NTP TECHNICAL REPORT ON THE TOXICOLOGY AND CARCINOGENESIS STUDY OF DIETARY ZINC (CAS NO. 5263-02-5) IN SPRAGUE DAWLEY RATS (Hsd:Sprague Dawley SD) (FEED STUDY) Scheduled Peer Review Date: July 13, 2017 NOTICE This DRAFT Technical Report is distributed solely for the purpose of predissemination peer review under the applicable information quality guidelines. It has not been formally disseminated by the NTP. It does not represent and should not be construed to represent NTP determination or policy. NTP TR 592 National Toxicology Program National Institutes of Health Public Health Service U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES FOREWORD The National Toxicology Program (NTP) is an interagency program within the Public Health Service (PHS) of the Department of Health and Human Services (HHS) and is headquartered at the National Institute of Environmental Health Sciences of the National Institutes of Health (NIEHS/NIH). Three agencies contribute resources to the program: NIEHS/NIH, the National Institute for Occupational Safety and Health of the Centers for Disease Control and Prevention (NIOSH/CDC), and the National Center for Toxicological Research of the Food and Drug Administration (NCTR/FDA). Established in 1978, the NTP is charged with coordinating toxicological testing activities, strengthening the science base in toxicology, developing and validating improved testing methods, and providing information about potentially toxic substances to health regulatory and research agencies, scientific and medical communities, and the public. The Technical Report series began in 1976 with carcinogenesis studies conducted by the National Cancer Institute. In 1981, this bioassay program was transferred to the NTP. The studies described in the Technical Report series are designed and conducted to characterize and evaluate the toxicologic potential, including carcinogenic activity, of selected substances in laboratory animals (usually two species, rats and mice). Substances selected for NTP toxicity and carcinogenicity studies are chosen primarily on the basis of human exposure, level of production, and chemical structure. The interpretive conclusions presented in NTP Technical Reports are based only on the results of these NTP studies. Extrapolation of these results to other species, including characterization of hazards and risks to humans, requires analyses beyond the intent of these reports. Selection per se is not an indicator of a substance’s carcinogenic potential. The NTP conducts its studies in compliance with its laboratory health and safety guidelines and FDA Good Laboratory Practice Regulations and must meet or exceed all applicable federal, state, and local health and safety regulations. Animal care and use are in accordance with the Public Health Service Policy on Humane Care and Use of Animals. Studies are subjected to retrospective quality assurance audits before being presented for public review. NTP Technical Reports are indexed in the NIH/NLM PubMed database and are available free of charge electronically on the NTP website (http://ntp.niehs.nih.gov). NTP TECHNICAL REPORT ON THE TOXICOLOGY AND CARCINOGENESIS STUDY OF DIETARY ZINC (CAS NO. 5263-02-5) IN SPRAGUE DAWLEY RATS (Hsd:Sprague Dawley SD) (FEED STUDY) Scheduled Peer Review Date: July 13, 2017 NOTICE This DRAFT Technical Report is distributed solely for the purpose of predissemination peer review under the applicable information quality guidelines. It has not been formally disseminated by the NTP. It does not represent and should not be construed to represent NTP determination or policy. NTP TR 592 National Toxicology Program National Institutes of Health Public Health Service U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES 2 CONTRIBUTORS National Toxicology Program NTP Pathology Working Group Evaluated and interpreted results and reported findings Evaluated slides and contributed to pathology report on 2-year rats (June 24, 2015) N.R. Catlin, Ph.D., Study Scientist A.E. Brix, D.V.M., Ph.D., Study Pathologist G.D. Hill, D.V.M., Ph.D., Coordinator Experimental Pathology Laboratories, Inc. ILS, Inc. C.R. Blystone, Ph.D. A.E. Brix, D.V.M., Ph.D. M.C. Cora, D.V.M. Experimental Pathology Laboratories, Inc. P.M. Foster, Ph.D. M.F. Cesta, D.V.M., Ph.D. National Toxicology Program R.A. Herbert, D.V.M., Ph.D. S.A. Elmore, D.V.M., M.S. M.J. Hooth, Ph.D. National Toxicology Program A.P. King-Herbert, D.V.M. R.A. Herbert, D.V.M., Ph.D. G.E. Kissling, Ph.D. National Toxicology Program B.S. McIntyre, Ph.D. T. Osborne, D.V.M., Ph.D. D.E. Malarkey, D.V.M., Ph.D. National Toxicology Program S.L. Smith-Roe, Ph.D. A.R. Pandiri, D.V.M., Ph.D. M.D. Stout, Ph.D. National Toxicology Program G.S. Travlos, D.V.M. C.C. Shackelford, D.V.M., Ph.D. Experimental