Document qgyNyR87vjDqMypJOoknjr5x

era - 0800 _ 373 AR126--0%6 ORIGINAL Geil FINAL REPORT Wo CBI PROTOCOL 418-011 ORAL (GAVAGE) DEVELOPMENTAL TOXICITY STUDY OF N-EtFOSE IN RATS `SPONSOR'S STUDY NUMBER: T-6316.7 FINAL REPOR' DATE: 17 DECEMBER 1998 : 2 epa-ors TRENUeIAR sa Ei3:5 owosnieon +7 do3ss7? PROTOCOL 418-011 ORAL (GAVAGE) DEVELOPMENTAL TOXICITY STUDY OF N-EtFOSE IN RATS SPONSOR'S STUDY NUMBER: T-6316.7 ABLE OF CONTENTS SUBJECT PAGE IL. SUMMARY AND CONCLUSION I=] A. Methods 1 B. Results 2 C. Conclusion 4 I. DESCRIPTION OF TEST PROCEDURES 1 A. Conduct of Study 1 Ad. Sponsor 1 A2. Testing Facilty 11 A3. Study Number 1 A4. Sponsor's Study Number Tr 1-1 AS. Purpose of the Study 1 AS. Study Design 003588 [E] 5 re SUBJECT . A7. Regulatory Compliance AB. Ownership of the Study AS. Study Monitor A.10. Alternate Study Monitor A.11. Study Director A.12. Technical Performance A13. Report Preparation A.14. Report Review A15. Date Protocol Signed A186. DatesofTechnical Performance AA7. Records Maintained B. B.1. Test Article Information Description B.2. Lot/Batch Number B.3. Date Received and Storage Conditions B.4. Special Handing Instructions B.S. Analysis of Activity C. Vehicle Information C.1. Description C.2. Lot Number | C.3. Date Received and Storage Conditions C.4. Special Handling Instructions + 003589 PAGE 1-4 2 2 1-2 1-2 2 1-2 n-2 2 3 3 1-3 3 1-3 1-3 3 4 1-4 14 1-4 14 1-4 SUBJECT . C.5. Analysis of Purity D. Test Article Preparation D.1. Sample Information E Test System E.1. Species E.2. Strain E.3. Supplier (Source) E4. Sex E5. Rationale for Test System EB. Test System Data E.7. Breeder Male Rat Data E.8. Method of Randomization E. System of identification F. Husbandry F.1. Research Facility Registration F.2. Study Room F.3. Housing F4. Lighting F.5. Sanitization F6. Feed F.7. Feed Analysis F.8. Water wo ". ! 0035390 PAGE 4 1-4 1-5 1-5 1-5 1-5 5 1-5 1-6 11-6 I-68 1-6 1-6 n7 7 7 7 7 7 7 7 1-8 SUBJECT . F.9. Water Analysis G. Methods G1. Dosage Administration G2. Rationale for Dosage Selection G.3. Route of Administration G.4. Rationale for Route of Administration G5. Frequencyof Administration G6. Length of Study G7. Method of Study Performance G.8. Gross Necropsy G9 Statistical Analyses I. RESULTS A Mortality, Clinical and Necropsy Observations Al. Mortality A2. Clinical Observations A3. Necropsy Observations B. Maternal Body Weights and Body Weight Changes C. Matemal Absolute (g/day) and Relative (g/kg/day) Feed Consumption Values D. Caesarean-Sectioning and Litter Observations E Fetal Alterations . EA. Summary of Fetal Alterations : E.2. Fetal Gross External Alterations iw 003591 PAGE Is 8 8 Io 1-9 n-9 9 9 Ie 1-10 "12 1-1 n-1 [I= 1 1 1 m2 n-2 3 n-3 n-4 SUBJECT . E.3. Fetal Soft Tissue Alterations E4. Fetal Skeletal Alterations REFERENCES APPENDIX A - REPORT FIGURE Figure 1. Maternal Body Weights APPENDIX B - REPORT TABLES Table 1. Table 2. Clinical Observations -- Summary Necropsy Observations -- Summary Table 3. Maternal Body Weights -- Summary Table 4. Table 5. Table 6. Maternal Body Weight Changes -- Summary Maternal Absolute Feed Consumption Values (g/day) Summary Maternal Relative Feed Consumption Values (g/kg/day) -- `Summary Table 7. Table 8. Caesarean-Sectioning Observations -- Summary Litter Observations (Caesarean-Delivered Fetuses--) Summary Table 9. Fetal Alterations -- Summary Table 10. Table 11. Fetal Gross External Alterations -- Summary Fetal Soft Tissue Alterations -- Summary Table 12. Table 13. Fetal Skeletal Alterations -- Summary Fetal Ossification Sites - Caesarean-Delivered Live Fetuses (Day 20 of Gestation) -- Summary . Table 14. Clinical Observations - Individual Data PAGE 1-4 n-5 nz A1 B-1 B2 B-3 B-5 B-6 B-7 B-8 B-9 B-10 B-11 B-12 B-13 B-16 B-17 v 003592 SUBJECT Table 15. Table 16. Table 17. Table 18. . Necropsy Observations - Individual Data Maternal Body Weights - Individual Data Maternal Feed Consumption Values - Individual Data Caesarean-Sectioning Observations - Individual Data PAGE B-23 B-29 B-44 B-50 Table 19. Litter Observations (Caesarean-Delivered Fetuses) -- Individual Data B-56 Table 20. Fetal Sex, Vital Status and Body Weight - Individual Data B-62 Table 21. Fetal Alterations - Individual Data B-74 APPENDIX C - PROTOCOL AND AMENDMENTS C-1t0C-33 APPENDIX D - DEVIATIONS FROM THE PROTOCOL AND THE STANDARD OPERATING PROCEDURES OF THE TESTING D-1 APPENDIX E - TEMPERATURE AND RELATIVE HUMIDITY REPORT E-1 APPENDIX F - PILOT REPORT F-1to F-87 APPENDIX G - HISTORICAL CONTROL DATA G-1t0G-16 APPENDIX H - STATEMENT OF THE STUDY DIRECTOR H-1 APPENDIX |- QUALITY ASSURANCE UNIT FINAL REPORT STATEMENT tol-5 003593 .. 418-011:PAGE 1 TITLE: ORAL (GAVAGE) DEVELOPMENTAL TOXICITY STUDY OF N-EtFOSE IN RATS ARGUS RESEARCH LABORATORIES, INC. PROTOCOL NUMBER: 418-011 SPONSOR'S STUDY NUMBER: T-6316.7 I. SUMMARY AND CONCLUSION A. Methods Twenty-five female rats Cri:CDBBR VAF/PIus (Sprague-Dawley) presumed were assigned to each of five dosage groups (Groups pregnant | through V). tNhienestaeteelnitaeddsittuidoyna(ltfhreemea,lfeivrea,ttshwreeer,etahsrseeigannedd ftioveonraetsofasfisviegdnoedsatgoeGgrroouuppss|for through V, respectively). The test article, N-EtFOSE, or vehicle, 2% Tween 80 WinatReerv)e,rwsaesOsadmmoisniisstMeeremdbrviaangeavPargoceeosnsceed dDaeiiloyntiozfeedmWaalteerrat(sR.oOn. dDaeyison6itzherdough 17 of presumed gestation (DGs 6 through 17). Dosages of 0 (Vehicle), 1, 5, 10 and 20 mg/kgiday were administered at a dosage volume of mL/kg, adjusted daily on the basis of individual body weights. The female rats were observed for viability at least twice each day of the study. The rats were also examined for clinical observations of effectsof the test article, aafbtoerrtidoonss,agpere(mDaGtsur6etdherloivuegrhie1s7)a,nadnddeoatnhcse bdeaifloyreduarnidngatphperopxoismtadtoeslyagoeneperhiooud.r a. Dpertoaviildeedd dienstchreipatpipornosporfiaatllepsreoccteidonusreosf tuhsiesdreipnotrhteacnodndinucAtPPofEtNhiDsIsXtuCdy are (PROTOCOL AND AMENDMENT) 003594 418-011:PAGE I-22 Body weights were recorded on DGs 0 and 4 and daily during the dosage and postdosage periods. Feed consumption values were recorded on DGs 0, 4, 6, 8, 10, 12, 14, 16, 18 and 20. All rats in the main study were sacrificed by carbon dioxide asphyxiation on DG 20 and a gross necropsy of the thoracic, abdominal and pelvic viscera was performed. The number of corpora lutea in each ovary was recorded. The uterusof each rat was examined for pregnancy, number and distribution of implantations, live and dead fetuses and early and ate resorptions. Each fetus. was identified, weighed and examined for sex and gross extemal alterations. Aalptperroaxtiiomnastealnydotnhee-hraelmfaionfitnhge ffeettuusseess iinn eeaacchhliltitteterrwweerree eexxaammiinneedd ffoorr sskoefltettiaslsue alterations. Rats in the satelite study were sacrificed on DG 18. Blood samples were collected and centrifuged. The liver was excised, weighed and sectioned. aFsestiugsneesdwteorteheexmaamiinnestdudgyr.ossFleyttuosetsheaenxdtepnltacpeonstsaieblweearsedpeosoclreidbepderfolrittrear.ts After completion of sample collection, serum, iver section, fetal and placental samples were shipped to the Sponsor for analysis. B. Results No deaths, abortions or premature deliveries occurred during the study. All rats survived until scheduled sacrifice on gestation day 20 (DG 20) Al clinical and necropsy observations were considered unrelated to the test article Maternal body weight gains were significantly reduced in groups administered 5 mg/kg/day and higher dosagesof the test article. The effect was minimal and transient in the 5 mg/kg/day dosage group, occurring only on DGs 8 to 10. In the 10 mg/kg/day dosage group, significant reductions in maternal body weight gains occurred on DGs 6 to 8 and 10 to 12, followed by a significant increase in weight gain on DGs 14 to 16. The 20 mg/kg/day dosage group had significant weight loss followed by significant reductions in maternal body weight gain on DG 8 to 14 and 16 to 18. These effects of the test article resulted in a tendency for reduced weight gain in the 10 mg/kg/day dosage group and significant reductions in the 20 mg/kg/day dosage group for the entire treatment period (DGs6 to 10), the entire interval after initiation of treatment (DGs 6 to 20) and the entire gestation period (DGs 0 to 20). Maternal body weights were 003595 418-011:PAGE I-3 significantly reduced in the 10 and 20 mg/kg/day dosage groups on DGs 11 through 13 and 8 through 20, respectively. Body weights and body weight gains were unaffected by the 1 mg/kg/day dosage of the test article. The absolute feed consumption value was significantly reduced in the 10 mg/kg/day dosage group on DGs 6 to 8 and absolute and relative feed consumption values were significantly reduced in the 20 mg/kg/day dosage group for the entire dosage period and at all intervals within this period. The absolute feed consumption value continued to be significantly reduced and the relative feed consumption value tended to be reduced in the 20 mg/kglday dosage group during the postdosage interval. These effects of the 20 mg/kg/day dosage of the test article resulted in significantly reduced absolute and relative feed consumption values on DG 6 to 20 and DGs 0 to 20. Absolute and relative feed consumption values were unaffected by dosages of the test article as high as 5 mg/kg/day. Fetal body weights (total, male and/or female) were significantly reduced in the. 10 and 20 mg/kg/day dosage groups, as compared to the control group values. Dosages of N-EtFOSE as high as 20 mg/kg/day did not affect any other Caesarean-sectioning or litter parameters. The litter averages for corpora lutea, implantations, ter sizes, live fetuses, early resorptions, percent resorbed conceptuses and percent male fetuses, as well as the numbers of dams with any resorptions or with viable fetuses were comparable in the five dosage groups and did not significantly differ. No dams had litters with all conceptuses resorbed, and there were no dead fetuses or late resorptions. All placentae appeared normal. All of these values were within the ranges observed historically at the Testing Facility. fReetvaelrbsoidblyewdeeilgahytss iinn ftehteal1o0ssainfidca2t0iomnga/sksgo/cdiaayteddowsiatghethgerosuipgnsi,fiwcearntelyevrieddeuncteads. significant 10 and 20 reductions mg/kg/day in the litter averages for ossified dosage groups and a significant caudal vertebrae in increase in the fetal the incidence of wavy ribs in the 20 mg/kg/day dosage group. Aalnldotvhaerrifateitoanls)grwoesrseexctoenrsniadle,rseodftutnirsesluaeteadndtostkheleetteasltaalrtteircaletiboensca(umsael:for1m)atthieons incidences were not dosage-dependent; and/or 2) the incidences were within ranges observed historically at the Testing Facility. 