Document 2NB1p0GwnjwL2O5G5xb5OYz6a

AR)6-02CH Pathology Review of Reported Tumorigenesis in a Two Year Study of FM-3924 in Rats November 25, 1998 BY Pathology Associates International 6217 Centre Park Drive West Chester, Ohio 45069 Telephone (513) 779-9600 Fax (513) 779-9603 603099 Pathology Review of Tumors Reported in Rats Given FM-3924 Introduction: This report presents an independent assessment of tumorigenesis data obtained from a study to determine the chronic toxicity and carcinogenic potential ofthe fluorochemical Carcinogenicity FM-3924 Study of in rats. The study, entitled "Two Year (Diet) Fluorochemical FM-3924 in Rats," was sponsored Toxicity / by the 3M Company, St. Paul, Minnesota. Biophase procedures were conducted at Riker Laboratories, Inc., between April, 1981 and May, 1983 and were in compliance with FDA Good Laboratory Practice (GLP) guidelines. At Riker Laboratories, Leonard J. Sibinski, BA, served as the Study Director. Following in-life procedures, all major organs and tumors were processed into microslides and examined by Robert G. Geil, DVM, DACVP. Dr. Geil's findings and interpretations were incorporated into the final report for the study. For the tumorigenesis review, the complete study report, including all relevant pathology data was forwarded to Pathology Associates International (PAI), West Chester, Ohio for `examination by Richard H. Bruner, DVM, DACVP. The sponsor (3M) regarded original `pathology interpretations by Dr. Geil as adequate, and examination of `microscopic tissue sections was not included in the review process. Specific objectives ofthe review were to evaluate tumor data and to provide an opinion relative to the potential relationship of reported neoplasms with the test material based upon: 1. The incidence and morphology of observed tumors and associated proliferative and non-proliferative lesions. 2. A. literature review to examine the biologic behavior and carcinogenetic potential of similar fluorochemicals, 3. Contemporary knowledgeoftumor mechanistic data, especially with respect to possible epigenetic pathways, 4. Consideration of chronic toxicity, immunosuppression, hormonal modulation or ancillary biochemical interactions which may serve as modulating factorsin the development of tumors in this study, and 5. Personal experience in evaluating rodent carcinogenesis bioassays. Review Procedures and Findings 1. Reported tumorigenesis and ancillary pathologic changes for FM-3924: Based upon reported pathology findings, the review pathologist concurred that treatment-related changes were present at both the 1-Year (interim) and 2-Year (terminal) sacrifices as well as in some unscheduled deaths. At the 1-Year interim sacrifice, treatment-related changes were generally restricted to the liver and were characterized by a dose-dependent increase in hepatocellular cytomegaly, vacuolation `and necrosis along with slightly increased inflammatory cell infiltrates. Cytomegaly ' 6o0a100 alinvder vcaelcluomleattaiboonliwsemreresguelnteirnaglliyn hcyopnesristtreonpthywivitah ptrroeliaftemreantti-oinnodfucpeedropxeirstoumrebastainond/oorf aswnmadoslooltrahrrgeeednluydcoeaptdltarbislbmouiotcdedpreterotficuchsuirloounnmi(c(hPyl-pi4voe5rr0iacee.lnlzyswmeelliinndgucwtiitohn).coHmeppraotmoicesleldulmaertanbecorloissims Treatment-related microscopic findings in unscheduled deaths and animals continued uvonaftciultohtlehaetiltioevnremriinnwabalo(st2h-ssYieegxanerisf).icsaaAncdtrdliifytiicioenniacnlrclelya,usdecedydsptieonrisdihsidtgeehgnetndehoerspaeattioomcnaell(el'susl.paorngcMiyootssotmtsehgneapolabytliasyn,d) haydpeenroplmaasstiacndnocdaurlceisnowmearsewienrcereianscerdeasined