Pathology Laboratories, Inc. M.K. Vallant, B.S., M.T. J.M. Ward, D.V.M., Ph.D. S. Waidyanatha, Ph.D. Global Vet Pathology N.J. Walker, Ph.D. K.L. Witt, M.S. Social & Scientific Systems, Inc. M.E. Wyde, Ph.D. Provided statistical analyses Battelle Columbus Operations M.V. Smith, Ph.D., Principal Investigator Conducted study and evaluated pathology findings L.J. Betz, M.S. S.F. Harris, B.S. M.R. Hejtmancik, Ph.D., Principal Investigator D.K. Gerken, D.V.M, Ph.D. Biotechnical Services, Inc. K.A.B. Knostman, D.V.M., Ph.D. Prepared Technical Report Experimental Pathology Laboratories, Inc. S.R. Gunnels, M.A., Principal Investigator Provided pathology review B.F. Hall, M.S. L.M. Harper, B.S. M.H. Hamlin, II, D.V.M., Principal Investigator J.I. Irving, M.A.P. A.E. Brix, D.V.M., Ph.D. D.C. Serbus, Ph.D. C.C. Shackelford, D.V.M., Ph.D. Dynamac Corporation Prepared quality assessment audits S. Brecher, Ph.D., Principal Investigator S. Iyer, B.S. V.S. Tharakan, D.V.M. NOT FOR ATTRIBUTION Peer Review Draft 3 CONTENTS ABSTRACT ................................................................................................................................................................. 5 EXPLANATION OF LEVELS OF EVIDENCE OF CARCINOGENIC ACTIVITY .......................................... 9 PEER REVIEW PANEL .......................................................................................................................................... 10 SUMMARY OF PEER REVIEW PANEL COMMENTS ..................................................................................... 11 INTRODUCTION ..................................................................................................................................................... 13 MATERIALS AND METHODS .............................................................................................................................. 33 RESULTS ................................................................................................................................................................... 47 DISCUSSION AND CONCLUSIONS ..................................................................................................................... 61 REFERENCES .......................................................................................................................................................... 69 APPENDIX A Summary of Lesions in Male Rats in the 2-Year Feed Study of Dietary Zinc ............................................................................................................................. A-1 APPENDIX B Summary of Lesions in Female Rats in the 2-Year Feed Study of Dietary Zinc ............................................................................................................................. B-1 APPENDIX C Genetic Toxicology ...................................................................................................................... C-1 APPENDIX D Hematology Results and Trace Metal Methods and Results ................................................... D-1 APPENDIX E Chemical Characterization and Dose Formulation Studies ................................................... E-1 APPENDIX F Feed and Compound Consumption in the 2-Year Feed Study of Dietary Zinc ..................... F-1 APPENDIX G Ingredients and Nutrient Composition of AIN-93M Modified Low Zinc Feed .................... G-1 APPENDIX H Sentinel Animal Program ......................................................................................................... H-1 Peer Review Draft NOT FOR ATTRIBUTION 4 Dietary Zinc, NTP TR 592 NOT FOR ATTRIBUTION Peer Review Draft 5 ABSTRACT DIETARY ZINCa CAS No. 5263-02-5a Chemical Formula: C2H6O12Zn5 Molecular Weight: ~549.0 Synonym: Zinc hydroxide carbonate Zinc is a naturally occurring element and is ubiquitous in the environment. Zinc itself is stable in dry air, but exposure to moist air results in the formation of zinc oxide or basic carbonate. Due to the reactivity of zinc metal, it is not found as a free element in nature but as a variety of different compounds including zinc chloride, zinc oxide, and zinc sulfate. Zinc and zinc compounds are used across a wide range of industries that include rubber production, animal feed supplementation, as a fertilizer additive, in cosmetics and drugs, as a paint pigment, in dental cements, as a wood preservative, in batteries, in galvanizing and metal work, in textile production, in television screens and watches, and in smoke bombs. Of the zinc compounds, zinc oxide is the most widely used. Zinc was nominated by the Agency for Toxic Substances and Disease Registry (ATSDR) for carcinogenicity and genotoxicity testing based on the increasing size of the population exposed to zinc through dietary