003596 418-011:PAGE I-4 Cc. Conclusion On the basis of these data, the maternal no-observable-effect-level (NOEL) of N-EtFOSE is 5 mg/kg/day (the 10 and 20 mg/kg/day dosages caused biologically important and statistically significant reductions in body weight gains or weight losses, and the 20 mg/kg/day dosage also persistently reduced the absolute and relative feed consumption values). The developmental NOEL is also 5 mg/kg/day (the 10 and 20 mg/kg/day dosages significantly reduced fetal body weights and caused minimal, but statistically significant reversible delays in ossificationof the caudal vertebrae; the 20 mg/kg/day dosage also significantly increased the incidence of wavy ribs, an additional reversible delay in ossification associated with the reduced fetal body weights). WO bJd z fosppeers Mildred S. Christian, Ph.D., Fellow, ATS ~~ Date Executive Director of Research Lobb Alan M. Hoberman, Ph.D., DABT. Director of Research rms Date nd G. York, Ph. Associate Director ofR and Study Director 17: dy BT Date rch 003597 418-011:PAGE Il-1 [I CRI OF TEST PROCEDURE! A. Conduct of Study: A. Sponsor: 3M CorporateToxicology, 3M Center, Building 220-2E-02, St. Paul, Minnesota 55144-1000 A.2. TestingFacility: Argus Research Laboratories, Inc., 905 Sheehy Drive, Building A, Horsham, Pennsylvania 19044-1297 A3. StudyNumber: 418-011 A4. Sponsor's Study Number: T6316.7 AS. Purpose of the Study: The purposeofthis study was to detect adverse effects of N-EtFOSE on Cr:CDBR VAF/Plus presumed pregnant female rats and development of the embryo and fetus consequent to exposure of the dam from implantation to closure of the hard palate. This study evaluated ICH Harmonised Tripartite Guideline stages C and D of the reproductive process. AS. Study Design: The requirements of the International Conference on Harmonisation (ICH) Harmonised Tripartite Guideline were used as the basis of study design. A7. Regulatory Compliance: The study was conducted in compliance with Good Laboratory Practice (GLP) regulations of the U.S. Food and Drug Administration (FDA)? the Japanese Ministry of Health and Welfare (MHW) and the European Economic Community (EEC). There were no significant deviations from the GLP regulations that affected the quality or integrity of th study. Quality Assurance Unit findings derived from the inspections during the conduct of this study are documented 003598 . 418-011:PAGE Il2 and have been provided to the Study Director and the Testing Facility Management. A.8. Ownerof sthehStiudpy: The Sponsor owns the study. All raw data, analyses, reports and preserved tissues are the property of the Sponsor. A.9. StudyMonitor: Marvin T. Case, D.V.M., Ph.D. A10. AlteStrudnyMaonittoer: Andrew M. Seacat, Ph.D. A.11. StudyDirector: Raymond G. York, Ph.D., DABT (Associate Director of Research) A12. Technical Performance: JKroihsnteFn. lBaanmdeotlta,SBh.eSr.er(,DiBr.eSc.to(rRoefseLaarbcorhaAtsosroyciOapteer/aFteitoanls)Evaluation) Sharon Adamski (Laboratory Technician) A.13. ReportPreparation: Raymond G. York, Ph.D., DABT Jo Ann Susan Frazee, M.S. K. Bradshaw, (Study Coordinator) B.S. (Data Management Specialist) Karen G. Parker, AA. (Administrative Assistant) AA4. Report Review: Alan M. Hoberman, Ph.D, DABT (Director of Research) Mildred S. Christian, Ph.D., Fellow, ATS (Executive Director of Research) A15. Date Protocol Signed: 29 July 1998 003599 . 418-011:PAGE I1-3 A16. Datesof Technical Performance: Rat Arrival Date Cohabitation Period Day 0 of Presumed Gestation (DG 0) Dosage Period (DGs 6 through 17) Toxicokinetic Sample Collection and Caesarean-Sectioning Period Ca(eDsGar1e8a)n--SSaetcetliliotneiSngtuPdeyriod Main Study (DG 20) 11 AUG 98 18 AUG 98 PM - 23 AUG 98 AM 19 AUG 98 - 23 AUG 98 25 AUG 98 - 09 SEP 98 10 SEP 98 08 SEP 98 - 12 SEP 98 AA7. Records Maintained: The are original retained report, in the raw data and reserve samples of the test archives of Argus Research Laboratories, article and vehicle Inc. Any preserved imasisluiensg aorfethreetdarianfetdfiinnalthreepaorrtc,hiavfetserowfhtihcehTteismteintgheFaSciplointysoforrwoilnledyeecairdeafttheerirthfienal disposition. Prepared formulations were discarded at the Unused bulk test article will remainat the Testing Facility Testing Facilty. until its disposition is decided by the Sponsor. B. TestArticle Information: B.A. Description: N-EtFOSE - a waxy solid B.2. LotBatch Number: FM-3929 [30035, 30037, 30039 (Expiration date: May 2000)] B3. Date Received and Storage Conditions: The test article was received on 20 May 1998, and stored at room temperature. Prepared formulations were stored refrigerated. B.4. Special Handling Instructions: Standard safety precautions respirator, safety goggles or (use of protective clothing, gloves, dust-mist safety.glasses and a face-shield) were taken when handling the bulk test article and prepared formulations. 003600 418-011:PAGE Il-4 BS. Anal Purity: Information regarding the identity, composition, strength, and purity of the test article is on file with the Sponsor. C. Vehicle Information: C1. Description: 2% Tween 80 in Reverse Osmosis Membrane Processed Deionized Water (R.O. Deionized Water). C.2. LotNumber: MO3H05 C3. Date Received and Stora tions: The New vehicle was Jersey, and received on 8 July 1998 from stored at room temperature. J.T. Baker, Phillipsburg, R.O. Deionized Water is available from a continuous source at the Testing Facility and is maintained at room temperature. ca s ndling Instructions: Standard safety precautions respirator, safety goggles or (use of protective clothing, gloves, dust-mist safety glasses anda face-shield) were taken when handling the vehicle. C.5. AnaloyfPsuriitsy: Neither the Sponsor nor the Study Director was aware of any potential contaminants likely to be present in the vehicle that would interfere with the results of this study. D. TestArticle Preparation: `aSnudsp4emnsgi/omnLs. ofTNh-eEttesFtOaSrEticwleerweapsrecpoanrsieddedraeidly1a0t0c%onpcuenrterfaotritohnes pofur0p,o0s.e2,o1f,2 dosage calculations. 003601 418-011:PAGE II-5 D1. SampleInformation: [sammorie|comuoens|ste|names|coStsortagoe |snepesto|shiTagtees| [CoGnceentraation | [mIle|S28 AUGR88 [ReeAsUG]SE EGoAk Test I NR RlFoo0n00 cTsivngiFacidty|B0a10CaT8l6 ReRseevreve Temperature| Archives Water Temperature| Archives a. sDuapmlipclatoefseaamcphlseestwwearsetsahkiepnpferdotmo tthheeSfpirostnsaonrdfloarstanparlyespisa.raTthioeonretmhaeidnainygpsraepmaprleeds. wOernee b. rFiertsatindeadyaotfthprepTaersattiinogn.Facilty as backup samples. c Lastday of reparation Homogeneity and stability of prepared formulations are on file with the Sponsor. D.2. Analytical Results: Concentration samples (2 mL) were taken on the first and last days of preparation for analyses by 3M Environmental Technology and Safety Services. The resultsof these analyses were not available at the time of this report. E. TestSystem: . EA. Species: Rat E2. Strain: Crl:CDBR VAF/Plus (Sprague-Dawley) E.3. Supplier(Source): Charles River Laboratories, Inc., Raleigh, North Carolina Ed. Sex: Female (Note: Male rats were used only considered part of the Test System) for the purposes: of breeding and are not 003602 418-011:PAGE 16 ES. Rationale for Test System: TShyestCermlb:eCcDauBsRe:VA1F)/iPtIisusone(Smparmamgaulei-Daanwlspeeyc)ireast awcacsepsteeldecatendd awsidtehleyTuessetd ttohxriocuitgyhlotuetraitnogdeunsitcriytyf)o;r2n)ontchliisnsitcraalinsthuadsiebsoefenddeveemloonpsmternattaeld ttooxibceitsye(nesmibtriyvoe-tfoetal dFeacvielliotpym"e;natnadl t4o)xtinhse;t3e)sthiasrttiocrliecailsdbaiotlaogaincdalleyxpaecrtiiveencinetehxiisstspaetctiheesTaenstdisntgrain. E.6. TestSystemData: Number of Rats Approximate Date of Birth Approximate Age at Arrival Weight (g) on the Day after Arrival Weight (g) at Study Assignment 190 08 AUG 98 64 days 195-234 210-251 E7. Breeder Male Rat Data: Shipment 1 `Shipment 2 Number of Rats. Approximate Date of Birth Approximate Age at Arrival Weight (g) on the Day after Arrival Weight (g) at Study Assignment 110 120 13 JAN 98 26 JAN 98 87 days 75 days 300-356 498-784 300-356 ES. Method of Randomization: gUepnoenraatrreidvalr,anradtosmweunriets.assFiegmnaeldetoraitnsdwiveirdeuaalshsoiugsniendgtoonotnheeobfafsiivse odfocsoamgpeuter- groups (Groups | through V), 25 rats per dosage group, using a computer- generated (weight-ordered) randomization procedure based on body weights recorded on DG 0. dosage groups for tNheinseatteelelinteadsdtiutdioyn(atlhrfeeem,afliever,atthsrewee,retharseseiagnndedfitvoe one rats of five raasnsdiogmniezdattoioGnrobuapssed| tohnrobuogdhyVw,eirgeshptesctrievceolryd)eudsionngDaGco0.mputer-generated ES. System of Identification: Each rat was individually identified with a Monel self-piercing ear tag (Gey Band and Tag Co, unique permanent Inc., No. number. MSPT 20101) inscribed Cage tags were marked with with the the rat's designated study number and permanent rat number. . 003603 418-011:PAGE Il7 F. Husbandry: FA. Research Facili istration: USDA Registration No. 23-R-099 under the Animal Welfare Act, 7 U.S.C. 2131 ot seq. F.2. StudyRooms: The study rooms were maintained under conditions of positive airflow relative to a10h0al%lwfaryesahnadiritnhdaetpheanddebnetleynspuapspslieeddtwhirtohuaghm9i9n.i9m7u%m HofEtPeAn cfihlatenrsge(sAirpoerClheouarnof room). Room temperature and humidity were monitored constantly throughout the study. Room temperature relative humidity was targeted was targeted at at 30% to 70%. 64F See to 79F (18C APPENDIX E to 26C); (TEMPERATURE AND RELATIVE HUMIDITY REPORT). F3. Housing: Rcoahtasbiwteartieoni,ndeivaicdhuapllayirhoofumsaeldeeaxncedpftedmuarliengratthsewcaoshahboituasteidoninpetrhieodm. alDeurriatn'gs cage. All cage sizes and housing conditions were in compliance with the Guide for the Care and Use of Laboratory Animals . F.4. Lighting: Alinght:aut1o2m-ahtoiucraslldya-rcko,ntwriotlhleedacflhudoraerskcepnetriloidghtbecgyicnlneiwnagsatma1i9n0t0aihnoeudrsatES1T2-.hours F5. Sanitization: Cage pan liners were changed approximately three times each week. Cages `were changed approximately every other week. F6. Feed: : Rats were given ad libitum access to Certified Rodent Diet #5002 (PMI Nutrition International, St. Louis, Missouri) in individual feeders. F.7. Feed Analysis: Alenvaellysseexscweeedrienrgoutthienemlayxpiemrfuomrmceodncbeynttrhaetifoenedfosrucpeprltiiefri.ed Nfoeecdoonrtadmeivinaatnitosnsatfrom 003604 418-011:PAGE II-8 expected nutritional requirements were detected by these analyses. Copies of the results of the feed analyses are available in the raw data. Neither the Study Director nor the Sponsorwas aware of any agent present in the feed that was known to interfere with the results of this study. F.8. Water: Lmoecmablrwaanteer(tRh.aOt. hwaadtebre)ewnapsraovcaeislsaebdlebtyo ptahessraatgseatdhrloiubigthumafrreovmerisndeiovisdmuoasliwsater bottles attached to the cages and/or from an automatic watering system. Chlorine was added to the processed water as a bacteriostat. F.9. WaterAnalysis: The processed water is analyzed twice annually for possible chemical fcoorntpoasmsiinbalteiobnac(tLerainaclasctoenrtaLmaibnoartaitoornie(sAn,aLlyatniccaaslteLra,boPreatnonrsiyelsv,anInica.), and monthly Chalfont, Pennsylvania). Copies of the results of the water analyses are available in the raw data. tNheeitwhaetretrhtehSattuwdaysDkirneocwtonrtnooirnttherefeSrpeownistohrthweasresauwlatrseooffthaisnystaudgye.nt present in G. Methods: GA. DosageAdministration: cone | NoToonrt|| moasnggee | Gannll| vSeess S| oi | | EE [oe cee] [ovToveJ 0Ts vaeorvaeas [vasrs.vans |} [Tv[wee[7Toa |5 assvss[vaseamo| Co eoTe +Toms [om [[oovvI[moese[ [ 0T 1 z11 55 [|avwooorvvaamsm|[vavsosswerivaasommsr||] R2 RRRatwsasnSoareeesaeeTOgDropu rboorbrosrctoescson.653938 Eo The test article was considered 100% pure for the purpose of dosage calculations. 