itnhefelmiavleers.of both sexes and hepatocellular Based upon reported pathology data, dietary levels of 100 ppm FM-3924 for two ydaeedagerensnoemrreaastsuiloatnneddascianwreuclnilenqoaumsiavisoncciarnelhasihegedhpahdtyoopcseeerlplfluealmsaatrlieccsy.ntoodAmuellgteahsloyui,nghvbaoictnuhcorlseaeatsxieeodsnhaenpadantdionccceryleslatusoleiaddr nnoedouplleass"ms(iwnerebotnhot reported in males, sexes) would be irtegias rldiekdelyasthahtepsaotmoceeollfutlhare"haydpeenropmlaasstiicf cscuyogtngotemesemtgpiaonlrgyatrhywatadsiaaNgonObosEsetLrivcfecodrrihtieenrpiaamtawolecerysetoagmpiepvgleaineldy1(w01a).sannIdtoti3sa0ncohptieepwvmoerdtofhfoyr,tmhaaellsoet,essttahsamstahitgeenpreaedtiiatclo this investigation 2. Literature review of the biologic behavior and carcinogenic potential of fluorochemicals The reviewer regarded the test material (N-ehylperfluorooctanesulfonamido ethanol) laistearatuunrieqrueevixeewnobwiaosticconwdiutchteudnktnoodwenterstmriunceturife-tahcteivbiitoylorgeilcaatliobneshhaivpsi.or, Ainclliumdiitnegd tiunmcolruidgeedneasissu,rvoefysiomfilraordefnltuorcoacrhceimniocgaelnseshisadbiboeaesnsaryesporctoemdp.leTtehde lbiytertahteureNarteiovniaelw TAdiolsxtcihocovouelgroehdg,ynoP.srevocegarrraalcimnro(eglNeeTnvePas)nitsanredbpioosraetslsseactycsosncicweeinmttihifnigc"cjsooumubrpcnlhaerlxosn"iac.nfdluiobnrivooelcsothgieigmcaiatcliaolnessxtrwwaceetrrsee. lffollcuuaootrreiiddd.ee ewAxedprdoeistuairvoean.iallaRlbeyls,eulstteovseporrfaotlvhiredespeeorpstetsru,sdpiieencsctliauvrdeeisnbgroinetfhtlehyeNsTbuiPmolm2oag-iyrceiaalrzaeebfsdifoefacotslsslaooyfwsol:fonsgo-dtieurmm 1. Subchronic toxicity studies in rats with "complex" fluorochemicals have identified treatment-related hepatocellular hypertrophy (cytomegaly) similar to findings with FM-3924. In studies conducted by Van Rafelghem et al, a single intraperitoneal injection of perfluoro-n-decanoic acid (PFDA) in several rodent 603101 species resulted in persistent hepatocellular swelling which, ultrastructurally, was characterized by peroxisomal proliferation(12). In subchronic studies sponsored by the 3M Company, Griffith and Long raodumtiensist(e3)r.edAatmmdoosneisumofpe3rf0luporpomoctoarnoagtreeattoer,ratsr,atmsicdeisapnladyemdonhkeepyastocveilaluolraarl hypertrophy. This change was more prevalent and severe in males. Noteworthy was the observation that all mice given doses of 1000 ppm ad libidum and all monkeys monkeys given given 31000mgm/gk/gk/gd/adyaydi(sgpalvaaygeed) andoireedxipar,eteemremisniasl,lyb.laAcdkdisttiooonlasl,lyf,aciaalll pallor, and prostration; and one monkey given 10 mg/kg/day displayed anorexia, black stools and facial pallor. = Liver effects, however, were not observed in monkeys assigned to these studies. Furthermore, microbial assays using five Salmonella stains and one Saccharomyces strain, with and without metabolic activation, did not reveal mutagenic activity for the test material. 2. Review of studies relating to the chronic toxicity of fluoride and fluoride- containing compounds was limited to the NTP 2-year carcinogenesis bioassay of sstouddiy,umtrfelautomreindte-arenldatseedlepctatehnovliorgoincmcehnatnaglesstuwdeirese of fluoride in not observed cattle. in the In the NTP liver of rats given dietary concentrations of up to 175 ppm sodium fluoride in the drinking water for two years (11). Furthermore,inan extensive survey of cattle exposed to high environmental fluoride concentrations for lifetime periods, increased liver disease or neoplasia was not reported although many animals exhibited dental and skeletal changes typicalofadvanced fluorosis (9). 