for the different background incidence rate for LCT am ong species stem from both quantitative and qualitative differences in the Leydig cell respons e to hormonal stimuli. The rat Leydig cell is extremely sensitive to slight changes in circul ating luteinizing hormone (LH) levels. Among rat strains, the F344 rat is particular ly susceptible to this process a nd is not considered a relevant model for studying LCT. Because of the differences present between the human and the rat Leydig cells, non-genotoxic compounds that cause LCT in ra ts have little relevance to humans. By contrast, the incidence rate in humans of LCT is very low (i.e., 0.00004%). Similarly, rats are known to have approximately 13-times more LH receptors per cell than humans (Hutaniemi, 1983).PC Code: 138831 FINAL

25 The data with Cyflumetofen supports this specie s and strain-specific effect on the Leydig Cells. Other than at a high dose with th e F344 rat, there was no treatmen t related effects observed on the Leydig cell in other tested rat strains (Wistar) an d species (dog, mouse), no matter the dose level or the duration of dosing.

The carcinogenic potential of cyflumetofen wa s evaluated following dietary administration to

male and female CD-1 mice. Two studies were conducted in the mice. One study was conducted at doses of 0, 150, 500 and 5000 ppm while a second high dose study was c onducted at 0 and 10,000 ppm. There was no evidence of carcinogenicity in male or female mice at any dose level. The

doses tested were determined to be adequate to assess the carci nogenic potential of cyflumetofen

in male and female mice.

2. Mutagenicity

The mutagenic potential of cyflumetofen, its metabolites, and an impurity was tested in in vitro

and in vivo studies. There was no concern for mutagenic ity for the parent, the metabolites or the impurity. Consequently, mutagenicity can be ruled out as a possible m ode of action for tumor

formation.

3. Structure Activity Relationship

No structure-activity relationship information is available in the OncoLogic expert system.

4. Mode of Action

No studies were submitted to evaluate the MOA for thyroid tumors. However, MOA data were submitted for the Leydig cell tumors (LCT). The MOA for the Leydig cell tumors was not reviewed since this tumor type has a high backgr ound rate in the Fischer 344 rats and is of low

relevance for humans

VI. CLASSIFICATION OF CARCINOGENIC POTENTIAL

In accordance with the EPA's Final Guidelines for Carcinogen Risk Assessment (March, 2005), the Cancer Assessment Review Committee (CARC) classified cyflumetofen as “Suggestive Evidence of Carcinogenic Potential”. This classifi cation is based on the presence of a single tumor type (thyroid c-cell) in one sex (male) and one species (rat), and the lack of concern for mutagenicity.

VII. QUANTIFICATION OF CARCINOGENIC POTENTIAL

When there is suggestive evidence of car cinogenicity, the Agency does not attempt a dose-response assessment as the nature of the da ta generally would not su pport one. Therefore, the Agency has determined that quantification of ri sk using a non-linear approach (i.e. the chronic reference dose) will adequately protect for all chr onic toxicity, incl uding carcinogenic ity, likely to result from exposure to cyflumetofen.PC Code: 138831 FINAL

26 VIII. BIBLIOGRAPHY

MRID CITATION

48542682 Repeated Dose 90-day Oral Toxicity Study in Rats. The Institute of Environmental Toxicology. Laboratory report number: IET 01-0061. Study report date: 17-June-2004. Applicant Report Number: OTSA-0014 (EN)-FR. DACO 4.3.1 48542683 Repeated Dose 90-day Toxicity Study in Mice. The Institute of environmental Toxicology. Laboratory report number: IET 01-0111. Study report date: 21-June-2004. Applicant Report Number: OTSA-0042 (EN)-FR.DACO 4.3.1. 48542696 Repeated Dose 1-year Oral Toxic ity Study in Rats. The Institute of Environmental Toxicology, Ibaraki, Japan. Laborato ry report number IET 01-0124. June 22, 2004. DACO 4.4.1 48542697 Carcinogenicity Study in Rats. The Institute of Environmental Toxicology, Ibaraki, Japan. Laboratory report nu mber: IET 01-0125. Study report date: 29-July-2004. Applicant Report Number: OSTA-0046(EN)-FR. DACO 4.4.2. 48542698 Carcinogenicity Study in Mice. The In stitute of Environmental Toxicology, Ibaraki, Japan. Laboratory report number: IET 02-0039. Study report date: 27-July-2004. Applicant Report Number: OTSA-0061 (EN)-FR. DACO 4.4.3. 49226601 BAS 9210 I (Cyflumetofen): Carcinogenic ity Study in Rats. The Institute of Environmental Toxicology, Ibarak i, Japan, October 7, 2013. IET 11-0002 49209101 BAS 9210 I (Cyflumetofen): Carcinogenic ity Study in Mice. The Institute of Environmental Toxicology, Ib araki, Japan. Laboratory report number: IET 11-0016. Study report date: 08-August-2013. Applicant Report Number:

2013/1251154. DACO 4.4.3.