003605 418-011:PAGE Il-9 G.2. Rationalefor Dosa ction: Dosages were selected on Laboratories, Inc., Protocol the basis of a dosage-range study 418-011P) that tested 0, 1, 5, 10, (Argus 25 and Research a3n5dmgh/ikggh/edrady.osaIgnetsh,atanstdudfye,edbocdoynswuemipgthitognaivnalwuaess wdeercereraesdeudceatd 1a0t amlgl /dkogs/adgaeys tested. G3. Route of Administration: Oral (gavage) G4. Rationale for Route of Administration: The oral (gavage) route dietary route, the exact was selected for use because: 1) in comparison with dosage can be accurately administered; and 2) itis a the proposed route of human exposure. G5. Frequency of Administration: Appropriate dosages of the once daily to female rats on test article were DGs 6 through administered 17. Dosages orally (via gavage) of 0 (Vehicle), 1, 5, 10 a5nmdLi2k0g,mga/dkjgu/sdteadyodfatilhyeotnestthearbtiacslieswoefrteheadimnidniviisdtuearlebdoadtyawdeoigshatgserveoclourmdeedof before dosage. The rats were dosed at approximately the same time each day. G.6. LenofgSttudh y: Approximately 4 weeks G.7. Method of Study Performance: Awfittehr1a9c0clbirmaeteidoenr, m1a9l0ehreaatlst(hyonveirmgianlefermaatlpeerraftesmwaelreerpatlaincetdheinmtaolceohraatb'istactaigoen) Female rats with spermatozoa andlor a copulatory plug in situ observed in a smear of were considered to be the vaginal contents at DG 0 and retumed to individual housing. Tahnde ffoermgaelneerraatlsawpepreearobasnecrevweedefkolryvidaubriilintgy aactclleiamsatttiwonicaenedacohn dDaGy 0o.f tThheestruatdsy were also examined for clinical observations of effects of the test article, 003606 418-011:PAGE 11-10 aafbtoerrtidoonss,agpere(mDaGtsur6e tdherloivuegrhie1s7%a)n, danddeaotnhcsebedafiolryedaunridngaptphreoxpiomsattdeolsyaogneepehroiuord. dBaoidlyy dwueriignhgttshweedroesraegceoardneddpwoesetkdloysdaugreinpgeraicocdlsim(aDtiGosn,6 otnhrDoGugsh 020a).nd F4eaendd consumption values were recorded on DGs 0, 4, 6, 8, 10, 12, 14, 16, 18 and 20. G8. GrossNecropsy": Rats Assigned to the Main Study: Al rats were sacrificed by carbon dioxide asphyxiation on DG 20, Caesareansweacstipoenrefdoramnedd.a gUrteorsis onfeacprpoaprseynotflytnhoentphroreagcniac,ntabradtosmwienarle astnadinpeedlvwiictvhis1c0e%ra agrmomsosnliesuimonssulwfeirdeetporceosnefrivremdtihnenaeubtsreanlcbeufoffeirmepdl1an0t%atfioornmsailtiensf.or Tpiossssiubelsewfiutthure evaluation; all other maternal tissues were discarded. "wTahse nexucmibseerdoafncdorepxoarmainluetdeafoirn peraecghnaonvcayr,y nwuamsbreercoarnddedd.istTrhibeutuitoenroufs of each rat rimepsloarnpttaitoinownsa,sldiveefiannedddaesaodnfeetinuswehsicahndoregaralnyogaenndeslaitse wreassornpottiognrso.ssAlyneevairdleynt wAalsatgerroesssolrypteviiodnewnta.sAdefliivneefdetaussownaesidnewfhiincehd athseaotcecrumrrfeentcues tohfaotrrgeasnpoognedneedsitso sdteiamudlif.etuNsoens)r.espDoenaddinfgettuesrems faentdusleastearreescoornpstiidoenreadretodibfefedreenatdiat(etdhebrye twheere no degree of autolysis present; marked to extreme autolysis indicated that the fetus was a late resorption. Each fetus was removed from the uterus, placed in an individual container and iadnedntfiifxiaetdivwei.thEaactahgfnetoutsinwgatshesusbtsuedqyuneunmtbleyrw,eliigttherednuamnbdere,xautmeirniende dfiosrtrsiebxutainond gross external alterations. Live fetuses were sacrificed by an intraperitoneal a See APPENDIX D (DEVIATIONS FROM THE PROTOCOL AND STANDARD OPERATING PROCEDURES OF THE TESTING FACILITY), item 1. b. A table of random units was used to select one control group rat from cwohnitcrholaltlitsissuseusefsorexaanmyipneodsastenehcirsotoppsaythwoelroegirceatlaeinveadl,uaitnioorndsoerftgorporsosvide lesions. 0a3607 418-011:PAGE Il-11 iPnhjoetctoigornaopfhsBeouftghraonsasseixate-rDnaSlpfeectiaallal(tSecrhaetrioinnsg-aPrleoauvgahilAanbilmeailn Htehaeltrha).w data. Approximately one-half of the fetuses in each litter were examined for soft tissue alterations using an adaptationof Wilson's sectioning technique". The fetuses were initially fixed in Bouin's solution; sections were stored in alcohol. The rreedmaSiTMni,ngfifxeetduisnesalicnoheoalchalnidtteerxwaemrieneedvifsocresrkaetleedt,alclaelaterreadt,iosntsa.inSekdewlietthalalizarin preparations were retained in glycerin with thymol added as a preservative. igned to the Satelli ; fOonllDowGing18s,armaptlseasssciogllneecdtetdo.thBelotooxdicsoakmipnelteisc e(vaaplpuraotxiiomnatweelrye4smacLrifpiecredraat)nwdetrhee Tcohlelercetseudltfirnogmstehreuimnf(earpioprrvoxeinmaatcealvya2inmtLo)swearusmimsmepeadriaattoerlytufbreoszeannodncednrtyriifcuegaedn.d maintained frozen (-70C) until shipment to the Sponsor for analysis. The liver was excised, weighed, and a sample section (lateral lobe) was frozen and retained at -70C until shipment to the Sponsor for analysis. Rats were Caesarean-sectioned and fetuses were examined grossly to the extent possible as described above for rats assigned to the main study. Fetuses and placentae were pooled per litter and retained frozen (-70C) until shipment stoectthieonSp(loantseroarl lfoorbea)n,alfyestials.anAdftpelracceonmtpalletsiaomnplofessawmeprleeschoilplpecetdio(nf,roszeernumo,n ldirvyer ice) to the Sponsor for analysis. 0Q3608 418-011:PAGE 11-12 G9. Statistical A : The following schematic represents the statistical analyses of data: Type of Test I. Parametric I. Nonparametric A. Bartlett's Test A. Kruskal-Wallis Test (75% ties) Significant at p<0.05 Nonparametric Not Significant | AnalysisofVariance Significant at p0.05 | Dunn's Test Not Significant Significant at p<0.05 Not Significant B. Fisher's Exact Test (>75% ties) Dunnett Test lll. TPesrt foorportiDoatna Variance Test for Homogeneity of the Binomial Distribution a. Statistically significant probabilities are reported as either p<0.05 or p<0.01 b. Proportion data are not includedin this category. c. Used only to analyze data with homogeneity of variance d. Testfor homogeneity of variance. : 603609 418-011:PAGE 1113 Clinical observation and other proportion data were analyzed using the Variance Test for Homogeneity of the Binomial Distribution"?. cCoonntsiunmupotuisondavtaalu(ee.sg.a,nmdaltietemraalvebroadgyewseifogrhtpse,rcbeondtymwaelieghftetcuhseasn,gepse,rcfeenetd resorbed conceptuses, fetal body weights, ossification site data) were analyzed using fetal anomaly Bartlett's Test data and fetal of Homogeneity of VTeasrtiwaanscenostasnigdnitfhiecaAnntal(py>s0i.s05o)f].VaIrfitahencAnea,lywshiesnofapVparroiparnicaetewa[is.e.,siBganritfliectatn'ts (inpd<i0v.i0d5u)a,l Dgurnonuepst.t'sIfTtehsetA"nawlayssiussoefdVatoriiadenncteifwyatshensottataipsptircoaplrsiiagtneif[ii.ce.a,ncBeartolfettth'es Ttehsatn woraseqsuiaglnitfoic7a5nt%(pt<i0es.0w5e)]r,etphreesKernuts.kalIn-WcaalsliessTwehsetre" twhaesKruussekda,l-wWhaelnlisleTsesst wwaass sutsaetidsttiocaildleynstiigfnyitfhiceansttat(ips<t0i.c0al5)s,igDnuinfnic'asncMeeotfhothdeoifndMiuvlitdiupalle gCroomuppsa.riIsf othnesr"e were greater than 75% ties, Fisher's Exact Test" was used to analyze the data. tChoeunptrodcaetdauorbetsadiensecdraitbeCdaeasbaorveeanf-osretchteioKnruisnkgalof-WtahlelidsaTmesswte"re evaluated using aDnadmt1w2o8e6a8rl(y5rmesgo/rkpgt/idoanys adnodsadgaemgr1o2u8p8)9h(a1d0amgli/ttkegr/cdoansyidstoisnaggoeffgirvoeupli)vehafedtuases Jsikteerwctohnesidsitsitnrgibouftitohnroefe dliavetfae,tussetast.istBiceaclaaunsaelyssuecshwoecrceurmraednecewsicthananadbnwoirtmhaolulty VthaeluveasluaensdfCoraetshaerseeanra-tsseactnidolnitdearts.a fMoarttehrensael dbaodmyswaenidghltist,terfseweedrceonesxculmupdteidon from summarization and statistical analyses; all values are presented on the individual tables. 003610 418-011:PAGE IlI-1 mn. RESULTS - A. Mortality, Clinical and Necropsy Observations (Summaries - Tables 1 and 2; Individual Data - Tables 14 and 15) AA. Mortality No deaths, abortions or premature deliveries occurred during the study. All rats survived until scheduled sacrifice on gestation day 20 (DG 20). A.2. Clinical Observations All clinical observations were considered unrelated to the test article because: 1) the incidences were not dosage-dependent; 2) the observations occurred in only one rat; and/or 3) the observations are common events in the laboratory environment. Clinical observations included localized alopecia on the underside, limbs and/or neck, ungroomed coat, cold to touch and fused second and third digits on the right forepaw. A.3. Necropsy Observations The only necropsy finding was a tan area (0.6 cm x 0.8 cm) on the median lobe of the liver in one 20 mg/kg/day dosage group dam (12913). This observation was considered unrelated to the test article because it occurred in only one rat. B. Maternal Body Weights and Body Weight Changes (Figure 1; `Summaries - Tables 3 and 4; Individual Data - Table 16) Maternal body weight gains were significantly reduced (p<0.05 or p<0.01) in groups administered 5 mg/kg/day and higher dosages of the test article. The effect was minimal and transient in the 5 mg/kg/day dosage group, occurring only on DGs 8 to 10. In the 10 mg/kg/day dosage group, significant reductions (p<0.05) in maternal body weight gains occurred on DGs 6 to 8 and 10 to 12, followed by a significant increase (p<0.05) in weight gain on DGs 14 to 16. The 20 mg/kg/day dosage group had significant weight loss (p<0.01) on DGs 6 to 8 followed by significant reductions (p<0.05 or p<0.01) in maternal body weight gain on DGs 8 to 14 and 16 to 18. These effects of the test article resulted in a tendency for reduced weight gain inthe 10 mg/kg/day dosage group and significant reductions (p<0.01) in the 20 mg/kg/day dosage group for the entire. treatment period (calculated as DGs 6 to 18), the entire interval after initiation of treatment (DGs 6 to 20) and the entire gestation period (DGs 0 to 20). Maternal 903611 418-011:PAGE IIl-2 body weights were significantly reduced (p<0.05 or p<0.01) in the 10 and 20 mg/kg/day dosage groups on DGs 11 through 14 and 8 through 20, respectively. Body weights and body weight gains were unaffected by the 1 mg/kg/day dosage of the test article. C. Maternal Absolute (g/day) and Relative (g/kg/day) Feed Consumption Values (Summaries - Tables 5 and 6; Individual Data - Table 17) The absolute (g/kg/day) feed consumption value was significantly reduced (<0.05) in the 10 mg/kg/day dosage group on DGs 6 to 8 and absolute (g/day) and relative (g/kg/day) feed consumption values were significantly reduced (p<0.01) in the 20 mg/kg/day dosage group for the entire dosage period (calculated as DGs6 to 18) and at all intervals within this period. The absolute feed consumption value continued to be significantly reduced (ps0.01) and the relative feed consumption value tended to be reduced in the 20 mg/kg/day dosage group during the postdosage interval (DGs 18 to 20). These effects of the 20 mg/kg/day dosage of the test article resulted in significantly reduced (p<0.01) absolute and relative feed consumption values on DGs 6 to 20 (the entire interval after the first dosage was administered) and DGs 0 to 20 (the entire gestation period). Absolute and relative feed consumption values were unaffected by dosages of the test article as high as 5 mg/kg/day. The significant reduction (p<0.05) in the relative feed consumption value in the 1 mg/kg/day dosage group on DGs 6 to 8 was considered unrelated to the test article because the value was not dosagedependent. D. Caesarean-Sectioning and Litter Observations (Summaries -Tables 7 and 8; Individual Data - Tables 18 through 20) Pregnancy occurred in 24 (96%), 23 (92%), 24 (96%), 25 (100%) and 24 (96%) ofthe rats in the 0 (Vehicle), 1, 5, 10 and 20 mg/kg/day dosage groups, respectively. One 5 mg/kg/day dosage group liter consisted of five live fetuses and two early resorptions, and one 10 mg/kg/day dosage group litter consisted of three live fetuses. Because such occurrences can abnormally skew the distributionof the data,values.for these dams and litters were excluded from data summarization and statistical analyses. As a result, Caesarean-sectioning observations were based on 24, 23, 23, 24 and 24 pregnant