3. Mechanisms neoplasia in rats resulting in increased given FM-3924: hepatocellular nodular hyperplasia and pRaetshuolltosgiosft.posItsiwbalse rgeeansootonxeidcithtayt stthuidsicesomwpiltehxFfMl-u3o9ro2c4hewmeircaelnportopbraobvliydweadstonotthestrreovnigelyw mutagenic and that most biologic effects were due to perturbation of liver cell p`ametthawbaoylsi.sm,Acwciotrhdipnegrloyx,isitoimsallikperloylitfheartattihoendaenvdelgoepnmereanlt doifsrluivpetriocenlloftummuoltrisp(leanmdentoadbuollairc hyerplasia) was associated with epigenetic mechanisms, possibly including peroxisomal proliferation and genetic damage (ploidy) associated with oxidative stress and/or altered regulation of the cell cycle. Following publication of the final report for this study (1988), numerous journal articles have been published which identify carcinogenesis in rodents following exposure to non-genotoxic test materials. Subsets of these chemicals `which induce liver cell tumors in rodents are characterized by antecendent hepatocyto`megaly and peroxisomal proliferation (2,5,6,8). 603102 : 4p.romEoftfeecrtss:ofancillary biochemical interactions which may have served as tumor Comments relative to all metabolic aberrations which may have influenced liver cell tthuamtorhiegpeanteicsitsoxiincitthyiswsatsuduynewqouuilvdocbaelllyarlgienlkyedspweictulhatiinvgee.stiIotnsohofuFlMd-b3e92n4ot,eda,ndhoawtevheirg,h dpsreoorslveiefdleeravasetliasontliuovmfeorhrecepplarltooamcloyttteerearsti(wo1o0nu)s.lhdaAdlbetrheesoxuuplgethecdtietidnshtnooeuclrredopsaibiser.edmCapomhraargseeisdzpeotndidstishnuaget,lyia,nncdraecmacsaeelyderhlaiatvveeedr dceolelsprnooltifienrvaatiroinabwlhyircehsumltaiyn oicnccureraisnedrotduemntosr damage which promotes in increased mitolic ffoolrlmoawtiionng,emxpoosstupraethtool`omgiasntystaogxriecemathtaetricaellls activity may contribute to tumor formation (4,10,13). 5. Personal opinion relative to a relationship between proliferative lesions and FM-3924 observed in this study: cItoimsbminyedopwiintihoninthcarteadsiesdtinhcytpeirnpclraesatsiecs niondhuelpeastoicnelbloutlharsenxeeosplaarsemcsleianr hiingdhicdaotosres ftehmaatletsh,e tpersotlimfaetraetriivael lsehsoiuolnds baendrengeaorpdledasamss awelirveerccoanrsciidneroegdentoinbSeprsapgounetaDnaeowulseyaltraetrsa.tioOntshoerr secondary to the systemic effects of altered hepatocellular metabolism (7). Based upon histomorphologic changes observed in this study, it is likely that epigenetic mechanisms t(heespceeclilalclyyclpee)rwoexirseokmeayl fparcotloirfseriantitohne, doexivdealtoipvmeesnttroesfs haenpdatootcheelrluflaacrtoprrsolwihfeircahtidveerleegsuiloantse. Ancillary data which would support epigenetic pathways for tumorigenesis in this study r`ewfoeurlednpcerolviitdeeraaturreataivoaniallaeblfeorfsoerletchteiponreopfareaxtpioosnuorfe tthhirsesrheovlidesw,inlihvuermadnasm.ageB,asinecdluudpionng chaerpcaitnoocgyetneosmiesg,alhyasannodtnbeeopelnasrieapoirnterdodiennthsumvaianspeerxopxoissoemdatlo ptreosltifmeartaetriioanlsantdhaatspsorcoimaotteed `metabolic perturbations Additional Information which might Contribute to the Safety Assessment of FM-3924 1. Mutagenesis assays or ancillary procedures to establish the genotoxic potential of the test material. cycle. 2. Cell proliferation studies to provide an index treatment-related increases in the cell 3. Ultrastructural analyses or contemporary analytical procedures to confirm the possible peroxisomal proliferation and ancillary metabolic perturbations. 