48542687 Bacterial Reverse Mutation Test. The Institute of Envir onmental Toxicology, Ibaraki, Japan. Laboratory report nu mber: IET 01-0033. Study report date: 06-August-2001. Applicant Report Number: OTSA-0009(EN)FR. DACO 4.5.4. 48542691 BAS 9210 I (Cyflumetofen) – In Vitro Chromosome Aberration Assay in V79 Cells. Experimental Toxicology and Ecology, BASF SE, Germany. Laboratory report number: 32M0510/094436. Study report date: 25-January-2011. Applicant Report Number: 2010/1151897. DACO 4.5.6.

48542689 OK-5101: In Vitro Mammalian Chromosome Aberration Test. The Institute of Environmental Toxicology, Ibaraki, Japan. Laborato ry report number: IET 01-0034. Study report date: 15-December-2003. Applicant Report Number: OTSA-0011 (EN)-FR. DACO 4.5.6. 48542692 Evaluation of the Mutagenic Activity of OK-5101 in an In Vitro Mammalian Cell Gene Mutation Test with L5178Y Mouse Lymphoma Cells. Noton B.V., The Netherlands. Laboratory report num ber: 485622. Study report date:

06-November-2007. Applicant Report Number: OTSA-0362-FR. DACO 4.5.5.

48542694 OK-5101: Micronucleus Test in Mice. The Institute of Environmental Toxicology. Laboratory report numb er: IET 01-0035. Study report date: 16-December-2003. Report Number: OTSA-0019(EN)-FR. DACO 4.5.7. 48542695 Evaluation of DNA Repair Inducing Ab ility of OK-5101 in Male rat Hepatocytes ( In Vivo Rat Hepatocyte DNA-Repair Assa y). Notox B.V., Hertogenbosch, ThePC Code: 138831 FINAL

27 Netherlands. Laboratory report num ber: 486284. Study report date:

30-October-2007. Applicant Report Number: 2007/1067386. DACO 4.5.8. 48542710 Evaluation of the Mutagenic Activity of B-1 in the Salmonella typhimurium Reverse Mutation Assay and the Escherichia coli Reverse Mutation Assay (With Independent Repeat). Notox B.V., He rtogenbosch, The Netherlands. Laboratory report number: 396113. Study report da te: 15-June-2004. Applicant Report

Number: OTSA-0171. DACO 4.5.4. 48542712 Evaluation of the Mutagenic Ability of B-1 in an In Vitro Mammalian Cell Gene Mutation Test with L5178Y Mouse Lympho ma Cells (With Independent Repeat). Notox B.V., Hertogenbosch, The Nether lands. Laboratory report number: 396179. Study report date: 22-December-2004. Applicant Report Number:

OTSA-0172. DACO 4.5.5. 48542711 Evaluation of the Ability of B-1 to Induce Chromosome Aberrations in Cultured Peripheral Human Lymphocytes. Notox B.V., Hertogenbosch, The Netherlands. Laboratory report number: 396146. Study report date: 21-June-2004. Applicant Report Number: OTSA-0173. DACO 4.5.6. 48542713 Evaluation of DNA Repair Inducing Ability of B-1 in Male Rat Hepatocytes ( In

Vivo Rat Hepatocyte DNA-Repair Assa y). Notox B.V., Hertogenbosch, The Netherlands. Laboratory name. Laborat ory report number: 490348. Study report date: 23-April-2009. Applicant Report Number: 2009/1126680. DACO 4.5.8. 48542715 Evaluation of the Mutagenic Activity of 2-(Trifluoromethyl) benzamide, B-3 in

the Salmonella typhimurium Reverse Mutation Assay and the Escherichia coli Reverse Mutation Assay (With Inde pendent Repeat). Notox B.V., Hertogenbosch, The Netherlands. La boratory report number: 489612. Study report date: 23-April-2009. Applicant Report Number: 2009/1126681. DACO