dams. 003612 418.011:PAGE IIl3 Fetal body or p<0.01) we in igh the t1s0(taontadl,2m0amlge/akngd//doaryfdeomaslaeg)e were sig groups, naisficcaonmtlpyarreeddutcoedt (p he <0.05 control dosage group values. groups were The reduced a reflection of fetal body body weights in the weight reduction of 10 the and 20mg/kg/day dams at these higher dosage levels. Dosages of N-EtFOSE as high as 20 mg/kg/day did not affect any other Caesarean-sectioning or litter parameters. The liter averages for corpora lutea, implantation, lter sizes, live fetuses, early resorptions, pe of rcent dams resorbed with any conceptuses and percent resorptions or with viable male fetuses, as well fetuses were compara as the numbers ble in the five dosage groups and did not significantly differ. No dams had litters with all conceptuses resorbed, and there were no dead fetuses or late resorptions. All philsatcoreinctaalleyaaptptehaerTeedstnionrmgalFa.cilAtlyl".of these values were within the ranges observed E. Fetal Alterations (Summaries - Tables 9 through 13; Individual Data able 21 Fetal alterations were defined as: 1) malformations (irreversible changes that `foicncduirngast ilnotwhiisncsipedceinecse/sstirnaitnhi,s asnpdecrieevserasnidblsetdraeilna);ysaonrd a2c)cevalreiraattiioonnss (incommon development). Litter averages were calculated for specific fetal ossification sites as part of the evaluation of the degree of fetal ossification. Fetal evaluations were based on 342, 349, 347, 354 and 326 DG 20 Caesarean- delivered live fetuses in 24, 23, 24, 25 and 24 litters in the 0 (Vehicle), 1, 5, 10 agrnodss20exmtge/rknagl/daaltyerdaotsioangse. gOrfoutphse,serersepsepcteicvteilvye. feEtaucsehsfe1t6u6s, w1a67s,e1x6a9m,i1n7e0d afnord 158 were examined for soft fissue alterations and 176, 182, 178, 184 and 168 were examined for skeletal alterations and fetal ossification site averages. EA. Summary of Fetal Alterations (Summary - Table 9; Individual Data [able 21 In the five respective dosage groups, litters with fetuses with alterations present numbered 6 (25.0%), 8 (34.8%), 4 (16.7%), 7 (28.0%) and 8 (33.3%). The numbers of fetuses with any alteration observed were 15 (4.4%), 10 (2.9%), 6alt(e1r.a7t%i)o,n 8we(r2e.24%.)4%a,nd2.182%,(31..7%6)%,, a2n.d1%thaenpder4c.e0n%t,aignetshoefsefestausmees wrietshpeacntiyve dosage groups. : a See APPENDIX G (HISTORICAL GONTROL.DATA) 003613 418-011:PAGE ll-4 Rreevdeurcseidbl(ep<d0e.l0a5ysorinpf<e0t.a0l1o)ssfeitfailcbaotdioynwe"iagshstosciinattheed w1i0thantdhe2s0igmngif/ikcga/ntdlayy davoesraaggeesgrfoourposs,siwfeirede ceavuiddaenltvaesrtseibgrnaifeicinantthere1d0ucatniodns20(pm<g0/.k0g5/)dianytdheosliatgteer groups and a significant increase (p<0.05) in the fetal incidence of wavy ribs in the 20 mg/kg/day dosage group. All other fetal gross external, soft tissue and skeletal alterations (malformations and variations) were considered unrelated to the test article because: 1) the. incidences were not dosage-dependent; and/or 2) the incidences were within ranges observed historically at the Testing Facilty. E2. Fetal Gross External Alterations (Summary - Table 10; Individual Data- Table 21) E.2.a. Malformations One 10 mg/kg/day dosage group fetus (12885-15) had a short trunk and absent tail. Subsequent skeletal examination of this fetus revealed that only four cervical vertebrae were present and that there were no thoracic, lumbar, sacral or caudal vertebrae, or ribs. This fetus also had a variation in pelvic ossification (the pubes were not ossified). E.2.b. Variations No gross external variations occurred in the fetuses in this study. E3. Fetal Soft Tissue Alterations (Summary - Table 11; Individual Data - Table 21) E.3.a. Malformations No fetal malformations were identified at visceral examination. E.3.b. Variations E3b.1. Vessels Three control group fetuses (12801-10; 12810-10; 1221-6), one 1 mg/kg/day dosage group fetus (12834-14) and tio 5 mg/kg/day dosage group littermates 003614 418-011:PAGE Ill-5 T(1h2e8s6e0-f5e,tu-s9)eshahaddthneouemxbtielrincaall fairntdeirnygdseasncdenndoinogthteortshoefletfitsosfuethaeltbelraatdidoenrs.. Tinwnoom1inmagtlekgalrtdearyy.doNsoagadedigtrioounpalfaelttuesreasti(o1n2s8o2c6c-u1r2r;ed12i8n3t7h-e9s)ehfaedtusaens.absent E3b.2. Lungs O1 nmeglckognltrdoalygdroosuapgfeetgurso(u1p2f8e2t2u-s4)(1h2a8d50a-n16a)bsheandtaanpiacbaslelnutngdilaopbeh,raagnmdatoincelung lobe. No additional alterations occurred in these fetuses. E33. Kidneys Two control group fetus (g1r2o8u7p7-fe1t4u)shesad(1s2li8g0h2t-o1r1;mo1d2e8r0a3t-e2)dialnadtioononfe 10 mg/kg/day dosage the pelvisof ane or both kidneys, a reversible developmental delay. No additional alterations occurred in these fetuses. Ed. Fetal SkeletalAlterations (Summaries -Tables12 and 13; Individual Data -Table 21] E.d.a. Malformations cEexrtveircnaalllvyermtaelbfroaremaendd1n0omtgh/okrgac/idca,ylduombsaarg,esgarcoraulp ofretcuasud1a2l88v5er-t1e5brhaaedoornlriybsf,ouars well as a variation in pelvic ossification (not ossified pubes), as previously described. E.4.b. Variations E.4b.1. Skull A(1l2a8r9g3e13n)as.al-Nfroonatdadlitsiuotnuarlealotcecruartrieondsion cocnuerr1e0d mg/kg/day dosage in this fetus. group fetus E4b.2. Ribs A cervical rib at rat ), occurred the 7th in 0, 3, cervical 1, 3 and v4erfteetbursae,safcroomm0m,o2n, variation 1, 3 and in this strain of 4 ltters in the f0e(tVuesheisclhea),d 1n,o5,ot1h0eraenxdte2r0namlgo/rkgsk/edlaeytadloaslategreatgiroonsu.ps, respectively. These 003615 418-011:PAGE Iil-6 Wdoasvaygeribgsr,oauprefveetrussib(l1e28d4e8l-a1y)inanosdsisfeivceanti(opns",0.o0c5c)u2r0remdg/inkgo/ndea1y mggr/okugp/fdeatyuses f(e1t2u9s04(-112;80192-870)9-a5l,so-7h,a-d10i,nc-o1m2p;l1e2t9e1l9y-o3s,si-f9i)e.d Orinbse. 2T0hmeg/sikggn/ifdiacyandtoisnacgreeagsreoiunp tchoensfietdaelreidncaidternecaetmoefnwt.arveylartiebds dinevtehelo2p0memngt/aklg/ddelaayy,doassasogceiagtreodupwiwtahsthe significantly reduced (p<0.05 or p<0.071) fetal body weights in this dosage group. E4b3. Sternum oDcecluaryreeddsitne7m,a2lo,ss1ifi,ca1t*i*onan(din1co"mpfleettuesleysofsrsoimfi3e,d 2o,r 1n,ot1 oasnsdifi1edlit1tsetrssitnetmheebra) 0 (Vehicle), 1, 5, 10 fetuses, one control and 20 mg/kg/day dosage group fetus (12802-1) had groups, respectively. incompletely ossified Of these pubes in addition to an unossified 1st sternal centrum. E4b.4. Pelvis The ischia and/or 0 fetuses from 1, pubes 0, 1, 2 were and 0 incompletely litters in the or not ossified 0 (Vehicle), 1, in 5, 3,0, 2, 10 and 2 and 2{012m8g0/2-k1g)/daanyddoonseag1e0 gmrgo/ukpgs/,dareyspdeoctsiavgeley.groOunpefefteutsus(1in28t8h5e-c1o5n)trhoaldgraodduiptional skeletal alterations, as previously described. E.4.b.5. Fetal Ossification Site Averages The litter reduced averages for ossified caudal vertebrae per fetus were (00.05) in the 10 and 20 mg/kg/day dosage groups. significantly These delays in wciatuhdatlhevesritgenbifriaclanotslsyifriecdatuicoendwe(pr<e0.c0o5nsoirdpe<r0e.d0e1f)fefcettaslobfotdhye wteesitgharttsicilnetahsessoeciated dosage groups. Arnevaelaylseasnoyfotthheerasvteatriasgtiecanlluymsbiegrnisfiocfanftetdailfofsesriefnicceastiaonmosintgestpheerffievetudsodsiadgneot groups. sacral, Ossification ribs, sternum of the hyoid, vertebrae (cervical, thoracic, lumbar and (manubrium, sternal centers and xiphoid), forelimbs (carpals, metacarpals and phalanges) and phalanges) occurred at similar incidences hindlimbs in litters in (tarsals, metatarsals and all dosage groups. All values were within the ranges observed historically at the Testing Facily. "Significantly different from the vehicle control group (ps 0.01). 003616 418-011:PAGE Ill-7 REFERENCES ~ 1 U.S. Food and Drug Administration (1994). Intemational Conference on Harmonisation; Guideline on detection of toxicity to reproduction for medicinal products. Federal Register, September 22, 1994, Vol. 59, No. 183. 2. U.S. Food and Drug Administration. Good Laboratory Practice Regulations; Final Rule. 21 CFR Part 58. 3. Japanese Ministry of Health and Welfare (1988). Good Laboratory Practice Standard for Safety Studies on Drugs, Pharmaceutical Affairs Bureau, April 1, 1983, amended October 5, 1988. 4. European Economic Community (1989). Council decision on 28 July 1989 on the acceptance by the European Economic Community of an (OECD decision/recommendation on compliance with principlesofgood laboratory practice. Official Journal of the European Communities: Legislation. 32(No. L 315; 28 October): 1-17. 5. Christian, M.S. and Voytek, P.E. (1982). In Vivo Reproductive and Mutagenicity Tests. Environmental Protection Agency, Washington, D.C. National Technical Information Service, U.S. Department of Commerce, Springfield, VA 22161. 6. Christian, M.S. (1984). Reproductive toxicity and teratology evaluations of naltrexone (Proceedings of Naltrexone Symposium, New York Academy of Sciences, November 7, 1983), J. Clin. Psychiat. 45(9):7-10. 7. Lang, P.L. (1988). Embryo and Fetal Developmental Toxicity (Teratology) Control Data in the Charles River Cri:CD7BR Rat. Charles River Laboratories, Inc., Wilmington, MA 01887-0630. (Data base provided by Argus Research Laboratories, Inc.) 8. Institute of Laboratory Animal Resources (1996). Guidefor the Care and Use of Laboratory Animals. National Academy Press, Washington, D.C. 9. Salewski, E. (1964). Farbemethode zum makroskopischen Nachweis von Implantationsstellen am Uterus der Ratte. Arch. Pathol. Exp. Pharmakol. 247:367. : 003617 418-011:PAGE III-8 10. Wilson, J.G. (1965). Methods for administering agents and detecting malformations in experimental animals. Teratology: Principles and Techniques, Wilson, J.G. and Warkany, J.(eds.), University of Chicago Press, pp. 262-277. 11. Staples, RE. and Schell, V.L. (1964). Refinement in rapid clearing technique in the KOH-alizarin red method for fetal bone. Stain Technol. 39:61-63 12. Snedecor, G.W. and Cochran, W.G. (1967). Variance test for homogeneity of the binomial distribution. Statistical Methods, 6th Edition, lowa State University Press, Ames, pp. 240-241 13. Sokal, RR. and Rohlf, F.J. (1969). Bartlett's test of homogeneity of variances. Biometry, W.H. Freeman and Co., San Francisco, pp. 370-371 14. Snedecor, GW. and Cochran, W.G. (1967). AnalysisofVariance. PSpt.ati2s5t8ic-a2l7M5ethods, 6th Edition, lowa State University Press, Ames, 15. Dunnett, C.W. (1955). A multiple comparison procedure for comparing several treatments with a control. J. Amer. Stat. Assoc. 50:1096-1129. 16. WSo.kHa.l,FrR.eRe.maanndaRnodhlCfo,.,F.SJ.an(1F9r6a9n)c.isKcrou,skpapl.-W3a8l8l-i3s89T.est. Biometry, 17. DTeucnhnn,oOm.eJt.ri(c19s664().3):M2u4l1t-i2p5le2.comparisons using rank sums. 18. Siegel, S. (1956). Nonparametric Statisticsfor the Behavioral Sciences, McGraw-Hill, New York, pp. 96-104. 19. Zambrama, MA. and Greenwald, G.S. (1971). Effects of fetal ovarian and placental weight of various number of fetuses in the rat. Biol. of Reprod. 4:216-223. 20. Collins, T.F.X., Welsh, J.J., Black, T.N., Whitby, KE. and O'Donnell iMn.Wu.te,roJr.to(c1a9f8f7e)i.ne.PotFedn.tiCahlermev.erTsoixbiilci.ty2o5f(0s)k:e6l4e7t-a6l6e2f.fects in rats exposed 21. Nishimura, M., lizuka, M., Iwaki, S: and KastA,. (1982). Repairability of Drug-induced "Wavy Ribs" in Rat Offspring. Arzneim -Forsch./Drug Res. 32(l), Nr. 12,, pp. 1518-1522. 003618 418-011:PAGE IIl-9 22. Woo, D. and Hoar, R. (1972). "Apparent Hydronephrosis" as a Normal Aspect of Renal Development in Late Gestation of Rats: The Effect of Methyl Salicylate. Teratology 6:191-196. 23. Khera, K.S. (1981). Common fetal aberrations and their teratologic significance: a review. Fund. and Appl. Toxicol. 