4. Recovery studies to establish the persistenceofliver cell effects following subchronic exposures to FM-3924, 603103 4 Conclusions: Based upon review ofthe information availableinthe study report, it is my opinion that dietary FM-3924 for 2 years resulted in chronic liver changes (megalocytosis) in males at all dose levels (10, 30 and 100 ppm) and for females at the high dose concentration (100 ppm). At the high dose level, hyperplastic nodules were increased in both sexes and hepatocellular tumors (adenomas and carcinomas) were increased in females. Incidence values for liver proliferative lesionsindicatedthat FM- - 3924 should be conditionsofthis regarded as a liver carcinogen study. The presenceofpersistent, for Sprague Dawley dose-dependent liver rats under the cell cytomegaly suggested that epigenetic mechanisms were causative for tumorigenesis in this study, and that safe exposure thresholds may be established providing that toxicokinetic and ancillary data do not indicate additional adverse effects such as reduced excretion and bioaccumulation. DechaIAT ore f [2s 5 [28 Richard H. Bruner, DVM, DACVP Date References 1. Goodman, D.G., Maronpot, R.R., Newberne, P. M., Popp, J.A. and Squire,AR. Proliferative and Selected Other Lesionsofthe Liver inRats(Draft Proposal). SSNDC Guidesfor Toxicologic Pathology. STP/ARP/AFIP, Wash., D.C. (1993). 2. Green, S. Receptor-Mediated Non-Genotoxic Carcinogenesis: New Models for Risk Assesment? Human Exp. Tox. (10): 390-395 (1991). 3. Griffith, F.D. and Long, J. E. Animal Toxicity Studies with Ammonium Perfluorooctanoate. Am. Ind. Hyg. Assoc. J. (41): 576-583 (1980). 4. Huff,J Absence of Morphologic Correlation Between Chemical Toxicity and Chemical Carcinogenesis. Environ. Health Perspect. 101 (Suppl. 5):45-54 (1993). 5. Lalwani, N.D., Dethloff, L.A., Haskins, J.R., Robertson, D.G., and De La Iglesia, F.A. Increased Nuclear Ploidy, Not Cell Proliferation Is Sustainedinthe Peroxisome Proliferator-Treated Rat Liver. Tox. Path. (25) No.2. 165-176 (1997). 6. McDonald, J.S., Lankas, G.R. and Morrissey, R.E. Toxicokinetic and Mechanistic Considerations in the Interpretationofthe Rodent Bioassay. Tox. Path. (22) No. 2. 124-140 (1994). 7. McMartin, D.N., Sahota, P.S., Gunson, D.E., Han Hsu, H.and Spaet, R.H. Neoplasms and Related Proliferative Lesions in Control Sprague-Dawley Rats from 603104 5 Carcinogenicity Studies. Historical Data and Diagnostic Considerations. Tox. Path. (20) No. 2, 212-225 (1992). 8. Schafer, K.A. Cell Cycle: A Review. Ver. Path. (35) No. 6. 461-478 (1998). 9. Shupe. J.L., Bruner, RH., Seymour, J.L. and Alden, C.L. The PathologyofChronic Bovine Fluorosis: A Review. Tox. Path. (20) No.2. 274-285 (1992). 10. Swenberg, JA. Cell Proliferation and Chemical Carcinogenesis: Conference Summary and (1993). Future Directions. Environ. Health Perspect. 101 (Suppl. 5) 153-158 11. US DepartmentofHealth and Human Services. NTP TR 393. NTP TechnicalReport on Toxicology and Carcinogenesis Studies of Sodium Fluoridein F-344 Rats and B6C3F1 Mice (drinking water) (1990). 12. Van Rafelghem, M.J., Mattie, DR., Bruner, RH. and Anderson, M.E.Pathological `and Hepatic Ultrastructural Effects ofa SingleDose ofPerfluoro-n-decanoic Acid in the Rat, Hamster, Mouse andGuineaPig. Fund. & Appl. Tox. (9), 522-540 (1987). 13. Ward, JM., Uno, H., Kurata, Y., Weghorst, C.M. and Jang, J. Cell Proliferation Not Associated with Carcinogenesis in Rodents and Humans, Environ. Health Perspect. 101 (Suppl. 5): 125-136(1993). 003105