4.5.4. 48542716 Evaluation of the Ability of 2-(Trifluoromethyl)benzamide, B-3 to Induce

Chromosome Aberrations in Cultured Peripheral Human Lymphocytes (with Repeat Experiment). Notox B.V., He rtogenbosch, The Netherlands. Laboratory report number: 489613. Study re port date: 16-April-2009. Applicant Report

Number: 2009/1126682. DACO 4.5.6. 48542717 Evaluation of the Mutagenic Ability of 2-(Trifluoromethyl)benzamide, B-3 in an

In Vitro Mammalian Cell Gene Mutation Test with L5178Y Mouse Lymphoma Cells (With Independent Repeat). Notox B.V., Hertogenbosch, The Netherlands. Laboratory report num ber: 489614. Study report date: 20-April-2009. Applicant Report Nu mber: 2009/1126683. DACO 4.5.5. 48542718 Evaluation of DNA Repair Inducing Ability of 2-(trifluoromethyl)benzamide, B-3 in Male Rat Hepatocytes ( In Vivo Rat Hepatocyte DNA-Repair Assay). Notox B.V., Hertogenbosch, The Netherlands . Laboratory report number: 490347. Study report date: 23-April-2009. Applicant Report Number: 2009-1126684.

DACO 4.5.8. 48542670 Assessment of Acute Oral Toxicity with AB-13 (Analytical Grade) in the Rat (Acute Toxic Class Method). Notox B.V., Hertogenbosch, The Netherlands. Laboratory report number: 407835. Study report date: 09-August-2004. Applicant Report Number: OTSA-0197. DACO 4.2.1.PC Code: 138831 FINAL

28 48542688 Evaluation of the Mutagenic Activity of AB-13 (Impurity of OK-5101) in the Salmonella typhimurium Reverse Mutation Assay and the Escherichia coli Reverse Mutation Assay (With Inde pendent Repeat). Notox B.V., Hertogenbosch, The Netherlands. La boratory report number: 407846. Study report date: 19-August-2004. Applicant Report Number: 2004/1038340. DACO

4.5.4. 48849413 Evaluation of the Mutagenic Activity of AB13 (Impurity of OK-5101) in an In

Vitro Mammalian Cell Gene Mutation Test with L5178Y Mouse Lymphoma Cells (With Independent Repeat). Notox B.V., Hertogenbosch, The Netherlands. Laboratory report number: 408408. Study report date: 14-July-2004. DACO 4.5.5. 48542690 Evaluation of the Ability of AB-13 (Impurity of OK-5101) to Induce

Chromosome Aberrations in Cultured Peripheral Human Lymphocytes (With Repeat Experiment). Notox B.V., He rtogenbosch, The Netherlands. Laboratory report number: 407857. Study report date: 13-July-2004. Applicant Report

Number: OTSA-0193. DACO 4.5.6. 49029701 BAS 9210 I (Cyflumetofen): Repeated dose 1-year oral toxicity study in rats.

Institute of Environmental Toxicolo gy, Japan. Laboratory report number IET 11-0001; 70C0510/09X067. Study report date: 21-Nov-2012. Applicant Report

Number 2012/1343340. 49209101 BAS 9210 I (Cyflumetofen): Carcinogenic ity Study in Mice. The Institute of Environmental Toxicology, Ibaraki, Japan. Laboratory report number: IET 11-0016. Study report date: 08-August-2013. Applicant Report Number:

2013/1251154.

49226601 BAS 9210 I (Cyflumetofen): Carcinogenicity Study in Rats. The Institute of Environmental Toxicology, Ibaraki, Japan. Laboratory report number: IET 11-0002. Study report date: 07 October, 2013. Applicant Report Number:

2013/1345430.

49250601 Mode of Action for Leydig Cell Tumors in a High Dose 2-yr Rat Cancer Study with Cyflumetofen. BASF Corporati on, 26 Davis Drive, Research Triangle Park, NC 27709. Study report date: Oc tober 23, 2013. BASF Registration

Document Number 2013/7002798.

US EPA Lori L. Brunsman (2013). Cyflumetofen Qualitative Risk Assessment Based On F344 Rat Dietary Study. TXR No.: 0056837

Boorman et al., 1990 Boorman GA, Chapin RE, and Mitsumori K (1990).Testis and epididymis. In: Pathology of the Fischer Rat—Reference and Atlas , GA Boorman, SL Eustis, MR Elwell, CA Montgomery,

and WF MacKenzie (eds). Academic Press, New York, San Diego, pp. 405–418.