1:13-18. 003619 APPENDIX A REPORT FIGURE 003620 . i} J -] - Eo / MATERNAL BODY WEIGHTS Fier A ) ) Co [= | = ee [|e Cd APPENDIX B REPORT TABLES 003622 }8E EEERohP E TEET E L REYLooAEri Pek Ones rest hoo SE CHETar So SF nero SAE, ; ; g EH & ou ev tien amreaeamatmg | ongorn wetawn Pr Sh i SIE EE g Tr 8 wm mm omnmomnm mmm mm meen i T g Mir mmm waren wane wpa view iim Sommommnomnnoonnnooannooonnomw ee Ea amas amsaaa aeaaies anos ae LE ss mn : eR e wanEi wre--------------t pe 8 LLTIonnnoommnonnnommnnoann ATL 2 Rb NL Lue an 3 2 3 3! B R EE R T Eine Hienfo ainsikh Tai ad'sLIbescomiueingof 7concptaven Bohn VI EL RE Ha LT RL en sine 3 2 Sovmrion tarearonn) array mm Beso wmwss 8 ows30-2 wannss.0. - 8 Bren wmmms TH wa: es asoan i Baise nase moses na altar moses ns es moses wa meson wae er sees ne em messes masse esi EI E E y SEeen "Significantly diferent from the vehicle conteo group value (ped. 05). .:i) i mi ------ ---- ------ ---- nan sc ne bE RE RE ET wysp ann} g 8 EE mn wean ein ann ene ane ER AREY i g 2 55 2 J me g8 3  BEE tw in ww dw in g CREE TH ne wat mum gm nam Ps chi a = He intnsiassinstsir ted SEE RIB ; Ea EEE ox on Esa mia moa hme TW wan mem wim wim nem 2 z gr Ww gm #1 Am 8 TEFERSTm wa wan wan pam nan tC TREES wa ww wim vi 8 pr fe "BEET mw wm mam mm gh nem g Ye TV 3  on ooo JRE IEEE TEE 8 mee BED OIE ELI IE ENE Om TEL ma oa we wr - wen i FE SEE rtd ,,, I ss LR, NLR i ggEE HEEE ee a S-- - rasserionse - i : s gu m rSEmRRsE g EET REE A A Sistastasis ses sessessessress esesereres i : g i g pro me a #OoE if piESEmES ; RL -- I =. A vvsssvrivsionss :  EEL fEEs ii FR 3 ; :S 2 mooEr 0 am 3 = rns ME THESES MTENE LIVER MEDIAN108,TANAEA (0.6 OU X0.8OO) prey be 20 1 ALL TISSUES APPEARED WORAL. 3& 8 gs gS H mom oop aBnneOEwRuDp1 EERE EiI mEmmER I i EE ~ i gOMF es mae. Bom EER Ba ome ome aoRw ETRE OaBmE aBm oBom aBml oBmomOBEam oEw H REEEEE EGE Rl rie sa siteprtet everer runs RE ; z : &  Eg = EEE E g mom obon on i i % g EomLoEEEEEEE i g | g EOE EE PPEEEEREEonogBoBEEBREEEBEEE 3 ; & i: Jp He ge g:FE. = 0.o50BE I i 2 g :  HERSEEi ELi L is ik 2 i : t iI EEaEET i : i  HE HESSEEEEEEEEEFFREES mma LAD nnn aa SURE EERE ; 5 [3 SERRE I OE NE BEOEEE NE RA I EEE EEEEEEEEEEEERE Someil pri i iiiliiiiiitdd Bis 3 iii iti i111; s z3 & E gs BA ITEe EERs E EEt REEEa EE tN HENSEEEEEEEEE NEEEE Fm aterm bunt dn dn EB... 2 i: i g HEr bbbi bibo iio todp ihE iR @ oEm LLr L o Lbr li T iR n : 8 : t V Bodh li IIIA RE ER Wai EEE NE 8 & Te i re PSB ; o mmb pod B PEEELEE F FE E rt SE RRR i & > ine : 5 won in dh i To i & g es Emo EOE 2 Time PPGPo H b EEoPOop E PE 3 : nove EEE am om H FoE H Ce -- ntinasentSY Xe masse it, aseoomsaat g : i 3 g s : 2 > 3. z 2 en a an TT ne a cersereessiriass 8 Ey i i i113 i3 Fy g-- c UT.e .. SOE R SN H wnELsLEBorers 2: % ; ToETEEIEEERERIERR EommmRaRmRhan 2 - ne Jpel a Ped Pl =pesped = fr Eh. bh, oan Th a oh heh sh 0 nsota srensounar "Fos sooYRSGkTSwn Sacco 18 ones wr g B EERE RLY g -- : py 3ped pdped Bdpepers aT BY umhey - a 2 OwestmsaeRaRwnRT Le IEwaRaIaRaTaTmeRmLe LE mr ERERRREERARA UL HER] rr : 3 i g DmIa EEaRETEEASRARRRERaALRLAhNEhE ELL] BRE iE i ii ] o mo RRRRRRRLRRR LSS 4 ane or semua Sn Caron uimiavovaet "PETRL to WEIGHTS WERERRCce1hocrovse nO CE ; 3 g B nue mannmnnnnn nEnEE Ena ama 8 Landa LR ARAGA SEES, en SERRE ERR RR g TR rr BR1E0 RchATAe ANNTAahRIDTe oenrgs FB Th RA 0EES . wn A DALLA LLS Tmen R RR RRR S DRLRR, 3 & || P; oSrnniitiiiiiniiainieneneen n i Ho a UN... SO mi omSuEURREREER AEAERENhEh Boon TRaadehbhe heh. g 3 i  mmeullmERERReERTR ERREEETS. T mrEEREE REA os ara PO Te heTh oT TT - Sham" CR LAE ma ae aE a =i ui Ouro TEA/OUART PRIN BOY WEIGHTS RENE RRCORDRD Th Ghat (3) 2 g me ie Ln maREn ERR RRE EERS ncn dl g wm AREA RRS NAELSE, EB oomue dRHI R E NR RER2Eila lan aE E GA RREAI, RAN I g 23 BE Lhhdd dFad g wn ny 8 - " diedndododad biloba baba do dada fob bbvbbbbl obit ctu. Ri i z me iE Dibble dnd wh . dp 5 = = . b 23h fos B a H = a ih . g monphbbbbbndhbans 3 ameasa Ue h d Ao RMdMadndad Saad dRaAdRADaDbaYdake a CTR Lm a. he Ln i IRIEL UR INIIIIN ..eeeeecsrrrnee . g : 2 Foe Ho OOO TOO cin NO. JOR-... errperee dll SER on ann sg & " Boa g mm Ewege EBiSiune. Bm Ele. cr & [EO cr 3 Be Fl rs i... Boom mom 8 - un E Rims ums eo oe ons or oe NL Op SPRANGWITH ATEATIONS We orsrecivmsmwas 3? - & be mai A ---------- S CompaSriser |@88J vom r TEm cEar Sa aotr.i ve may een ane wn ws wa wen om we we as ssn om ws ws in io Sis ro R sin Eon - co 3 3 8 a eam ene a sem um oe oe jr -- Erm, os ight " hrm | or g or 3 3 Bug TTT an mem om wma eee g FEimlamn ,in, TREE wnen nwm wuee uove z Bren MTR pe oscar wn owscarrmion LL wn osscnmon o Wo seo om 8Sse am ene 2Q3A anonesaerins i Arai or ur To, sie n+ os we We RpmEsSmmonric va menco, sen. ee 7 SrinsBAERerosOeL vebtueeeessssss sre HSALEAL ae ---- & 2a 2 Tr TIE artr g oe mn we emt FE wr nn om Ee NL AER OF SPECIMENS VET ALTIRATNIOORN0S7 A moe s Ee, ET) g i Cm - tm | Bn UE mn os ---- 58 L HiTm vn - : nl Si ii ae cruel mri Lecrirs eeeeererenenenn Bee masses bensramsenasers Bo SE me mn ose ons S ume wen ems wn or on ee we oe iBE ni riS RR 5 i P Hy ! 8 g 3 igh dpi ipbo gon Sm, ane 3 :g ooa uwne wow hwss I Err U "o mo ii g in 2 u Ss em ;; ] Bo TNT mes aan oe omy 3g3 aes ot eo em A, we wee or 'oe "- SR wnwma one' Efron vn -' wr Trg 23 8 | nw e -- I, we we & Pnu Dllm 5 an %So-- :: |i APPENDIX C PROTOCOL AND AMENDMENTS 2003711 418-011:PAGE C1 OPRIMEDICA _ a,HorSshaRm PEA 19B044 TeleToplhoornaex:: ((221165)) 444433--88751807 PROTOCOL 418-011 SPONSOR'S STUDY NUMBER: T-6316.7 STUDYTITLE: PURPOSE: JESTINGFACILITY: STUDYDIRECTOR: SPONSOR: STUDY MONITOR: ALTERNATE STUDYMONITOR: iOnraRlat(sGavage) Developmental Toxicity Study of N-EtFOSE NTh-eEpFOuSrEpoonofCtsrhie:sCsDtBuBdyRiVstAoFd/ePtleucstapdrveesrsuemeefdfepcrtesgonfant female rats and development of the embryo and fetus consequent to exposure of the dam from implantation to closure of the hard palate. This study evaluates ICH Harmonised Tripartite Guideline stages C and Dofthe reproductive process. 9Ar0g5uSshReeesheyarDrcihveL,abBourialtdoirnigesA, Inc. Horsham, Pennsylvania 19044-1267 Telephone: (215) 443-8710 Telefax: (215) 443-8587 ARsasyomcioantde GDi.rYeocrtko,r oPfhR.eDs.e,aDrAcBhT 3M Toxicology Services 3M Center, Building 220-2E-02 St. Paul, Minnesota 55144-1000 TMealrevpihnoTn.e:Case(6,51D).V7.3M.3,-5P1h8.D0. Telefax: (651) 733-1773 ATenlderpehwonMe.: Se(a6c5a1t), Ph.D. 575-3161 Telefax: (651) 733-1773 003712 418-011:PAGE C2 : Protocol 41P8a-g0e112 REGULATORYCITATIONS: U.S. Food and Harmonisation; DGruuigdeAldimnieniosntrdaettieocnti(o1n99o4f)t.oxiIcnitteyrntaotrieopnarlodCuocntfieonrefnocremeodnicinal products. FederalRegister, September22, 1994, Vol. 59, No. 183. U.S. Food and Drug Administration. Good Laboratory Practice Regulations; Final Rule. 21 CFR Part 58. JfoarpSaanfeestey MSitnuidsitersyoonf HDeraulgtsh,aMnHd WWelOfradriena(n1c99e7N).umGboeord2L1,abMoarractohr2y6P,r1ac9t9i7c.e Standard aEcucreoppteaannceEcboyntohmeiEcuCroompmeuanniEtcyo(n1o8m8i9)c.CCoomumnucniiltdeyocifsainonOoEnC2D8 Jduelcyis1i9o8n9/roencotmh-e mendation on the European cCoommmpulniiatniceesw:itLhepgriisnlcaitpiloens.of32g(oNood.lLab3o1r5a;to2r8y practice. October): Official 1-17. Journal of REGULATORYCOMPLIANCE: This study will be conducted in compliance with the Good Laboratory Practice (GLP) regulations cited above. ADilrecchtaonrgaensdotrhereSvpiosinosnosro,ftdhaitsedpraontodcmolaisnhtalalinbeeddwoicthumtehnetperdo,toscoilg.ned by the Study aTnhde wQiulalliintsypAecstsucrriatinccalepUhnaitse(sQoAfUt)hewilsltauuddyitinthaeccporortdoacnolc,etwhietrhatwhedaSttaanadnadrdthOeperreapotritn,g Procedures of Argus Research Laboratories, Inc. TachceurfaitnaellyrerpeofrltecwtislltihnecrlauwdedaatsataotbetmaeinntedsidgunreidngbtyhtehpeeSrtfuodrymaDnirceeoctfortthheastttuhdeyraenpdortthat aslilgnaipfpilciacnatbdleeviGaLtPionrsegfurloatmiGonLsPwerergeulfaotliloonwsedocicnutrh,eecaocnhdwuicltlobfethdeessctruidbye.d Should in detail, together with howthe deviation might affect the quality or integrityofthe study. `SCHEMATICOFSTUDYDESIGNANDSTUDYSCHEDULE: See ATTACHMENT 1 to the protocol. 003713 418-011:PAGE C-3 : Protocol 418.011 Page3 TA ESTRTIANC DVEL HICE LE: Identification: TestAtticle: Name: Physical Description: ~~ N-EtFOSE. Waxy solid. Lot/Batch Number: Specific Gravity: FM-3929 (30035, 30037, 30039). ~17. Purty: Expiration Date: 99.1%: May, 2000. Information on the identity, composition, strength and purityofthetest article is on file with the Sponsor. Vehicle: 2% Tween 80 in Deionized Water). RSeuvpeprliseerOasnmdosloitsiMdeemnbrtainfeioPcfroaTcwteesiesonednD8e0lownililzebde Water (R.O. documented in the raw data. NetoitbheeprrtehseenStpionntshoervneohrictlheetShattudwyoDuilrdecitnotreirfseraewwairteh tohfearneysuplottseonfttiahliscosnttudaym.inTahnetrsefliokreel,y no analyses other than those mentioned in this protocol will be conducted. SafetyPrecautions: fGolromvuelsa,timoanspkr,epaaprpartoiporniaatnedeyademipnriosttercattiioonn.anTdhaeuMnaitfeorrima/llaSbafceotayt DaarteatoShbeeewto(rMnSdDuSri)ngis attached to the protocol (see ATTACHMENT 2). Storage: Bulk Test Article: Vehicle Components: Prepared Vehicle: Prepared Formulations: Room Temperature. Room Temperature. RReoforimgeTreamtpeedra(tsuarmep.les to be frozen). AJlullitaenstGaurltbiicnlseksih,iMpamnenatgsertootfhFeoTremsutliantigoFnasc,ilaittytshheopurledviboeusaldydcrietsesdeadddtroetshseaanttdention of telephone number. `Shipments should cartons should be include labeled information conceming storage conditions and appropriately. The recipient should be notified shipping in advance of shipment. 003714 418-011:PAGE C4 : Protocol 41P8a-g0e141 [EORMULATION: EreqofuPerepnaractioyn: Formulations suspensions wil be prepared weeklyatthe Testing Faciity. Detailed preparation procedures are attachedto this protocol (ATTACHMENT 3). AdjusftomrPeurnitty: Thetest article will be considered 100% pure for the purposeof dosage calculations. Testing FacilityReserveSamples: TduhreinTgestthiencgoFurasceotftwhililsrsetsuedryv.e aThseamTpesltein(1g gF)aocifletaycwihlllortesoefrbvuelkatessatmparlteicl(e5 umis)eodfeach lotof vehicle components used during the courseofthe study. Samples will be stored under the previously cited conditions. ANALYSES: Samples additional to those described below may be takenif deemed necessary during the courseofthe study. BulkTest ArticleSampling: INnofoarnmaaltyisonesoonftthhee sbtuabliklitteystofartthieclbeuwliklltbeset conducted during the course of this article is on fle with the Sponsor. study. AnalysesofPreparedFormulations: ActontdhuecrteeqduedsutrionfgtthheeScpoounrssoero,fntoheansatluydsy.esHoofmporgeepnaerietdyteasntdarsttiacblielftoyrimnufloartmiaotnisowniils boen fle with the Sponsor. However, records will be maintained to document how the test article formulations were prepared. `ConcoefTensttArtriclae Fotrmuilatoionns: Cstoundcye.ntDruaptliiocnatoef tshaempplreespa(r2edmfLoremauclha)tiwiolnlsbweitllabkeevnefrriofmiedthdeurfiirnsgt tahnedcloaustrsperoefpatrhaitsion orenmtahienidnagysparmeppalreesd.willObneerseatamipnleedaotftehaechTessettiwnigllFbaceilsthyipapsedbafcorkuapnaslyasmipsl;etsh.e Backup `samples wil be stored under the previously cited conditions and discarded at the Testing Facility upon requestofthe Sponsor. 003715 418-011:PAGE C-5 : Protocol 41P8a-g0e11s `ShippingInstructions: `Samples to be analyzed will be shipped (frozen on dry ice) to: K3rMisEJn.vHiaronnsmeenn,taPlh.TDe.chnology and Safety Services. 9Bu3i5ldBiunsgh2-A3v-e0nu9e STte.lePapuhlo,neM:inn(e6s1o2t)a77585-163031-83331 Telefax: (612) 778-6176 The recipient will be notified in advance of sample shipment. DISPOSITION: ParrteicplaerweildlfboermruelattuimoendstwoitlhbeeSdtiusdcyaMrodneidattortahte tTheestpirnegviFoaucislliytyc.itAeldlardedmraeisnsi.ng bulk test TESTSYSTEM: `SpecianedRsea/soSnftorrSelaecitionn: TbehceauCs:e:CD1B) BitRisVoAnFe/mPlaumsmal(iSparnasgpueec-iDeaswlaecyc)eprtaetdwaansdsweliedcetleyduassetdhetThreosutghSoyusttem fionxdiucsittryyRefroartnoognecnliicniitcya)l; 2s)tutdhiiessstorfadinevhealsopbmeeenntadlemtooxnisctitrya(teemdbrtoyob-efesteanlsitive to dFeavceilloiptmye;natnald t4o)xtihnes;te3s)thiasrttoircliecailsdpahtaarmaancdoleoxgpiecrailelnycaecetxiivsetianttthihsesTpeesctiiensg and strain. Number: Initial population acclimated: 190 virgin female rats. Population selected for study: 125 mated female rats (25 per dosage group). Population assigned to satelite study: a1n9dmtahtreede fpeemraGlreoruaptss (|f,iIvlle apnerd GIrV)ouapsssiIginaenddtVo toxicokinetic evaluation. BWodyeiganhdAgte: Fwheimcahletirmaettshweiyllwbiel obredeerxepdectotehdavtoebbeodaytlweeaisgth6t0sodfay2s00ofgatgoe.22A5ctguealabcohdaytwreeicegihptt,sawtil be recorded the day after receipt and will be documented in the raw data. The weight range will be included in the final report. 003716 418-011:PAGE C6 Protocol 41P8a.g0e11s Sex Femaleratswill be given the test article. Male ratsofthe same source and strain will be used only as breeders and are not considered paofrtt he Test System. Source: Charles River Laboratories, Inc. "The rats will be shipped in filtered cartons byairfreight and/ortruck from Charles River Laboratories, Inc., tothe Testing Facility. Identification: Rats are permanently identified using Monel self-piercing ear tags (Gey Band and Tag Co., Inc., numbers No. MSPT 20101). upon assignment to Male rats are given unique permanent the Testing Facily's breeder malerat identification population. Female rats are assigned temporary numbers at receipt and given unique permanent identification numbers when assigned to the study on the basisofday 0of presumed gestation body weights. ANIMALHUSBANDRY: All cage sizes and housing conditions are in compliance with the Guideforthe Care and Useof Laboratory Animals. Housing: "Theratswill be individually housed in stainless steel, wire-bottomed cages except during the cohabitation period. During cohabitation, each pair of rats will be housed in the male rat's cage. No nesting materials will be supplied because the female rats will be sacrificed before parturition is expected. RT ooe mAim r,peraat ndu Humr idie ty: The animal room is independently supplied with at least ten changes per hour of 100% fRroeoshmatiretmhpaetrahtausrbeeweilnl bpaesmsaeidnttahirnoeudgaht9694.9F7%(1H8ECP)Atfoil7t9ersF(A(i2r6oCCl)eaanndmroonoimt)o.red constantly. Room humidity will also be monitored constantly and maintained at 30% to 70%. Light `mAaninatuationmeadt.icaElalychcodnatrroklpleedri1o2d-whiollurbleiggihtn'a1t2-1h9o0u0r h`odaurrksfElSuoTr.escent light cycle will be 003717 418-011:PAGE C-7 : Protocol 418.011 Page7 Diet: Rats will be given Certified Rodent Diet #5002 (PMI Nutition Intemational) available ad libitum from individual feeders. Water: Water will be available ad fibitum from individual battles attached to the cages or from an automatic watering access system. All water willbefrom a local source and passed pthrroocuegshseadrweavteerrseasosamobsacitsermioesmtbatr;apnreocbeefsosreedwuaste.er Cihsioerxipneecwtieldl btoecaodndteaidtnontohmeore than 1.2 ppm chlorine at the timeofanalysis. Water is analyzed monthly for possible bacterial contamination and twice annually for possible chemical contamination. Contaminants: Neither the Sponsor nor the Study Director is aware of any potential contaminants likely to be present in the certified diet or in the drinking water at levels that would interfere wpietrhftohremerdesbuylttsoheftfheiesdsstuupdpyl.ieTrhoerretfhoorsee,mneontainoanleydseisn otthihserprtohtaoncotlhowislel broeuctionnedluycted. RANDOMAINDZCOAHAT BII TATOION N: cUopmopnutarerri-vagle,nmearalteeadnrdafnedmoamleunriattss. wiAlfltebreaacscsliimganteidont,oviinrdgiivnifdueamlahloeursaitnsgwiollnbteh.ebasisof wciolhlacbointseidstwoitfhabmraeexdiemrummaolfefrivaets,daoynse. mFaelmearlaterpaetrsfweimtahlsepreatr.maTthoezocaohoabbisteartvieodn ipneraiod csomnesairdoefretdhetovbageinataldcaoynt0eonftpsraensdu/omreda gceosptualtaitoonryapnldugasosbisgenrevdetdoiinnsdiitvuidwuilallbheousing, Healthy mated female rats will be assigned to dosage groups generated (weight-ordered) randomization procedures. based on computer- ADMINISTRATION: RR outee andasfoo rChon ice: `The oral (gavage) route was selected for use because: 1) in comparison with the dietary route, the exact dosage can proposed routes for clinical use. be accurately administered; and 2) itis oneof the Meat nd Fh reqo uend cy: Femaleratswill be given the test article once daly on days 6 through 17 of presumed rgeesctoartdieond,btohdeypweeriiogdhtofanodrggainvoegnenaetsaipsp.roDxiomsaatgeelsy will the be adjusted for the most same time each day. recently 003718 418-011:PAGE C8 ' Protocol 41P8a.g0e1s1 RatfoirDo osan geSa elel ctie on: Dosages were selected on the basis ofa dosage-range study (Argus Research Laboratories, Inc., Protocol 418-011P) that tested 0, 1, 5, 10, 25 and 35 mg/kgiday. In that study, body weight gainwas decreased at 10 mg/kgiday and higher dosages, and feed consumption values were reduced at all dosagestested. DC osaogen Levcelse, ntratai ndVooln umess: oo[me |ome[BVorEa pn| [0 eee| +[02 [5|cusoroommnv|n [0[oer| 5[+[5|susonaommnm| [wv oer| 0[2[6|sunorrconmmr|n [ov {oe| = [ 5[| eusonoommemve | ThReeosaavricdwitohoecocnlteaa ro10s0%foo orsdorCtohbpern.c ofdasa clans. TA ESTSN , AL ANY DMES ASURE EMENS TS: Viability: All Periods: Atleast twice daily. CO linib cal serv andla orGt enei ralAo ppean rans ce: Acclimation Period: Weekly. Predosage Period: Day 0 of presumed gestation. Dosage Period: Twice daily.Priorto dosage administration and once approximately one hour postdosage. Postdosage Period: Once daily. `CalpinpircoaplrioabtseebrvyatthieonSstumdayyDbireecrteocroradnedd/omroSrteudfyreMqouneinttolry.than cited above,if deemed BodyWeights: Acclimation Period: Weekly. : Predosage Period: Day 0 and 4 of presumed gestation. 003719 418-011:PAGE C-9 : Protocol 418.011 Pages Dosage Period: Daily. Postdosage Period: Daily. [EeedConsumptionValues (recorded and tabulated): Predosage Period: Day 0 and 4of presumed gestation. Dosage Period: Days 6, 8, 10, 12, 14 and 16of presumed gestation. Postdosage Period: Days 18 and 20 of presumed gestation. Feed consumption values may be recorded more frequentlyif it is necessary to replenish the feed. These intervals will not be tabulated. MatingPerformance: Mating wil be evaluated daily during the cohabitation period and confirmed by observation of spermatozoa in a smearofthe vaginal contents and/oar copulatory plug observed in situ. Caesarean-SectioningObservations: Rats will be Caesarean-sectioned on day 20 of presumed gestation. The fetuses will be removed from the uterus and placed in individual containers. The rats will be. examined for number and distribution of: Corpora Lutea. Implantation Sites. [Placentae appearance (size, color or shapeif abnormal) will be noted in the raw data]. Live and Dead Fetuses. (Alive fetus is defined as one that responds to stimuli; a dead fetus is defined as a term fetus that does not respond to stimuli and that isnotmarkedly autolyzed; dead fetuses demonstrating marked to extreme autolysis are considered to be late resorptions.) Early and Late Resorptions. (A conceptus is defined as a late resorptionifitis grossly evident that organogenesis has occurred;ifthis is not the case, the conceptus is defined as an early resorption.) 003720 418-011:PAGE C-10 : Protocol P41a8g-e01110 EetalObservations: GrossExternalAlterationsandSex: aFentdudseesadwifleltbueseesxaalmsionweidlfboer seexxamainndefdorfogrrossesx eaxntdemfoarlgarlotsesraetixotnesm.alLaatleterreastoiropnstitoonsthe extent possible but such or statistical analyses. observations will not be included in either data summarization ification: `The body weightofeach fetus will be recorded. Only body weights of ive fetuses will be used to determine litter fetal body weight averages. Fetuses will be tagged with identification noting study number, liter number, uterine distribution and fixative. SoftTissueExamination: Aalptperroaxtiiomnasteblyyuosinngea-nhoaadfatlphtfeatfieotnuosfesWiinlseoanc'hs litter wil be examined for soft tissue sectioning technique. The fetuses will be initially fixed in Bouin's solution; sections will be retained in alcohol. SkeletalExamination: eTxhaemrienmeadinfionrgskfeelteutsaelsal(taeprpartoixoinmsaatfetleyr one-halfofthe fetuses in each liter) will be staining with alizarin red S. The fetuses will be initially `added fixed in alcohol; skeletal as a preservative. preparations will be retained in glycerinwiththymol Representative photographsoffetal gross, soft tissue and skeletal alterations will be taken. MET OFSH ACRO IFID CE: Rats willbesacrificed by carbon dioxide an intraperitoneal injection of euthanasia assoplhuytxiioant(ioBne.utLhiavneafseitaus-eDs will be sacrificed Special, by manufactured by Schering-Plough Animal Health). NECROPSY: Gross lesions wil be retained in neutral buffered 10% formalin for possible future evaluation (a table of randomunitswill be used to select one control group rat from which all tissues examined at necropsy will be retained, in order to provide control tissues for any possible histopathological evaluations of gross lesions). Unless specifically cited below, all other tissues wil be discarded. 003721 418-011:PAGE C-11 : Protocol 4P1a8g.e01111 `SatelliteRatsAssignedtoToxicokineticSampleCollection: tOhne tdoaxyic1o8kionfetpirceesvualmueadtigoenstwialtlibone(stahceridfaiycefdoallnodwtihnegftohlelloawsitndgossaagmep)l,esractoslalescstiegdn.ed to iBnltooosdesraummpsleepsar(aatpoprrotxuibmeastealnyd4cmenLtripfeurgerda.t) wTilhleberecsoullltiencgtesdefrruomm(tahpepirnofexriiomravteenl2ay cmaLv)a wtihlel bSepoinmsmoerdifaortealnyalfyrsoizse.n Tohnedlriyveircweilalnbdemeaxicnitsaeidn,ewdefirgohzeedn,(a-7n0dCa)suanmtipllsehsiecptmieonntto (lateral lobe) will be frozen and retained at -70C until shipmenttothe Sponsor for analysis. pRoastssibwillel absedCeasecsrairbeeadn-asbeocvteiofonrerdaatsndasfseitgunseedstwoiltlhbeemeaxianmistnueddy.grFoestsluysetso tahnedextent placentae will be pooled per litter and retained frozen (70C) until shipment to the `Sponsor for analysis Apfltaecrenctoamlpslaetmipolnesofwsialmbpeleshciopllpeecdti(ofnr,oszeenruomn,dlriyveircse)ecttoioKnri(slaJt.erHaalnlsoebne),, Pfhe.tDal.,aantdthe previously cited address for analysis. Both the recipient and the Study Monitor will be: notified in advance of sample shipment. ScheduledSacrifice: Ogrnosdsayne2c0roofpspyroefstuhmeetdhogreasctiact,ioanb,dfoemmianlaelraantsdwpielllvbiec vCiasecsearraewainl-lsbeectpieornfeodr,meadn.d Uateri of `apparently nonpregnant rats will be stained with 10% ammonium sulfide to confirm the absence of implantation sites. RatsFoundDeadorMoribund: Rdealtisvetrhyatwidllieboereaxraemsianceridfifcoerdthbeeccaauusseeooffmdoeraitbhuonrdmcoornidibtuinodn,coanbdoirttiioonn oonr ptrheemdaatyurtehe oabnsderuvtaetriinoen ciosnmtaednets.ofTfheemraaltes wrialtlsbweillexbaemrienceodrdfeodr.grAobsosrtleesdiofnest.usPesreagnndalnorcydesltiavteursed fpeutpussewsi.ll bUteereixoafmaipnpeadretnottlhyeneoxntpernetgpnoasnstibrlaet,suwsililngbethsetasianemdewmiethth1o0d%s ademsmcorinbieudmfor sulfide to confirm the absenceof implantation sites. 003722 418-011:PAGE C-12 Protocol 4Pa1g8e01112 PROPOSEDSTATISTICALMETHODS: Averages and percentages will be calculated. Litter values will be used where appropriate. Additional procedures and/or analyses may be performed, if appropriate. TyofpTee st" |. Parametric A. Bartlett's Test=* Il. Nonparametric A. Kruskal-Wallis Test (75% ties) Significant at p<0.05 Not Significant Significantat p<0.05 Not Significant Nonparametric raeVariance Dunn's Test Significant at ps0.05 | Not Significant B. Fisher's Exact Test (>75% ties) Dunnett's Test Ih.TPesrt foorportiDoatna VofartihaenBcienoTmeisatlfoDrisHtormibougteionneity ab.. PSrtoaptoisrttiicoanlldyastiagnairfiecannottpirnocblabuidleidtiienstahirsecraetpeogrotreyd. as either p<0.05 or p<0.01. c. Used only to analyze data with homogeneity of variance. d. Test for homogeneity of variance. 003723 418.011:PAGE C-13 : Protocol 4P1a8g-e01113 DATAACQUISITION, VERIFICATIONANDSTORAGE: Data will be hand- and/or computer-recorded. Records will be reviewed by the Study Director and/or appropriate management personnel within 21 days after generation. All original records will be stored in the archivesofthe Testing Facility. All original data will be bound and indexed. A copyofall raw data will be supplied to the Sponsor upon ryeeqauresatf.terPmraeisleinrgvoefdtthisesudreasftwiflilnbalerseptoorrte,d aaftttehr ewhTiecshtitnigmeFatchileitSypaotnsnoorchwailrlgbeefocronotnaected to determine the dispositionof these materials. REC TOBOEMR AINTDAINS ED: Protocol and Amendments. Test Article, Vehicle and/or Reagent Receipt, Preparation and Use. `Animal Acquisition. Randomization Schedules. Mating History. "Treatment (if prescribed by Staff Veterinarian). GCleinneicraallOCbosmermveanttiso.ns andlor General Appearance. Blood and Tissue Sample Collection, Processing and Shipment. Body Weights. Feed Consumption Values. Caesarean-Sectioning and Fetal Observations. Gross Necropsy Observations. OPhrogtaongrWeaipghhsts(.i required). Study Maintenance (room and environmental records). Feed and Water Analyses. Packing andlor Shipment Lists. KPEEYRSONNEL: Executive Director of Research: Mildred S. Christian, Ph.D., Fellow, ATS Directorof Research: Alan M. Hoberman, Ph.D., DABT ADsirseocctioartoefDLiarbeoctroartoofryROepseeraartciohnas:ndJSothundyF. DiBraemcettort:, BR.aS.ymond G. York, Ph.D., Manager of Study Coordination: ValeriAe. Sharper, M.S. MaUnsaegeCroommfiAtntiemea:l DOepenraatCi.oLnesbaon,dV-MMe.m0b.er, Institutional Animal Care and Manager of Regulatory Compliance: Kathleen A. Moran, M.S. Consultant, Veterinary Pathology: W. Ray Brown, D.V.M., Ph.D., ACVP DABT 003724 418-011:PAGE C-14 : Protocol 4P1a8g-e01114 EINAL REPORT: AbecfoimnpalriezheednfsoillvoewidnrgacfotnfsiunalltarteipoorntwwiitlhbteheprSeppoanrseodr.onThcoemrpelpeotritonwoilfl tinhcelusdteudthyeand will following: `ExSpuemrmiamreyntaanldDCeosnicglnusainodn.Method. AEpvapleunadtiicoenso:f Test Results. Figures, Summary and Individual Tables Summarizing the Above DGaLtPa,CPormoptloicoalncaendStAastseomceinatt,edReApmoerntsdmofenSutpspoarntdinDgevDiaattiaon(isf,apSptruodpyriDaitree)ctaonrd's QAU Statement. INSTITUTIONALANIMALCAREANDUSECOMMITTEE STAEMN: TInhsetitpurtoiocneadluArneismdaelsCcrairbeedanidn tUhsisepCroomtomciotltehea.veAlblepernorceevdiuerweesddbeystchriebeTdesitnitnhgisFapcrioltiotyc'osl that involve discomfort, study animals will distress or pain to be the conducted animals. in a manner to avoid or minimize TnehceesSspiotnysfoorr'csosnidguncattiunrge tbheilsoswtuddoycaunmdentthse ftahcetftahcatttthhaitsiinsfnoortmaatniounnnceocnecsesranriinlgythe dpurpolciecdatuirveesswteurdey mavaayilbabeloebftoarimneeedtfirnogmtthheesStpatoendsopru.rpNosoeasltoefrtnahteisvteu(diyn.vitro) REFERENCES: 1. CThersitsst.iaEn,nvMi.rSo.nmaenndtaVloyPtreokt,ecPt.iEo.n(A1g9e8n2c).y,IWnaVsihviongRteoprno,dDu.cCt.ivNeaatinodnaMluTteacghenniiccailty Information Service, U.S. Department of Commerce, Springfield, VA 22161 2. Cnharlitsrteixaon,neM(.PSr.o(c1e9e8d4i)n.gsReofprNoadlutcrteixvoenetoSxiycmiptyoasnidumt,erNateowloYgoyrekvaAlcuaatdieomnsyooff Sciences, November 7, 1983), J. Clin. Psychiat. 45(2):7-10. 3. CoLanntgr,olP.DLa.ta(19i8n8t)h.e EChmabrrlyeosRainvdeFreCtHal:CDeDveBlRopRmaetn.taClhaTroxliecsitRyiv(TeerrLaatboolroagtyo)ries, Inc., Wilmington, MA 01887-0630. (Data base providedbyArgus Research Laboratories, Inc.) 4. InstituteofLaboratory Animal Resources (1996). Guideforthe Care and Use of Laboratory Animals. National Academy Press, Washington, D.C. 003725 418-011:PAGE C-15 : Protocol P41a8g-e01115 5. Wmiallsfoonr,maJt.iGo.n(s19in65e)x.peMreimtehnotdaslfoanriamdamlisn.isTteerraitnogloaggye:nPtrsiancnidpldeesteacntidngTechniques (Wilson, J.G. and Warkany, J., eds.), University of Chicago Press, pp. 262-277. 6. StthaepKlOeHs-,aRliEz.arainndreScdhSnelmle,tVh.oLd. (1964). for fetal Refinement bone. Stain in rapid clearing technique Technol. 39:61-63. in 7. Salewski, E. (1964). Implantationsstellen Farbemethode zum am Uterus der Ratte. makroskopischen Nachweis von Arch. Pathol. Exp. Phamakol. 247:367. 8. Sthneedbeicnoormi,alGWdi.strainbudtiCono.chSrtaant,isWti.cGa.l M(e1t8h6o7d).s,V6atrhiaEdnicteiotne,stlfoowrahSotmaotgeeUnneiivteyrsoifty Press, Ames, pp. 240-241. 9. SBoikoamle,trRy,R.W.aHn.dFRroehelm,aFn.Ja.n(d19C6o9.),.SBaanrtlFertatn'csitsecsot,opfp.ho3m7o0g-e3n71e.ityofvariances. 10. SMneetdheocdosr,,6tGh.WE.ditainodn,CloocwharaSnt,atWe.UGn.iv(e1r9s6i7t)y.PrAensasl,ysAimsoesf,Vaprpi.a2n5c8e-.27S5t.atistical 11. Dunnett, CW. (1955). A treatmentswith a control. muJ.ltAimpleer.coSmtpata.riAsssooncp.r5o0c:e1d0u9r6e-f1o1r2c9o.mparing several 12. SWoAkHa.l,FrReRe.maanndaRnodhCfo,.,F.SJ.an(1F9r6a9n)c.isKcrou,skpap.l-W3a8l8l-i3s89T.est. Biomery, 13. Dunn, O.J. (1964). Multiple comparisons using rank sums. Technometrics 6(3):241-252. 14. Siegel, S. (1956). Nonparametric Statisticsfor the Behavioral Sciences, McGraw-Hill, New York, pp. 96-104. 003726 418-011:PAGE C-16 : Protocol 418-011 Page 16 [PROTOCOLAPPROVAL: FOR THE TESTING FACILITY ) cl `Alan M. Hoberman, Ph.D.,DABT Director of Research pay. Rlyefdng G. York, 0.0), DABT Associate Director of Research `Study Director 23 Ju 8 Date dues Date SA WT SE Dena C. Lebo, V.M.D. Member, Institutional Animal Care and Date Use Committee FOR THE SPONSOR N/A Marvin T. Case, D.V.M., Ph.D. Study Monitor Phy fry Date 003727 418-011:PAGE C-17 ATTACHMENT 1 SCHEMATIC OF STUDY DESIGN AND STUDY SCHEDULE 003728 . ATTACHMENT 1 . 418-011:PAGE C-18 ProtocPoalg4e181.00/112 STUDY SCHEMATIC DEVELOPMENTAL TOXICITY STUDY Start of Cohabitation Dosage FReamtas.le PrDeasyu7moefd Gestation DEonsdaogfe Sectioningd PDraeys1u7m0efd PrDeays2u0m0efd Gestation Gestation mm = Dosage Period a = FMoerasadudrietmioennatlsd"etsaeicltsiosneeof"tTheestpsr,otAoncaollyses and b = Fsoefttalisesvauleuaotrisoknesle(taalll)- external, 172 per litte-r 003729 ATTAGHMENT 1 . 418-011:PAGE C-19 Proco a18.011 Page20f2 'SCHEDULE* 11 AUG 88 18 AUG 98 PM-23 AUG 98 AM 19 AUG 98 23 AUG 88 25 AUG 98 - 09 SEP 98 08 SEP 98 - 12 SEP 98 02 DEC 98 Arrival Date - Acclimation Begins. Cohabitation Period. First Possible Day0 of Presumed Gestation. Last Possible Day 0of Presumed Gestation Dosage Period (Days 6 through 17 of presumed gestation). Caesarean-Sectioning Period (Day 20 of presumed gestation). Draft Final Report. a The study initiation dateis the date the Study Director signs the protocol. 063230 418-011:PAGE C-20 ATTACHMENT 2 MATERIAL SAFETY DATA SHEET 5 003731 418-011:PAGE C-21 MDAATTEARISAHLEETSAFETY a3mM Center St. Paul, Minnesota N-E+FOSE 55144-1000 1-800-364-3577 or (612) 737-8501 (24 hours) ACopyrriigohhtt,s 1r9e9s8s,iveMdi.nneCsooptyainMginianngd/aonrddMoawnnulfoaacdtiunrginogf Ctohmipsany. * i15nfoarlmlaotwieodnpfroorvidtehde tphuartp:ose of properly utilizing 3M products 1) 2) ptShrheiiedtottrihrenirfbaougrttremhedeaetmiewconionttphyiisstnhoecorobpttiihaneeitdneeondirtniifgofrniuonlmaollfGwMei,isatrhanrniendsnoogldcahaopnrrgoeofstiuthenrtlwheiesrsseeon. DTIRVAFIDeS:EI1ON0NA:MFEL:U3OMRADCHEBMrIaCnAdLSFluorochemical Alcohol IDoNnUuMaBtEaRJ1U.1P7.5C..7: lire. 0000-.5511113355--0092419455-.23 988--00221111--16168230--06 1SSZUFE.D0:002J-a0n5u7a2r-y229, 1= 98 = - SDUOPCEURMSEENDTE:S:10N-o3v7e7m8b-e7r 05, 1987 0000--5511113355--0190544329--32 1. INGREDIENT C.A.S. NO. PERCENT PPEERFLLUOOORRROOMVOEECXTTAAANNNEEESSSUUULLLFFFOOONNNAAAMMMIIIDDDOOOAALALCCLOOCHHOOOHLLO..L............... ALCOHOL...... ~3164684955185-5-9-0973-3-2-37 ~34449-89-3 8302...000 2.0 --- 7960...000 - 6.0 PFEEROFRLOUSOUROTPAENNETSANUELSFUOLNFAONMAIMDIODOALCOHOL..... 68585-72-6 1.0 = 3.0 2. PHYSICAL DATA BOILING POINT:.......eeessseess VAPORPRESSURE:.......oeesssees VAPOR DENSITY: ......covveereees ESVOALPUOBRIALTIITOYN IRNATWEA:T.E.R.:...........e.s.e.e.s. SPECIFIC GRAVITY:.............. PPHER:CEeNv TvvVeOeLeAnTInLnEs:s.i.i.i.n.s.evevnenenveess VMELITINSG PC. oOOo.i.SuI ouseIerN essTnnoeTrnYeeee: ee:uess G2 111m8 CHo <Ca10lcmEHG20 C >Ca1l.c0 AP2=01 C. n<eg1l.i0g.BUOAG=1 Ca(.of 1.m7eltW)ater=t N0I%A NNIIDD 003732 418.011:PAGE C-22 JMSaDnSu:aryFC-213,0 F1L9U9O8RAD Brand Flyorochemical Alcohol PAGE 2 2. PHYSICAL DATA (continusd) APP`EAmAbReArNCwEaxAyNDsoOlDiOdR: 3. FIRE AND EXPLOSION HAZARD DATA FFFLLUAAAMSMMHAPABBOLLIEENTLL:IIMM.II.TT.SS.v-euLUeEEeLLui:..e..n..s..e..n..s..s.. >N/A148 N/A C Sotatlasn AUTOIGNITION TEMPERATURE: ...... N/A EXT`IaNtGeUr,ISHCIaNrGbonMEDdIiAo:xide, Ory chemical, Fos SPEeHCeaIarAiLtifveFelIlREpprFreIosGtsHeuTcrIteiNvGoerPRpcOlrCoeEtsDhsiUunRrgEe,S:dienmcanldudibnrgeathheilnmegt,appsaerlaft-ucso,ntabiunnekde,r coat apnrdotpeacnttisv,e cbaonvdesrinagroufnodr aerxmpso,sewdaiasrteaasndof letghse, hfeaadc.e mask, and UNUOShUALavFaIrRsEouAsNDDEeXcPomLpOoSsIiOtNioHAnZAsReDcSt:ion for products of combustion. 4. REACTIVITY DATA STABILITY: Stable INCNOotMPAaTpIpBliIcLaIbTlYe.- MATERIALS/CONDITIONS TO AVOID: HAZARDOUS POLYMERIZATION: Hazardous polymerization will not occur. H AZARDOUS DECOMPOS CSaaTrfbuorn, MoHnyodxriodgeen ITION PRODUCTS: anFdluoCrairdbeo,n DTiooxxiicdeV,a poOrxsi,desGasoefs NiotrroPgaernt,icuOxliadteess. of 5. ENVIRONMENTAL INFORMATION SPIRTPLeeoLfrveseolrRncEaSatPlsoONopoStnrEdho:etrehceatslietvcnetioennaqszuairpodmfse,ntth.irsesCMpoSilrDlaSetcofrtoyrspipirnlofltoeerdcmtaimtoainto,enrirvaeelng.tairldaiCtnligeoann, upand 003733 Pesidus. Place in a U.S. DOT-approved container. 418-011:PAGE C23 MS08: FC-10 FLUORAD Brand Fluorochemical Alcohol January 23, 1998 5. ENVIRONMENTAL INFORMATION (continued) PAGE 3 "REoICfnOcMaiMnEecNroDaEmtDbeuDsIdtSniPbOlaSeApLme:artmeirtitaeld.hazCaormdbouusstiownastperodiuncctisnewrialtloriinncltuhdee pHFr.esence Dispose of waste product in a facility permitted to accept chemical waste. . ENVLIaRbOoNrMaEtNoTrAyLtDeAsTtAs: showed (Pinephales promelas) - Nnoo mbioordteaglriatdyataitonw.ater96-sHart.uraLtDiSoOn.FathNeoad Minnow lsetnagttihstiicna3l0lydasyigFnaitfhiecaandtMienfnfoewctegogn told bioconcentration of FC-10 into %fryhatscthu,dy.% suLravbivtaels,tswesihgohwte,d 2a0n0d muscle fillets of channel catfish. "RE`GVUoLlAaTtOiRlYe VOC Less OIrNgFaOnRiMcATICOoNm:pounds: N/A. H20 & Exempt Solvents: N/A. TThSCiAs, prEoIdNuEcCSt, cCoOmSpLl,iesAIWCiSthantdheKorcehae.mical registration requirements of EPFCIRREA HHAAZZAARRDD:CLNAoSS:PRESSURE: No REACTIVITY: No ACUTE: Yes CHRONIC: Yes 6. SUGGESTED FIRST AID EYEInCmOeNdTiAaCtTe:ly flush eyes With large amounts of Water. Get immediate medical attention. SKITcNnoanCetOdaNimTaiAtnCeaTlt:yedwacslhothsiknign. wiIfthsisgonasp/saynmdptloamrsgeocacmuoru,ntscalolf awatpehry.sicRieamno.ve Wash contaminated clothing before reuse and dispose of contaminated shoes. INHIAfLAsTiIgOnNs:/syaptoms occur, remove person to fresh air. If signs/synptons continue, call a physician. IFCiaSmWlmAleLdLiaOaWtpEehDly:ysiacsiandirIeMcMtEeDdIATbEyLYm.ediIcfalswaplelroswoendn,el.inNdeuvceervgoimvietinagnything by Routh to an unconscious person. 003734 418-01:PAGE C-24 MJSaDnSu:aryFC-291,0 F1L9U98ORAD Brand Fluorochesical Alcohol 7. PRECAUTIONARY INFORMATION PAGE 4 EVEvoPiRdOTEeCyTeIOcNo:ntact. Wear safety glasses with side shields. SKIoNiPdROTsEkCiTnIOcNo:ntact. onset. A pair of Wgleoavresappmraodperifartoem gtlhoevfeosllwohweinnghanmdaltienrgialt(hsi)s are PPeerceoommneanldepdr:otecbtuitoynl irtuebbmesr.as Unseecesosnaeryortomorperevofenttheskfionllcoonnitnagct: coveralls. RECoVOouMnMtEiwNliaDtthEiDoanVpEpNrtTooIpLrmAiaTaiItnOetNa:ilnociaselminesoxtshiaoaundssetqubaveteleno,twilurasetecioomanmp.epnrdoepPdrrioavetixedpeosruserusefpfiirlcaiimteiotnrsty. Te exhaust ventilation protection. RESANPvIIoORiSAdHTObaRrpYepartPoRhvOieTndaEgnCdrToeIfisOnpNai:iraracbtcooorrrnsdeanbmcaaesteeWdriitoahnl.aOiSrHbSAoerlrneeecgtulcaootnniecoennosft:rtahteihoanlffoo-lfmlaoswkingdust Nfeesvptiarmaitnoarn,tsfull-face supplied air respirator. PRE`7DVoaEnNnTgoItOtNehaotOr,FouAdgCrhCilInyDkEWNioTtrAhLsmSooIkNaeGpESwaThnIedOnNW:autseirn.g thaisshprhoadnudcst.afWtaesrhheaxnpdolsiendg and before eating. RECSOtMoMrEeNDaEwDaySTfOrRoAmGEh:eat. Keep container closed when not in use. FIRNEonfAlNaDsnEaXbPlLeO.SION AVOIDANCE: OTHNCEooRnstmaPonRkiEinCnaAgtU:iToInSOwNoAokfRiYntgIheNwFOhtRioMlbAeaTcIcuOosNi:nagndp/rtohoridsucstmpsorkoedmuecnatntdicoanlneedardeistnuolsttehcetiniofnorm4atoifon tofhitsheMSDhSa.zardous decomposition MIS HAZARD RATINGS: HPEEARLSTOHN:AL1PROFTLEACMTMIAOBNI:LITXY:(Se1 e pRrEAeCcTaIuVtIiToYn:s, 0 section 7.) EXPOSURE LIMITS 003735 INGREDIENT . VALUE UNIT TYPE AUTH SKIN PERFLUOROOCTANESULFONAMIDO PERFLUOROHEXANESULFONAMIDO AALLCCOOHHOOLL...... 00..11 ~ MoM/GM/3M.3 TTWAa a3Mn PAELRGFOLHUOOLR.OHEvPTrANeEoSsUnLnFsOnNnAnMnIeDOunnnnnaens 0.1 MG/M3 TWA BMY 418-011:PAGE C-25 MSDS: FC-10 FLUORAD Brand Flyorochemical Alcohol Jaruary 29, 1998 PAGE 5 EXPOSURE LIMITS (continued) INGREDIENT VALUE UNIT TYPE AUTH SKIN "PPEERC RFFLLUUOORROOBPUETNATNAENSEUSLUFLOFL NOANMAMIIDDOOeALrCOrHOrL... 0.1 01 MG/M3 Mame THA aM TA 3 Y Y Th+heeS1K0pIaoNitneNgnOtTiAmaTuIlcOoNcu:sontmrLeiimbsburttaeindoensautbnsdotaetnyhece,esoveeiirntadhlielcratbeeyxdpaowsiiurtrbheornb'eyY' toruh,nedeocrurteSaKnIepNoaurstriecfrueolruatrteloy, Dy direc contact With the substance. Vehicles can alter skin absorption. SSOHUR:CE OF3mEXPROeScUoRmEenLdIeMdITEDxApToAs:ure Guidelines 8. HEALTH HAZARD DATA EVENoCOaNdTvAeCrTs:e health effects are expected from eye contact. SKIPNroCdOuNcTtACiTs: not expected to be irritating to the skin. Meaxytenbdeedabtsionreb.ed through the skin and persist in the body for an INHNAaLyATbIeONa:bsorbed by inhalation and persist in the body for an extended tine. IFISnHgAeLsLtOiWoEnD:is not a likely route of exposure to this product. qIulalnnteistsiemsayoofcotuhrisamfatteerriaals.ingle swallowing of relatively large MUTAGENICITY: Not mutagenic in in-vitro assays. REPRSuObDsUtCaTnIcVeEK[aDsEVnEoLtOPMtEeNrTaAtLogeTnOiXIcNSi:n the rat at doses as high as 30 ailligrans per kilogras per day via oral route. OTHER HEALTH HAZARD INFORMATION: TCahliisfoprrnoidauctPriosposniottioknnow6n5. to contain any substances regulated under A Product Toxicity Summary Sheet is available. 003736 418-011:PAGE C-26 MSDS: FC-10 FLUORAD Brand Flyorochemical Alcohol January 29, 1998 PAGE SECTION CHANGE DATES HEADING SECTION CHANGED SINCE November 05, 197 ISSUE Abbreviations: N/D - Not Determined N/A - Not Applicable CA - Approximately TbheecionrfroecrtmataisonofinthethidsatMeatiesrsuieadl. Saf3eHtyMADKaEStaNSOheWeAtRRA(NMTSIDESS), isEXbPeRlEiSeSvEeDdORto MIEMRPCLHIAEND,TABIINLCILTUYDINOGR, FBIUTTNESNSOTFOLRIMAITEPDARTTOI,CULAANRY IMPLIED PURPOSE WOARRCROAUNRTSYEOFOF wPhEeRtFhOReMrANtChEe 3ORM UpSrAoGdEuctOFisTRAfDiEt. forUsearpiasrtriecsuploanrsipbulrepofseoranddetesrumiitnaibnlge for cusaenr'asffemcetthotdheofuseuseandoraapppplliiccaattiioonn.of Gaiv3eMnprtohdeucvta,riestoymeofoffawchtiocrhsatrheat uniquely within the user's knowledge and control, it is essential that . tphaertiucsuelrarevpaulrupaotseethaend3Msuiptraobdlucetfotro duestere'rsminmeetwhohdethoefrusiet oirs fit for a application. 3DuMeptroovitdhees riemnoftoermaptoisosnibiinliteylectthartonieclefcotrrmoniacs atrasnesrfveircemtaoy its customers. have resulted 003737 irnepreersroernst,atioomnisssiaosnstooritsaltceormaptlieotnesnesins otrhisacciunrfaocrym.atioInn,ad3dMitmiaokne,s no iinmfforrrmmaattiionnm oibntatihneedMNfSromavaaildaabtlabeasdeiremacytlvnotfrboem as current 3M. as the 418-011:PAGE C-27 ATTACHMENT 3 TEST ARTICLE PREPARATION PROCEDURE 003738 418-011:PAGE C-28 ATTACHMENT 3 Version: 418-P0r1ot1oc(o2l84J1U8L-08161) Page 1/3 TEST ARTICLE AND CONTROL ARTICLE PREPARATION PROCEDURE Test Article: N-EtFOSE Vehicle: 2% Tween 80, in R.O. Water A. Purpose:oTfhdeospuargpeosseuosfptehnissiopnrsoocfedNu-reEiFsOtoSpEroavniddethae mcoentthrooldafrotrictlhefeoprreopraarlation administration to ratsonArgus Study 418-011. B. General Information: 1. Allsuspension containers will be labeled andcolorcoded. Each label will specify the protocol number, test article identification, Argus batch number, concentration, dosage level, preparation date, expiration date and storage conditions. 2a. Suspensions will be prepared: __ Daily _X_ Weekly _For__daysofuse 2b. Vehicle will be prepared: Daily _X_ Weeky _ For__daysofuse 3. Suspensions will be prepared aat final dosage volume of 5 mLikg. 4. Safety: X_ Gloves, lab coat, goggles or safety glasses and faceshield X_ Dust:Mist Respirator _ Halt-Face Respirator _ Full-Face Respirator/Positive Pressure Hood Tyvek SuivApron 5. Dosage suspensions adjusted for Free base and % Purity. Yes X_ No (Calculations based on 100%) __ FreeBase __ Pury 6. Sampling requirements: Cited in protocol. 7. Storage: Cited in protocol 003739 418-011:PAGE C-29 ATTACHMENT3 Version: 418P0r1o1toc(o2l84J1U8L-08181) Page20f3 TEST ARTICLE AND CONTROL ARTICLE PREPARATION PROCEDURE NOTE: Test article will be preparedas a serialdilutionfrom the high dosage to thelowdosage. Once the final volumes are achieved, stir bars are to be added to the containers; mixing should ocour during sampling and/or administration. C. Preparationof Vehicle 1. Aladbdeltehdecroenqtuaiirneedr.amH oune tthoefwa Ra.t0e.t rdteoio5n0iz+e5dwCa,teardtdothaenraepqpuriorperdiaatmeolyunt of Tween 80 and mix until uniform (See TEST ARTICLE CALCULATIONS). D. TestAticle Suspension Preparation: 1. To preparethe 4.0-mg/mL, Group V suspension, add the required amount oafpptreosptrairattieclley (siSzeeed,TElaSbTelAedRTcoInCtaLiEneCr.ALACdUdLAthTeIOreNqSui)reidntoamaonunt of vehicle andheatthe mixture to 80 +5C for approximately 30 minutes. 2. cOonolcse. th(eBetessutraerttihcelerehaissadivsissiobllveedv;orstpeixn,otvheisrwniilglhatchwiheivlee tthhee sdoelsuitrieodn emulsion.) 3. To prepare the 2.0-mgimL, Group IV suspension, remove the required amount of stock suspension (Group V) (See TEST ARTICLE CALCULATIONS), add the required amountofvehicle and mix. 4. To prepare the 1.0-mg/mL, Group lil suspension, remove the required amount of stock suspension (Group IV) (See TEST ARTICLE CALCULATIONS), add the required amountofvehicle and mix. 003740 418-011:PAGE C-30 ATTACHMENT 3 Version: 418.P0r1ot1oc(o2l84J1U8L.09181) Page 3of3 TEST ARTICLE AND CONTROL ARTICLE PREPARATION PROCEDURE 5. Topreparethe 0.2-mgimL, Group Il amount of stock suspension (Group suspension, remove the lf) (See TEST ARTICLE required CALCULATIONS), add the required amountofvehicle and mix. Written by: Approved by; Clarification: _X No __es Initials/Date : " `~ Date: _Sr-3utay. (See attached clarification form.) UC 9-25 003244. 418-011:PAGE C-31 "P. RIMED ICA _-- Argus50R5esSehaerocHnhoyrLsDarhbiaovrmea,,toBPrUiAeIs1n,5G0In4cA4. TelTaopthootanxe:: ((321158)) 444433-.88751807 PROTOCOL 418-011 ORAL (GAVAGE) DEVELOPMENTAL TOXICITY STUDY OF N-ETFOSE IN RATS SPONSOR'S STUDY NUMBER: T-6316.7 Amendment 1 - 12 August 1998 1. FreqoufPe repnarc atiyon (pag4e and page 1of Attachment 3 to the protocol): Formulations (suspensions) will be prepared daily at the Testing Faciity, rather than weekly. ReaforsChoangne: `This change corrects the protocol. 1 / Eg lan i. Hoberman, Ph.., DABT Date Director of Research nd G. York(Ph.3., DABT Associate Director 9fRsearch Date Study Director (CineClot aigrN/T MDeemnbaeCr.,LIenbstoi,tuVt.ioMn.aDl.Animal Care andte Marvin T. Case, D.V.M. Ph.D. Study Monitor Date Use Committee 0742 OPRIMEDICA> 418-011:PAGE C-32 Sr Research Laboratories, In TelephoHnoer:s(h2a1m5,) P44A31-98074140 Telefax: (215) 443-8587 PROTOCOL 418-011 ORAL (GAVAGE) DEVELOPMENTAL TOXICITY STUDY OF N-ETFOSE IN RATS SPONSOR'S STUDY NUMBER: T-6316.7 Amendment 2 - 11 December 1998 1. Sponsor (page 1of the protocol): The Sponsor is 3M Corporate Toxicology, rather than 3M Toxicology Services. Rea fos rCho angne: `This change was made at the requestof the Sponsor. 2. SpeciaendsRe/asSontfor rSealecitin on (page 5 of the protocol): The test article is biologically active, rather than pharmacologically active in this strain. Rea forsCho angn e: This change was made at the request of the Sponsor to correct the protocol. 3. RR outee andasfoo rChon ice (page 7of the protocol): `The oral (gavage) route is a possible route of human exposure, rather than the one proposed for clinical use. ~ 003743 418-011:PAGE C-33 . ProAtomceonld4Pm1a6eg.n0et1?12 Rea fos r Cho angne: This change was made at the requestof the Sponsotro correct the protocol. I-pse7) Alan 8. Hoberman, Ph.D., DABT Date Director of Research Raymond G. York, Associate Director Study Director 11D6C-98 . DABT Date arch Wesson llonled fo se Wars Tose 1120, 2 DenaC. Lebo, V.M.D. Date Marvin T. Case, D.V.M., Ph.D. Date Member, Institutional Animal Care and ~~ Study Monitor Use Committee 003744 APPENDIX D DEVIATIONS FROM THE PROTOCOL AND THE STANDARD OPERATING PROCEDURES OF THE TESTING FACILITY 003745 418-011:PAGE D-1 DEVIATIONS FROM THE PROTOCOL AND STANDARD OPERATING PROCEDURES OF THE TESTING FACILITY 1. 26 AUG 98 (Da4y of presumed gestation): Clinical observations and body weights were not recorded for the following rats: Dosage Gro1 up I ut wv v Dosage Assigned 0(m(aV/ekhai/cdlaey)) ~~ 128N1u4mb- e1r2s816 1 12839 - 12847 5 12864 - 12872 10 12883 - 12897 20 12912- 12922 "This deviation did not adversely affect the outcomeofthe study because it occurred before initiationof the dosage period and represents a small loss of data across all dosage groups. Al deviations are documented in the raw data. = ferred SAV Rhymphd G.Yor, PhB, OABT AssociateDirector of Reseprch and Study Director 110s Date 003746 APPENDIX E TEMPERATURE AND RELATIVE HUMIDITY REPORTS 47 - ARGUS 418-011:PAGE E-1 Temperature and Relative Humidity Report Location: Room 04 Protocol Number: 418-011 Range of Dates: 11-Aug-1998 13:45 to 12-Sep-1998 10:26 TSpaercgieetsR:aRnagte: TToottaall NNuumbmeoorbffHDeoauyrsrs:: Total Number of Data Points: TSemFpe1r0a1t0urFe | Rela3t0i%ve1H0uTm0i%dity 7643.349 T834340 76 76 Mean (25D): MMeadxiiamnu:m: Minimum: NNuummoobbffPPooeeiinnrrttss iHnigRhan(9g)e:(%): Numobf Peoinrts Low (%): 603 (04| 417 624) 679032 a5471 77 43 76 (00090)|| 766 (e10o00) oo o | o oo Report Genarated: 23-Oct-1998 at 08:15 cowwenrs: reviews ay: 7AUF onre:_fs it' Cumulative by Location (v04.01.97) 003748 APPENDIX F PILOT REPORT 003749: