Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 1 of 69 1 2 3 4 5 6 7 8 9 June x, 2006 10 11 EPA HSRB 06 01 12 13 George Gray, Ph. D. 14 Science Advisor 15 Office of the Science Advisor 16 1200 Pennsylvania Avenue, NW 17 Washington, DC 20460 18 19 Subject: April 4 6, 2006 Meeting EPA Human Studies Review Board Report 20 21 Dear Dr. Gray: 22 23 The United States Environmental Protection Agency ( EPA or Agency) requested the 24 Human Studies Review Board ( HSRB) to provide advice on Agency scientific and ethics 25 reviews of completed human studies concerning the following pesticide active ingredients: 26 aldicarb, amitraz, azinphos methyl, dichlorovos ( DDVP), ethephon, methomyl, oxamyl, and 27 sodium cyanide. The studies reviewed included both studies on which the Agency proposes to 28 rely in actions under the pesticide laws and studies that the Agency has decided not to use in its 29 risk assessments, either for ethical or scientific reasons. The enclosed HSRB report addresses 30 the Board's response to EPA charge questions for the Board's consideration at its April 4 6, 31 2006 meeting. 32 33 The HSRB was extremely impressed with the high quality scientific and ethical review 34 brought before the Board. A summary of the Board's conclusions on the scientific and ethical 35 considerations of the human toxicity studies for the eight pesticides under review are provided 36 below. 37 38 Aldicarb 39 40 Scientific considerations 41   The cholinesterase data from the aldicarb human study were reliable for use in the 42 aldicarb single chemical, aggregate risk assessment. 43   The cholinesterase data from the aldicarb human toxicity study were reliable for use in 44 the cumulative risk assessment for N methyl carbamates. 45 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 2 of 69 Ethical considerations 1   The aldicarb human toxicity study failed to fully meet the specific ethical standards 2 prevalent at the time the research was conducted. 3   There was no clear or convincing evidence that the research was fundamentally 4 unethical intended to seriously harm participants or that informed consent was not 5 obtained. 6   There was no clear and convincing evidence of significant deficiencies in the ethical 7 procedures that could have resulted in serious harm ( based on the knowledge available at 8 the time the study was conducted), nor that information provided to participants seriously 9 impaired their informed consent. 10 11 Methomyl 12 13 Scientific Considerations 14   The methomyl human study could be appropriately applied to the inter species risk factor 15 for methomyl and for use in cumulative risk assessment of N methyl carbamates. 16 17 Ethical Considerations 18   The methomyl human toxicity study failed to fully meet the specific ethical standards 19 prevalent at the time the research was conducted. 20   There was no clear and convincing evidence that the research was fundamentally 21 unethical intended to seriously harm participants or that informed consent was not 22 obtained. 23   There was no clear and convincing evidence of significant deficiencies in the ethical 24 procedures could have resulted in serious harm ( based on the knowledge available at the 25 time the study was conducted), nor that information provided to participants seriously 26 impaired their informed consent. 27 28 Oxamyl 29 30 Scientific Considerations 31   Although the Board had some questions about the Agency's conclusions regarding lack 32 of sex difference and of the difference between brain and RBC ChE based on only one 33 species, the Board supported the Agency's conclusion that there were no study 34 deficiencies identified that would have affected the outcome of conclusions of this study. 35 36   Considering the high quality of the design and the conduct of the study, the Board agreed 37 that this intentional human dosing study of oxamyl was sufficiently robust to be used for 38 reducing the 10x inter species ( i. e. animal to human) uncertainty factor in the cumulative 39 risk assessment for the N methyl carbamates. 40 41 Ethical Considerations 42   The oxamyl human toxicity study failed to fully meet the specific ethical standards 43 prevalent at the time the research was conducted. 44 45 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 3 of 69   There was no clear or convincing evidence that the research was fundamentally 1 unethical intended to seriously harm participants or that informed consent was not 2 obtained. 3 4   There was no clear and convincing evidence of significant deficiencies in the ethical 5 procedures that could have resulted in serious harm ( based on the knowledge available at 6 the time the study was conducted) nor that information provided to participants seriously 7 impaired their informed consent. 8 9 Azinphos Methyl 10 11 Scientific Considerations 12   Data from the 28 day repeat oral dose study of azinphos methyl should not be used in 13 developing a point of departure for extrapolation of risk to workers exposed to azinphos 14 methyl via the dermal and inhalation routes. 15 16   Data from the 28 day repeat oral dose study of azinphos methyl cannot be used to inform 17 the inter species factor in the cumulative risk assessment. 18 19 Ethical Considerations 20   The AZM human toxicity study failed to fully meet the specific ethical standards 21 prevalent at the time the research was conducted. 22 23   There was no clear and convincing evidence that the research was fundamentally 24 unethical intended to seriously harm participants or that informed consent was not 25 obtained. 26 27   There was no clear and convincing evidence of significant deficiencies in the ethical 28 procedures that could have resulted in serious harm ( based on the knowledge available at 29 the time the study was conducted) nor that information provided to participants seriously 30 impaired their informed consent. 31 32 DDVP 33 34 Scientific Considerations 35   The DDVP repeat dose human toxicity study was sufficiently robust for developing a 36 point of departure for estimating dermal, incidental oral, and inhalation risk from 37 exposure to DDVP in the single chemical risk assessment. 38 39   The DDVP repeat dose human toxicity study should not be used to support reducing the 40 default 10X inter species factor in the cumulative risk assessment of the organophosphate 41 pesticides. 42 43   The HSRB concluded that the other DDVP human toxicity studies available for the 44 Board's consideration should not be used for determining a reduction in the 10X 45 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 4 of 69 uncertainty factor to derive reference dose values for DDVP based on animal toxicity 1 endpoints. 2 3 Ethical Considerations 4   The DDVP repeat dose oral human toxicity study failed to fully meet the specific ethical 5 standards prevalent at the time the research was conducted. 6 7   There was no clear and convincing evidence that the research was fundamentally 8 unethical intended to seriously harm participants or that informed consent was not 9 obtained. 10 11   There was no clear and convincing evidence of significant deficiencies in the ethical 12 procedures that could have resulted in serious harm ( based on the knowledge available at 13 the time the study was conducted) nor that information provided to participants seriously 14 impaired their informed consent. 15 16 Ethephon 17 18 Scientific Considerations 19   The Board concluded that the scientific quality of both the ethephon 28 day oral toxicity 20 study and the ethephon 16 day human oral toxicity study was not adequate on its own. 21 The 16 day study can be used to inform the ethephon 28 day human oral toxicity study. 22 The Board approved the 28 day study for use in EPA risk assessments, emphasizing that 23 the dose level administered is almost certainly not the lowest dose at which adverse 24 effects are likely to be observed. However, its use in lieu of the animal studies will result 25 in greater protection for exposed human populations. 26 27 Ethical Considerations 28   Both the 28 and 16 day oral human toxicity studies failed to fully meet the specific 29 ethical standards prevalent at the time the research was conducted. 30 31   There was no clear and convincing evidence that the research was fundamentally 32 unethical intended to seriously harm participants or that informed consent was not 33 obtained. 34 35   There was no clear and convincing evidence of significant deficiencies in the ethical 36 procedures that could have resulted in serious harm ( based on the knowledge available at 37 the time the study was conducted), nor that information provided to participants seriously 38 impaired their informed consent. 39 40 Hydrogen Cyanide 41 42 Scientific Considerations 43   Data from the amygdalin trial could be used for establishing a point of departure in the 44 acute dietary risk assessment for hydrogen cyanide. 45 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 5 of 69 Ethical Considerations 1   The hydrogen cyanide human oral toxicity study appeared to meet the specific ethical 2 standards prevalent at the time the research was conducted. 3 4   There was no evidence that the research was fundamentally unethical intended to 5 seriously harm participants or that informed consent was not obtained. 6 7   There was no evidence of significant deficiencies in the ethical procedures that could 8 have resulted in serious harm ( based on the knowledge available at the time the study was 9 conducted) nor that information provided to participants seriously impaired their 10 informed consent. 11 12 Amitraz 13 14 Scientific Considerations 15   The results from the single oral dose study are informed by the human metabolism study 16 such that the single oral dose study is appropriate for developing a point of departure for 17 acute and chronic dietary risk and short term oral exposure. 18 19   The combined results from the single oral dose study and the human metabolism study 20 were not appropriate for developing a point of departure for chronic dietary risk, short 21 term oral exposure, or inhalation risk. 22 23   The majority of the Board concluded that the human dermal study was not appropriate 24 for developing a point of departure for dermal exposures of various durations. 25 26 Ethical Considerations 27   The amitraz acute oral and dermal human toxicity studies failed to fully meet the specific 28 ethical standards prevalent at the time the research was conducted. 29 30   There was no clear and convincing evidence that the research was fundamentally 31 unethical intended to seriously harm participants or that informed consent was not 32 obtained. 33 34   There was no clear and convincing evidence of significant deficiencies in the ethical 35 procedures that could have resulted in serious harm ( based on the knowledge available at 36 the time the study was conducted), nor that information provided to participants seriously 37 impaired their informed consent. 38 39 In conclusion, the EPA HSRB appreciated the opportunity to advise the Agency on the 40 scientific and ethical aspects of human subjects and looks forward to future opportunities to 41 continue advising the Agency in this endeavor. 42 43 44 45 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 6 of 69 Sincerely, 1 2 3 4 Celia Fisher, Ph. D. Chair 5 EPA Human Studies Review Board 6 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 7 of 69 NOTICE 1 2 This report has been written as part of the activities of the EPA Human Studies Review 3 Board, a Federal advisory committee providing advice, information and recommendations on 4 issues related to scientific and ethical aspects of human subjects research. This report has not 5 been reviewed for approval by the Agency and, hence, the contents of this report do not 6 necessarily represent the view and policies of the Environmental Protection Agency, nor of other 7 agencies in the Executive Branch of the Federal government, nor does mention of trade names or 8 commercial product constitute a recommendation for use. Further information about the EPA 9 Human Studies Review Board can be obtained from its website at http:// www. epa. gov/ osa/ hsrb/. 10 Interested persons are invited to contact Paul Lewis, Designated Federal Officer, via e mail at 11 lewis. paul@ epa. gov. 12 13 In preparing this document, the Board carefully considered all information provided and 14 presented by the Agency presenters, as well as information presented by public commenters. 15 This document addresses the information provided and presented within the structure of the 16 charge by the Agency. 17 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 8 of 69 United States Environmental Protection Agency Human Studies Review Board 1 2 Chair 3 4 Celia B. Fisher, Ph. D. Marie Ward Doty Professor of Psychology, Director, Center for Ethics 5 Education, Fordham University, Department of Psychology, Bronx, NY 6 7 Vice Chair 8 9 William S. Brimijoin, Ph. D., Chair and Professor, Molecular Pharmacology and Experimental 10 Therapeutics, Mayo Foundation, Rochester, MN 11 12 Members 13 14 David C. Bellinger, Ph. D., Professor of Neurology, Harvard Medical School 15 Professor in the Department of Environmental Health, Harvard School of Public Health 16 Children's Hospital, Boston, MA 17 18 Alicia Carriquiry, Ph. D., Professor, Department of Statistics, Iowa State University 19 Snedecor Hall, Ames, IA * 20 21 Gary L. Chadwick, PharmD, MPH, CIP, Associate Provost, Director, Office for Human Subjects 22 Protection, University of Rochester, Rochester, NY 23 24 Janice Chambers, Ph. D., D. A. B. T., William L. Giles Distinguished Professor, Director, Center 25 for Environmental Health Sciences, College of Veterinary Medicine, Mississippi State 26 University, Wise Center, Mississippi State, MS 27 28 Richard Fenske, Ph. D., MPH, Professor, Department of Environmental and Occupational Health 29 Sciences, University of Washington, Seattle WA 30 31 Susan S. Fish, PharmD, MPH, Associate Professor, Biostatistics & Epidemiology, Boston 32 University School of Public Health, Co Director, MA in Clinical Investigation 33 Boston University School of Medicine, Boston, MA 34 35 Suzanne C. Fitzpatrick, Ph. D., DABT, Senior Science Policy Analyst, Office of the 36 Commissioner, Office of Science and Health Coordination, U. S. Food and Drug Administration, 37 Rockville, MD 38 39 Kannan Krishnan, Ph. D., Professor, Département de santé environnementale et santé au travail, 40 Faculté de medicine, Université de Montréal, Montréal, Canada 41 42 KyungMann Kim, Ph. D., CCRP, Professor & Associate Chair, Department of Biostatistics & 43 Medical Informatics, School of Medicine and Public Health, University of Wisconsin Madison, 44 Madison, WI 45 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 9 of 69 Michael D. Lebowitz, Ph. D., FCCP, Professor of Public Health & Medicine. University of 1 Arizona, Tucson, AZ 2 3 Lois D. Lehman Mckeeman, Ph. D., Distinguished Research Fellow, Discovery Toxicology, 4 Bristol Myers Squibb Company, Princeton, NJ 5 6 Jerry A. Menikoff, M. D., Associate Professor of Law, Ethics & Medicine, Director of the 7 Institute for Bioethics, Law and Public Policy, University of Kansas Medical Center, 8 Kansas City, KS 9 10 Robert Nelson, M. D., Ph. D., Associate Professor of Anesthesiology and Critical Care, 11 Department of Anesthesiology and Critical Care, University of Pennsylvania School of 12 Medicine, The Children's Hospital of Philadelphia, Philadelphia, PA 19104 13 14 Sean M. Philpott, Ph. D., Research Scientist, David Axelrod Institute, Wadsworth Center for 15 Laboratories and Research, New York State Department of Health, Albany, NY 16 17 Human Studies Review Board Staff 18 19 Paul I. Lewis, Ph. D., Designated Federal Officer, United States Environmental Protection 20 Agency, Washington, DC 21 22 * Not in attendance at April 4 6, 2006 Public Meeting 23 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 10 of 69 TABLE OF CONTENTS 1 2 INTRODUCTION.............................................................................................................................................. 11 3 REVIEW PROCESS.......................................................................................................................................... 12 4 CHARGE TO THE BOARD AND BOARD RESPONSE................................................................................. 13 5 1. ALDICARB.................................................................................................................................................... 13 6 2. METHOMYL ................................................................................................................................................. 19 7 3. OXAMYL ...................................................................................................................................................... 25 8 4. AZINPHOS METHYL...................................................................................................................................... 32 9 5. DDVP......................................................................................................................................................... 38 10 6. ETHEPHON ................................................................................................................................................... 47 11 7. HYDROGEN CYANIDE................................................................................................................................... 54 12 8. AMITRAZ...................................................................................................................................................... 58 13 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 11 of 69 INTRODUCTION 1 2 On February 6, 2006 ( 71 FR 24, 6137), EPA published its final rule for protection of 3 subjects in human research. This new rule included creating an independent Human Studies 4 Review Board ( HSRB) to provide advice, information and recommendations related to the 5 scientific and ethical issues of such research. 6 7 The Food Quality Protection Act ( 1996) mandated that the Agency reassess pesticide 8 tolerances ( i. e., the legal limit of pesticides in food) by August, 2006. The Agency is in the final 9 stages of reassessing all tolerances and has elected for the inaugural April 4 6, 2006 HSRB 10 meeting to focus the first meeting of the Board on those pesticides which are subject to the 11 August, 2006 statutory deadline for tolerance reassessment and which have a human toxicity 12 study being considered for incorporation in quantitative human health risk assessment. The 13 HSRB reviewed toxicity studies involving intentional exposure of human subjects to eight 14 pesticide active ingredients: aldicarb, amitraz, azinphos methyl ( AZM), ethephon, dichlorvos 15 ( DDVP), methomyl, oxamyl, and sodium cyanide. Three of these pesticides are members of the 16 N methyl carbamate ( NMC) common mechanism group ( aldicarb, methomyl, oxamyl); two are 17 members of the organophosphorus pesticides ( OP) common mechanism group ( AZM, DDVP). 18 The three remaining pesticides ( amitraz, ethephon, sodium cyanide) each have toxicity profiles 19 that differ from the other chemicals. 20 21 The Agency prepared a variety of documents which provided background information. 22 These included the proposed and final Human Studies Rule, report from the National Academy 23 of Sciences regarding human studies, EPA, Office of Pesticide Programs' ( OPP's) policy 24 regarding the use of cholinesterase data in human health risk assessments, and a slide 25 presentation providing a broad overview of the OPP's human health risk assessments. The 26 Agency used the framework provided in Emanuel ( 2000) as the basis for its ethics reviews. The 27 Agency described the approach for performing scientific review of human studies and for 28 incorporating human toxicity data into risk assessment. 29 30 For each of the human studies under consideration, the Agency provided the complete 31 study report as submitted to the Agency. For each chemical, the Agency developed a review of 32 the ethical conduct of the study. Each ethics review identified any deficiencies noted in the 33 conduct of the specific study compared to both current ethical standards and the ethical standards 34 prevailing at the time the research was performed. The Agency has intentionally deferred 35 making a final determination of whether an individual study satisfied the ethical standards for 36 acceptability in 40 CFR sections 26.1704   26.1706, pending the advice of the Board. 37 38 For most studies, the Agency develops documents, called Data Evaluation Records 39 ( DERs), containing a scientific review of the study. DERs contain summaries of the study 40 design, methods and results, describe potential deficiencies, and provide conclusions about the 41 usefulness of the study in risk assessment. In addition to the DERs, the Agency prepared for 42 each chemical a " Weight of Evidence" ( WOE) memorandum that discusses the differences and 43 similarities between the human and animal toxic responses to each chemical and characterizes 44 the usefulness of the human toxicity studies for human health risk assessment. The WOE memos 45 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 12 of 69 express the Agency's most current scientific conclusions on which the Agency solicited 1 comments from the Board. 2 3 The HSRB reviewed both studies on which the Agency proposes to rely on actions under 4 the pesticide laws and studies that the Agency has decided not to use in its risk assessments, 5 either for scientific reasons or because they do not meet the standards in EPA's final human 6 studies rule. The Agency asked the HSRB to advise the Agency on a range of scientific and 7 ethics issues regarding how the studies should be assessed against the provisions in 40 CFR 8 sections 26.1701   26.1704 of EPA's final human studies rule. The Board anticipates having a 9 series of meetings to review pesticide human studies relevant to pending re registration, 10 tolerance reassessment and new registration decisions, in accordance with the provisions of the 11 final rule. This report transmits the HSRB's comments and recommendations from its inaugural 12 April 4 6, 2006 meeting. 13 REVIEW PROCESS 14 15 On April 4 6, 2006 the Board had a public face to face meeting in Arlington, Virginia. 16 Advance notice of the meeting was published in the Federal Register " Human Studies Review 17 Board: Notice of Public Meeting and Proposed Candidates for Membership to the Board ( 71 FR 18 46, 12194). At the public meeting, following welcoming remarks from Agency officials, Celia 19 B. Fisher, HRSB Chair, proposed a set of scientific and ethics criteria consistent with the 20 language of 71 FR 24, 6137 to guide Board evaluation of each protocol. The Chair's scientific 21 criteria asked the Board to consider the following two questions: ( 1) Did the research design and 22 implementation meet scientific standards and ( 2) Did the data generated by the protocol have 23 implications for the Agency's Weight of the Evidence ( WOE) review and when applicable 24 aspects of the risk assessment? The Chair's ethics criteria asked the Board to consider three 25 questions: ( 1) Did the study fail to fully meet specific ethical standards prevalent at the time the 26 research was conducted; ( 2) Was the conduct of the study fundamentally unethical ( i. e., 27 specifically was there clear and convincing evidence that the research was intended to seriously 28 harm participants or failed to obtain informed consent); and ( 3) Was the conduct of the study 29 significantly deficient relative to the ethical standards prevailing at the time ( i. e., was there clear 30 and convincing evidence that identified deficiencies could have resulted in serious harm based 31 on knowledge available at the time the study was conducted or the information provided to 32 participants could seriously impair informed consent. The Board then heard presentations from 33 EPA on the following topics: ( 1) introduction ( summary of EPA protections for subjects of 34 human research, science and ethics considerations of human subjects research); ( 2) carbamate 35 pesticides ( science and ethics of aldicarb, methomyl and oxamyl human studies); ( 3) 36 organophosphate pesticides ( science and ethics of azinphos methyl and DDVP human studies); 37 ( 4) other pesticides ( science and ethics of ethephon, sodium cyanide and amitraz human studies). 38 The Board also heard oral public comments from the following individuals: 39 40 Carbamate pesticides 41 Mr. Angus Cameron representing BCG Europe 42 Neil Carmichael, Ph. D. representing Bayer Crop Science 43 Jennifer Sass, Ph. D. representing Natural Resources Defense Council 44 45 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 13 of 69 Organophosphate Pesticides 1 Monty Eberhart, Ph. D. and Mr. Dan Van Geothen representing Bayer Crop Science and 2 Mahktesheim Agan 3 Robert Levine, Ph. D. representing Amvac 4 Mr. Ian Chart representing Amvac 5 Laura Plunkett, Ph. D. representing Amvac 6 Thomas Starr, Ph. D. representing Amvac 7 Jennifer Sass, Ph. D. representing Natural Resources Defense Council 8 Ms. Shelley Davis representing the Farmworker Justice Fund 9 10 Other Pesticides 11 Neil Carmichael, Ph. D. representing Bayer Crop Science 12 13 Following Agency presentations and public comments, the Board deliberated on the 14 charge questions. For their deliberations, the Board considered the materials presented at the 15 meeting, written public comments and Agency background documents on each individual 16 pesticide ( i. e., pesticide human study, Agency data evaluation record ( DER) of the pesticide 17 human study, weight of evidence review, risk assessment and ethics review). 18 19 CHARGE TO THE BOARD AND BOARD RESPONSE 20 21 1. Aldicarb 22 23 Charge to the Board 24 25 Aldicarb is a N methyl carbamate ( NMC) pesticide whose primary toxic effect is 26 neurotoxicity caused by the inhibition of the enzyme acetylcholinesterase, via carbamylation 27 followed by rapid recovery. Aldicarb can, at sufficiently high doses, lead to a variety of clinical 28 signs. The Agency is conducting an acute, aggregate ( single chemical, multi route) risk 29 assessment of aldicarb. In addition, aldicarb is a member of the N methyl carbamate common 30 mechanism group and is thus included in the cumulative ( multi chemical, multi route) risk 31 assessment for the NMCs. 32 33 1.1 Scientific considerations 34 35 The Agency's " Weight of the Evidence" ( WOE) document and Data Evaluation Records 36 ( DERs) for aldicarb describe the study design and results of the aldicarb acute oral, human 37 toxicity study. The WOE document also discusses the Agency's conclusions regarding the 38 usefulness of the human study in the acute, aggregate, single chemical risk assessment and in the 39 cumulative risk assessment for the NMCs. Regarding the aldicarb human study, the Agency has 40 concluded that the study is sufficiently robust for reducing the inter species ( i. e., animal to 41 human) uncertainty factor in the aggregate and the cumulative risk assessments. 42 43 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 14 of 69 The Board was asked to comment on the scientific evidence that supports whether the aldicarb 1 human study is sufficiently robust for reducing the inter species ( i. e., animal to human) 2 uncertainty factor in: 3 a. single chemical, aggregate risk assessment and 4 b. cumulative risk assessment. 5 6 Board Response to the Charge 7 8 Brief Overview of the Study 9 10 The study ( Wyld et al. 1992) consisted of a double blind, placebo controlled, single oral 11 dose of aldicarb in orange juice, taken during the course of a meal. A variety of both subjective 12 and objective observations were accumulated, including red blood cell ( RBC) cholinesterase 13 activities, blood pressure, and respiratory parameters among others. Several dose levels were 14 investigated and several time points were also investigated. 15 16 Critique of Study 17 18 The strengths of the study were: it was designed with multiple doses so a dose response 19 relationship could be studied; it was double blind; a large number of parameters that are relevant 20 to anticholinesterase were studied ( including both subjective data such as headache and several 21 objective physiological measures such as blood pressure, pulse, pulmonary function and saliva 22 production); there were exclusion criteria; there were frequent measurements of cholinesterase 23 over a 1 day period in addition to pre treatment measurements; the observations of clinical signs 24 were made by trained observers; both sexes were studied ( although there were fewer doses and 25 individuals in the female group); and the cholinesterase depression was both dose and time 26 dependent in both males and females. 27 28 The weaknesses of the study were: there were typically only 4 8 individuals per dose; 29 there were fewer female subjects and they were only tested at the middle dose levels; there were 30 a large number of reported signs across all groups, including the placebo, making interpretation 31 difficult; the cholinesterase methodology may have not assayed all of the cholinesterase in 32 RBCs; and the first time point of cholinesterase measurement may have been after the time of 33 peak cholinesterase inhibition. 34 35 The Board also noted that, with respect to statistical analysis, the study was likely to be 36 under powered. With the large number of endpoints and the relatively large between subject 37 variances in almost all endpoints, the number of subjects was probably insufficient to guarantee 38 adequate statistical power. In addition, no adjustments were made for multiple comparisons. 39 Since each endpoint was analyzed separately and because many comparisons were carried out 40 across genders, time points, doses, etc., the overall confidence level is probably much below the 41 nominal 95%. It was also noted that the univariate statistical models that were fitted to the data 42 account for the repeated measures structure of the design, but do not account for the multivariate 43 structure in the measurements. Because endpoints were measured on the same subjects, one 44 could expect meaningful correlations between the various endpoints that could contain additional 45 information about the treatment effects and the effects of covariates. Consequences of not using 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 15 of 69 multivariate methods on this data set include a decrease in the power of statistical tests and 1 erosion of confidence levels that occur when many models are fitted to many endpoints 2 independently as was done in this study. These considerations throw some doubt on some of the 3 study results that suggest no statistically significant treatment effects. 4 5 Nevertheless, while it is clear that the statistical power of the study was low, the data do 6 show a very clear and predictable dose and time dependency in the RBC cholinesterase data. 7 The response of the cholinesterase data is what would be expected of a transient carbamate 8 anticholinesterase. Additionally, it would be expected that blood cholinesterase would be the 9 most sensitive endpoint. RBC cholinesterase is well recognized as a sensitive biomarker of 10 exposure to anticholinesterases. Its inhibition is not responsible for clinical signs of 11 anticholinesterase toxicity, so no cause and effect relationship should be expected between RBC 12 cholinesterase inhibition and changes in any of the physiological parameters measured. 13 Therefore, it is also reasonable that inhibition of the RBC cholinesterase would occur whereas 14 the other parameters related to clinical signs, such as salivation or blood pressure, would not. 15 Lastly, the occurrence of numerous clinical signs in the placebo group gives credence to the 16 conclusion that the RBC cholinesterase inhibition was a treatment related effect whereas clinical 17 signs were not. 18 19 The Board concluded that the results of the study could be used in the WOE analysis to 20 determine a NOEL for RBC cholinesterase and clinical signs in males and that the RBC 21 cholinesterase demonstrated a dose dependent and time dependent pattern of inhibition in both 22 males and females. 23 24 HSRB Consensus and Rationale 25 26 The Board concluded that the cholinesterase data from the aldicarb human study were 27 reliable for use in the aldicarb single chemical, aggregate risk assessment. 28 29 The Agency Data Evaluation Report for Aldicarb ( Sette 1992) suggested that the NOAEL 30 and LOAEL were based upon sweating in males. While sweating is a possible clinical sign 31 resulting from cholinesterase inhibition, the responses in the subjects were not consistently dose 32 related. Although sweating is an objective endpoint, the WOE document indicates that the RBC 33 cholinesterase inhibition would be the more appropriate objective endpoint. There are additional 34 reasons why there is more confidence in the cholinesterase data. First, the blood cholinesterase 35 data, despite their weaknesses, are the most consistent responses to aldicarb exposure, because 36 they follow time and dose dependent patterns, as would be expected of a transient 37 anticholinesterase. Second, the cholinesterase data, though probably based on incomplete 38 fractions of the entire RBC cholinesterase population due to the nature of the analysis method, 39 would still be expected to be internally consistent within this study between samples of treated 40 and placebo individuals, and therefore the inhibition levels can probably be believed. There was 41 a dose and time to recovery related relationship to cholinesterase inhibition in both sexes, so the 42 cholinesterase data are also consistent between the sexes. 43 44 The Board concluded that the cholinesterase data from the aldicarb human toxicity study 45 were reliable for use in the cumulative risk assessment for N methyl carbamates. Aldicarb, 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 16 of 69 being an N methyl carbamate, belongs to this common mechanism group which acts via 1 acetylcholinesterase inhibition. Therefore, the endpoint in the cumulative risk assessment must 2 be cholinesterase inhibition. Moreover, the dose response data from the human study appeared 3 such that BMD and BMDL can be calculated. The HSRB concluded that the aldicarb human 4 study appears to be a scientifically valid study, suitable for use in both the aggregate risk 5 assessment and the cumulative risk assessment. 6 7 While the Board concluded that the cholinesterase data could be reliably used in the 8 cumulative risk assessment, it did note some limitations. The animal data were not supplied so 9 the HSRB cannot address the accuracy of the corresponding animal data. Because of the rapid 10 reactivation of carbamylated cholinesterase, it is unclear whether accurate cholinesterase 11 inhibition values were obtained in the human study or that peak inhibition was measured because 12 the earliest time point measured was 1 hour post dose. The WOE document stated that the 13 human and rat cholinesterase inhibition were comparable at the 0.05 mg/ kg dose level; a 14 question arises as to whether this comparison was made at the same point in the 15 inhibition/ recovery patterns of both species. However, it should be possible to make 16 extrapolations of the possible peak from animal data on the time course pattern of cholinesterase 17 inhibition and recovery, and the Agency is urged to make certain that BMD and BMDL 18 comparisons between animals and humans are being made on comparable times in the 19 inhibition/ recovery pattern. 20 21 1.2. Ethical considerations 22 23 Charge to the Board 24 25 a. The Agency requests that the Board provide comment on the following: 26 27 In light of the ethics committee's instruction that the lay summary be " greatly expanded," 28 and the fact that the materials used to obtain informed consent listed a limited range of 29 symptoms of carbamate toxicity ( excluding some reported as adverse effects in the study), 30 included multiple references to the test material as a drug, and failed to identify dose levels to be 31 administered to male subjects, whether, the materials used to obtain informed consent should be 32 considered significantly deficient relative to the ethical standards prevailing when the study was 33 conducted 34 35 Whether the absence from the protocol of discussion of the potential risks to subjects or 36 benefits to society of conducting the proposed research ( as required by the 1989 Declaration of 37 Helsinki, Principle # 4, with which the research asserted compliance) should be considered 38 significantly deficient relative to the ethical standards prevailing when the study was conducted; 39 40 b. The Agency asks that the Board provide comment on the following, taking into account all 41 that is known about the ethical conduct of this study: 42 43 OPP's conclusion that there is not clear and convincing evidence that the conduct of the 44 research was fundamentally unethical. 45 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 17 of 69 Whether there is clear and convincing evidence that the conduct of the study was 1 significantly deficient relative to the ethical standards prevailing when the study was conducted. 2 3 Board Response to the Charge 4 5 Study Overview 6 7 The study was completed on March 11, 1992. The study sponsor was located in France 8 and the performing laboratory was located in Scotland. The protocol documents specifically 9 stated that the research was conducted in compliance with the Principles of Good Clinical 10 Practice as promulgated by the EMEA CPMP in May 1990, and the Declaration of Helsinki ( in 11 either the 1983 or 1989 version, as cited in two different portions of the study documents). 12 Although the May 1990 document on good clinical practice was the precursor for the subsequent 13 1997 document promulgated by the International Conference on Harmonization, the earlier 14 document does not include any specification of the contents of the informed consent form. 15 However, the supporting and supplementary study documents claimed that the research ethics 16 committee providing independent review and oversight of the research functioned according to 17 the standards of the FDA regulations found in 21 CFR 56. These regulations cross reference the 18 informed consent requirements found in 21 CFR 50 Both FDA regulations were effective on 19 January 27, 1981. As such, the research ethics committee should have applied the FDA 20 regulations from nearly a decade before the conduct of this particular research protocol. It 21 should be noted that the sponsor subsequently claimed that the study also was performed 22 according to the requirements of 40 CFR 26, based on a retrospective analysis published in 2003. 23 24 Critique of Study 25 26 The Board concurred with the Agency on the factual observations of the strengths and 27 weaknesses of the study, as detailed in Carley ( 2006a). However, further comments are 28 warranted on ( 1) the documentation and process of informed consent, and ( 2) the minimization 29 of risk with respect to the study design. 30 31 It is clear that the written documentation for informed consent is not up to the standards 32 found in the above regulations. The Volunteer Information that was given to research subjects as 33 an appendix to the signed informed consent document did not have an adequate discussion of the 34 risks of the research. At the request of the research ethics committee ( dated December 24, 35 1991), the Volunteer Information included the possibility that administration of the test product 36 could result in " gut motility effects ( abdominal pains), effect on eye pupil size and muscle 37 weakness"). In addition to the concerns that the risk information was insufficient in the written 38 consent documentation, selected subjects were enrolled a second time. This was not discussed or 39 in the protocol or the informed consent documentation and therefore raises doubts to the Board 40 about the adequacy of the consent process. In effect, the re dosing of selected subjects suggests 41 that the investigator was unblinded to group assignment, reducing the supposedly double blind 42 study to a single blind study and raising doubts ( unanswered by the documentation) about the 43 sufficiency of informed consent. In particular, the re use of selected subjects suggests that some 44 study participants were randomized to receive placebo versus active compound while other 45 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 18 of 69 subjects were not. As a result, the burden of risks was unlikely to have been distributed evenly 1 among all study participants. 2 3 There were several observations that raised concerns about the extent to which the study 4 design minimized risk to research subjects. First, the dosing schedule did not adhere to a strict 5 dose escalation design. As a result, it is possible that subjects dosed at higher levels would 6 experience significant adverse events that could have been anticipated in a more traditional dose 7 escalation design. The Board recognized that the public comments by Bayer CropScience 8 presented during the public comment period at the HSRB meeting suggests that there was 9 previous human testing data available to indicate that such risks would be unlikely. Second, the 10 stopping rule based on a 70% inhibition of cholinesterase activity is fairly liberal ( i. e. not 11 protective), given that 70% inhibition would likely result in significant adverse effects. 12 Although no subject in the research actually experienced this level of cholinesterase reduction, 13 the selection of such a stopping rule increases the risk to subjects. Finally, the letter to the 14 subjects' general practitioner does not contain any information about the actual study and the 15 product being administered other than the name of the compound. As such, this minimizes the 16 extent to which the opinion of the general practitioner could protect the research subject. 17 18 HSRB Consensus and Rationale 19 20 The Board concluded that: 21 22 a) The aldicarb human toxicity study failed to fully meet the specific ethical standards 23 prevalent at the time the research was conducted. 24 25 b) There was no clear or convincing evidence that the research was fundamentally 26 unethical intended to seriously harm participants or that informed consent was not obtained. 27 28 c) There was no clear and convincing evidence of significant deficiencies in the ethical 29 procedures that could have resulted in serious harm ( based on the knowledge available at the 30 time the study was conducted), nor that information provided to participants seriously impaired 31 their informed consent. 32 33 Informed Consent: While informed consent was obtained, the informed consent 34 documents did not include adequate discussion of the research risks as documented by Carley 35 ( 2006a) and did not fully meet ethical standards prevalent at the time the study was conducted. 36 There was a suggestion that additional aspects of risk may have been discussed in the Volunteer 37 Information. Although the Board recognized that the documentation would not meet today's 38 standards, the lack of documentation does not in and of itself demonstrate a significant 39 deficiency that would have seriously impaired the participants' ability to provide informed 40 consent. 41 42 Participant risk: The Board concurred that the stopping rule based on 70% reduction in 43 cholinesterase activity raised the possibility of exposing subjects to inappropriate risk. However, 44 the Board acknowledged that data existed at the time to suggest that the doses used in the study 45 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 19 of 69 would likely not achieve this level. Thus, there is no evidence that serious harm was intended nor 1 that serious harm could have resulted from the implementation of the study. 2 3 The Board concluded that there was no obvious reason why the Agency cannot rely on 4 the results of this study, as appropriate under current pesticide laws, given the absence of clear 5 and convincing evidence that the research was fundamentally unethical or significantly deficient 6 relative to the ethical standards prevailing when the study was conducted. 7 8 2. Methomyl 9 10 Charge to the Board 11 12 Methomyl is a member of the N methyl carbamate ( NMC) common mechanism group 13 based on its ability to inhibit acetylcholinesterase via carbamylation. The Agency has 14 previously completed the acute, aggregate ( single chemical, multi route) risk assessment of 15 methomyl. At the present time, the Agency is considering the use of the methomyl acute oral, 16 human toxicity study to inform the inter species uncertainty factor used in the cumulative risk 17 assessment of the NMCs. 18 19 Scientific considerations 20 21 The Agency's WOE document and DER for methomyl describe the study design and 22 results of the methomyl acute oral human study. The WOE document also discusses the 23 Agency's conclusions regarding the usefulness of the human study in the cumulative risk 24 assessment for the NMCs. For methomyl, the Agency has concluded that the human toxicity 25 study supports a 10X inter species uncertainty factor for methomyl in the cumulative risk 26 assessment of the NMCs. 27 28 Please comment on the scientific evidence that supports this conclusion. 29 30 Board Response to the Charge 31 32 Study Overview 33 34 A study with methomyl was conducted in human subjects using a double blind, placebo 35 controlled, single ascending dose design ( McFarlane et al. 1998). The defined goal of the study 36 was to determine an acute no adverse effect level ( NOAEL) based on inhibition of plasma and 37 RBC cholinesterase, and in the original protocol, the doses were projected to be 0.1, 0.3, 0.5, 38 0.75, 1 and 1.5 mg/ kg. Methomyl was administered as a bolus dose in a capsule immediately 39 ( about 5 minutes) following a meal, and post dose blood sampling commenced at 15 minutes 40 after dosing. Collection of blood samples for analysis of plasma and RBC acetylcholinesterase 41 activity was continued for 24 hours after dosing. Plasma and RBC acetylcholinesterase activities 42 were compared to a baseline measurement, determined from two predose samples collected 16 43 hours and 30 minutes prior to dosing, for each subject. The results were compared to the placebo 44 group. In addition, numerous physiological parameters including ECG, heart rate, pulse, blood 45 pressure, body temperature, clinical chemistry and hematology parameters, determination of 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 20 of 69 pupil size and salivation were assessed during the entire 24 hour period after dosing. The 1 criterion for not escalating to the next dose was inhibition of RBC cholinesterase exceeding 40%. 2 3 Critique of Study 4 5 The scientific validity of this study was determined by its comparative strengths and 6 weaknesses. The strengths of this study are determined by the comprehensive analysis of 7 cholinesterase inhibition that was obtained including: 8 9   The design and results established a clear dose response and time dependent relationship 10 for inhibition of RBC cholinesterase. This was based on the execution of three dose 11 levels in the ascending dose paradigm. The dosages evaluated were 0.1, 0.2 and 0.3 12 mg/ kg. 13   The analysis of the temporal pattern of RBC cholinesterase inhibition was comprehensive 14 and complete. Specifically, blood samples were collected at 15 minutes after dosing, 15 with 15 minute sampling intervals for the first 2 hours, followed by sample collection for 16 evaluation at 2, 3, 4, 6, 8, 12 and 24 hours after dosing. This time course established the 17 peak of cholinesterase inhibition and assured recovery of enzyme activity in all subjects. 18   The inclusion of clinical endpoints associated with increased cholinergic stimulation as 19 additional endpoints provided additional relevant measures of potential adverse effects 20 during the course of the 24 hour study. 21 22 Several weaknesses of the study were noted, and these weaknesses are also based on aspects 23 relating to its design and conduct and include: 24 25   All subjects were male, with no data obtained from females 26   The group size ( 5/ dose level) was small 27   The establishment of the baseline RBC cholinesterase level from which all subjects were 28 compared was the mean of 2 determinations, which, given the general pattern of 29 variation, reduces the confidence in the calculation of percent inhibition of the enzyme. 30 31 One additional issue concerning the design and execution of this study was the initial 32 proposed dose escalation which ranged from 0.1 to 1.5 mg/ kg ( 0.1, 0.3, 0.5, 0.75, 1, and 1.5 33 mg/ kg) and the criterion to not escalate dose was inhibition of RBC cholinesterase exceeding 34 40%. There was one instance in which this criterion was not adhered to. In the first session of 35 the study, one subject was dosed with placebo and the other received methomyl at 0.1 mg/ kg. 36 The subject receiving methomyl showed evidence of RBC cholinesterase inhibition exceeding 37 40% ( approximately 43%), but this change was determined at 8 hours after dosing. The study 38 sponsor concluded that, based on the general knowledge and understanding of the rapidity of 39 enzyme inhibition observed with the carbamates, this finding was likely to be a spurious result. 40 In review of the data for RBC cholinesterase inhibition for this session, it was also noted that the 41 placebo treated subject in this session showed a 20% decrease in baseline enzyme activity, 42 thereby suggesting a possible, albeit unconfirmed, issue regarding the accuracy and reliability of 43 the analysis of enzyme activity in these samples. Therefore, the sponsor's conclusion that the 44 effect observed in the subject dosed with 0.1 mg/ kg methomyl was most likely to be a spurious 45 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 21 of 69 result was considered scientifically reasonable and justifiable, and the decision to escalate the 1 dose in the next session was not in violation of the protocol. 2 3 In the second session of the study, the lead subject dosed with methomyl at 0.3 mg/ kg also 4 showed a greater than 40% inhibition of RBC cholinesterase. Consequently, the next session 5 completed the five subjects to be used in this group but did not escalate the dose further as per 6 the study protocol. Therefore, the highest dosage evaluated in the study was 0.3 mg/ kg, which 7 was much lower than the original intended escalation up to 1.5 mg/ kg. To establish a better 8 dose response relationship, the study was unblinded, and a dosage of 0.2 mg/ kg was added to the 9 protocol after which the study was re blinded and completed. 10 11 HSRB Consensus and Rationale 12 13 The Board concluded that the methomyl human study could be appropriately applied to the 14 inter species risk factor for methomyl and for use in cumulative risk assessment of N methyl 15 carbamates. 16 17 The HSRB concluded that data from the study in human subjects exposed to methomyl 18 provided a meaningful assessment of the dose response relationship and time course for 19 inhibition of RBC cholinesterase in human subjects. The strengths of the study design and 20 conduct outweigh the indicated weaknesses, and the results are scientifically valid. Therefore, 21 the data can be used to inform the assessment of interspecies uncertainty factors for methomyl. 22 23 Ethical considerations 24 25 Charge to the Board 26 27 2.2.1. The Agency requests that the Board provide comment on the following: 28 29   Whether the investigators' decision to administer a dose to additional subjects in session 30 3, when one subject receiving that dose in session 2 displayed RBC ChEI greater than 31 40%, a response that triggered the protocol's anti escalation provision, should be 32 considered significantly deficient relative to the ethical standards prevailing when the 33 study was conducted; 34 35   Whether the timing of the investigators' report to the ethics committee of the adverse 36 effects observed in one subject during session 2 should be considered significantly 37 deficient relative to the ethical standards prevailing when the study was conducted; 38 39   Whether the failure of the investigators to request approval from the ethics committee for 40 certain amendments to the approved protocol, as required by the protocol, when the 41 changes were administrative and had no effect on the safety of the subjects should be 42 considered significantly deficient relative to the ethical standards prevailing when the 43 study was conducted; and 44 45 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 22 of 69   Whether the absence from the protocol of discussion of the potential risks to subjects or 1 benefits to society of conducting the proposed research ( as required by the Declaration of 2 Helsinki, Principle # 5) should be considered significantly deficient relative to the ethical 3 standards prevailing when the study was conducted; and 4 5 2.2.2. The Agency asks that the Board provide comment on the following, taking into account all 6 that is known about the ethical conduct of this study: 7 8 9   OPP's conclusion that there is not clear and convincing evidence that the conduct of the 10 research was fundamentally unethical. 11 12   Whether there is clear and convincing evidence that the conduct of the study was 13 significantly deficient relative to the ethical standards prevailing when the study was 14 conducted, 15 16 Board Response to the Charge 17 18 Brief Overview of the Study 19 20 The study was performed by a commercial research organization, Inveresk Clinical 21 Research of Edinburgh, Scotland and sponsored by the Haskell Laboratory for Toxicology and 22 Industrial Medicine, E. I. du Pont de Nemours and Company, of Newark, Delaware. The protocol 23 documents specifically state that the research was conducted in compliance with the Principles of 24 Good Clinical Practice promulgated by the International Conference on Harmonization 25 ( CMPM/ ICH/ 135/ 95), and the 1996 version of the Declaration of Helsinki. 26 27 Critique of Study 28 29 The Board concurred with the Agency's factual observations of the strengths and 30 weaknesses of the study, as detailed in Carley ( 2006b). However, further comments are 31 warranted on certain events that took place during the conduct of the study. 32 33 1. Determination That Certain Data Were Spurious 34 35 In session one of the study, one of the subjects exhibited a greater than 40% inhibition in 36 RBC cholinesterase activity at one point, without any signs or symptoms. According to the 37 protocol, such inhibition of cholinesterase was a criterion for not escalating to the next dose 38 level. Because this occurred at 8 hours post dose, study personnel considered this to be a 39 spurious result and went ahead with escalation to the next dose without consulting with the ethics 40 review board ( It is somewhat unclear when the results of the assay became available; the 41 observation about this result is described as being noted " retrospectively at the time of reporting, 42 when there was no sample available for retesting."). If this determination of spuriousness had 43 been incorrect, then giving the following subject an escalated dose might have placed that 44 subject at considerable risk. 45 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 23 of 69 The Board concluded that the determination of spuriousness was reasonable at the time, 1 given the late timing of the change in cholinesterase levels, and the similar change observed in 2 cholinesterase levels for a subject in session one who received placebo. Nonetheless, the Board 3 determined that a far more appropriate action would have been for the researchers to have first 4 consulted with the ethics review committee about repeating that 0.1 mg/ kg dose before 5 proceeding with dose escalation. Moreover, the report of the study results stated that " laboratory 6 tests showing abnormal values for any subject were repeated as often as deemed necessary by the 7 clinical investigator until the test values returned to accepted limits or until an explanation other 8 than compound effect was given." In spite of this, it does not appear that any additional tests 9 were performed; study participants only received scheduled laboratory tests at the 12 hour and 24 10 hour points. Although there is a possibility that these events might have placed subjects at an 11 increased risk of harm, the Board determined that there was not clear and convincing evidence 12 that this could have resulted in serious harm to subjects, particularly given the information 13 known about the compound being tested and the other protections in place as part of the 14 protocol. 15 16 2. Decision to Proceed with Third Session 17 18 In session two of the study, one subject who received an escalated dose of 0.3 mg/ kg 19 exhibited a greater than 40% inhibition in RBC cholinesterase, together with self reported 20 clinical symptoms ( i. e. headache). As noted above, the protocol indicated that dose escalation 21 would not occur if any subject experienced such a level of inhibition. The protocol was followed 22 in this regard: additional subjects were tested at the 0.3 mg/ kg dose, but not at any higher doses. 23 However, the protocol also stated that the study would stop if a subject both had greater than 24 40% inhibition and also demonstrated signs or symptoms of carbamate toxicity ( one of which 25 might be considered to be a headache). Although the Board was concerned that this decision to 26 continue the study and to test additional subjects at the 0.3 mg/ kg dose was made without 27 consulting the ethics review committee, it determined that the provisions of the protocol were 28 sufficiently unclear as applied to these facts ( particularly regarding what constitute signs and 29 symptoms of toxicity); the actions of the investigators were thus consistent with the protocol as 30 written. In addition, given the protections that were in place as part of the protocol, and the 31 information then known about the compound being studied, the Board concluded that there was 32 not clear and convincing evidence that this decision presented a risk of substantial harm to the 33 subjects. Nonetheless, the Board concluded that a more appropriate course of action would have 34 been for the researchers to have consulted with the ethics review board before proceeding with 35 the study. This conclusion and the desirability of having taken similar actions after the 36 apparently " spurious" result in session one is underscored by public comments presented to the 37 Board that the observation of acetylcholinesterase depression at 0.3 mg/ kg was " surprising." 38 There is a danger in explaining away unexpected findings based on prior expectations, and ethics 39 review boards should play an important role in preventing investigators from falling prey to such 40 expectations. 41 42 3. Amendment of the Protocol 43 44 Following the testing of the subjects at the 0.3 mg/ kg level as part of session three, the 45 protocol was amended to allow additional subjects to be tested at the 0.2 mg/ kg level. This 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 24 of 69 change was approved by the chair of the ethics review committee, consistent with procedures for 1 the committee which allowed the chair to approve amendments that did not increase the risk 2 level ( i. e., did not involve testing subjects at a dose level above those approved by the convened 3 board). Although the Board did not conclude this was a formal violation of then existing ethical 4 standards, some members of the Board were concerned that the ethics committee chair's actions 5 were inappropriate and that the protocol change should have been reviewed by the full 6 committee because additional subjects were being exposed to a study compound beyond those 7 contemplated by the unamended protocol. 8 9 HSRB Consensus and Rationale 10 11 The Board concluded that: 12 13 a. The methomyl human toxicity study failed to fully meet the specific ethical standards 14 prevalent at the time the research was conducted. 15 16 b. There was no clear and convincing evidence that the research was fundamentally 17 unethical intended to seriously harm participants or that informed consent was not obtained. 18 19 c. There was no clear and convincing evidence of significant deficiencies in the ethical 20 procedures could have resulted in serious harm ( based on the knowledge available at the time the 21 study was conducted), nor that information provided to participants seriously impaired their 22 informed consent. 23 24 ( 1) The Board concluded that the decision to administer a dose to additional subjects in 25 session three could not be determined to be significantly deficient relative to the ethical standards 26 prevailing when the study was conducted because, as discussed above, that decision could be 27 interpreted as consistent with the terms of the protocol and was not likely to expose the subjects 28 to additional serious harm. 29 30 ( 2) The Board concluded that the timing of the investigators' report to the ethics 31 committee regarding what happened to the subject who experienced cholinesterase inhibition 32 during session two should not be considered significantly deficient relative to the ethical 33 standards prevailing when the study was conducted, given the subject's minimal symptoms and 34 the fact that RBC cholinesterase inhibition by itself is not considered an adverse event. 35 36 ( 3) The Board determined that the failure of the investigators to request approval from the 37 ethics committee for certain amendments to the approved protocol, while deficient relative to the 38 ethical standards prevailing at the time the study was conducted, should not be considered 39 significantly deficient, given that these changes were purely administrative. 40 41 ( 4) The Board determined that the absence from the protocol of discussion of the 42 potential risks to subjects or benefits to society of conducting the proposed research ( as required 43 by the Declaration of Helsinki, Principle # 5) should not be considered significantly deficient 44 relative to the ethical standards prevailing when the study was conducted. The potential risks and 45 benefits are described in the research subject information sheet/ informed consent document. 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 25 of 69 While not in the protocol itself, they are nonetheless included in the materials provided to the 1 ethics committee ( and to the potential research subject). Thus, the ethics committee and the 2 potential research subjects had the opportunity to weigh the risks and benefits. 3 4 The Board concluded that there was no obvious reason why the Agency cannot rely on 5 the results of this study, as appropriate under current pesticide laws, given the absence of clear 6 and convincing evidence that the research was fundamentally unethical or significantly deficient 7 relative to the ethical standards prevailing when the study was conducted. 8 9 3. Oxamyl 10 11 Charge to the Board 12 13 Similar to aldicarb and methomyl, oxamyl is a member of the N methyl carbamate 14 ( NMC) common mechanism group based on its ability to inhibit acetylcholinesterase via 15 carbamylation and is thus included in the NMC cumulative risk assessment. The Agency has 16 previously completed the acute, aggregate ( single chemical, multi route) risk assessment of 17 oxamyl. The Agency is now considering the use of the oxamyl acute oral, human toxicity study 18 to inform the inter species uncertainty factor in the cumulative risk assessment of the NMCs. 19 20 3.1. Scientific considerations 21 22 The Agency's WOE document and DER for oxamyl describe the study design and results 23 of the oxamyl acute oral human toxicity study. The WOE document also discusses the Agency's 24 conclusions regarding the usefulness of the human study in the cumulative risk assessment for 25 the NMCs. For oxamyl, the Agency has concluded that the human toxicity study is sufficiently 26 robust for reducing the 10X inter species ( i. e., animal to human) uncertainty factor in the 27 cumulative risk assessment. 28 29 Please comment on the scientific evidence that supports this conclusion. 30 31 Board Response to the Charge 32 33 Study Overview 34 35 The stated primary objective of this randomized, double blind, ascending oral dose study 36 with oxamyl ( McFarlane and Freestone 1999a) was to estimate the no observable adverse effect 37 level ( NOAEL) for oxamyl in humans after oral administration. 38 39 The study used 40 healthy male subjects, aged 19 39. Each was given a single oral dose 40 of oxamyl in a gelatin capsule at doses of 0 ( placebo), 0.005, 0.015, 0.03, 0.06, or 0.15 mg/ kg 41 BW. With the exception of dose sessions 1 ( 2 placebos), 2 ( 1 placebo and 1 at 0.005), 7 ( 1 42 placebo and 4 at 0.09), 8 ( 1 placebo and 1 0.15), and 9 ( 1 placebo and 4 at 0.15), all other dose 43 sessions had 1 placebo, 4 at current dose, and 1 at the next higher dose. Plasma and RBC 44 cholinesterase activity were assayed at screening and 2 days, 16 hours and 30 minutes prior to 45 dosing and at 0.25, 0.5, 0.75, 1.0, 1.25, 1.5, 1.75, 2, 3, 4, 6, 8, 12, and 24 hours post dose and 7 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 26 of 69 days post dose. Other clinical parameters indicative of carbamate exposure, such as urinalysis, 1 pupillometry, saliva increase etc were also measured. 2 3 No treatment related effects for ECG, heart rate, pulse, blood pressure, respiratory rate, 4 body temperature, hematology, clinical chemistry, urinalysis, or pupillometry were recorded. 5 6 At 0.09 mg/ kg b. w., 7 12% plasma and RBC ChE inhibition was observed with three of 7 five volunteers exhibiting greater than 20% plasma ChE inhibition. Therefore, 0.06 mg/ kg b. w. 8 was considered the NOAEL. The Agency generated BMD and BMDL estimates of 0.083 mg/ kg 9 and 0.069 mg/ kg, respectively, based on the RBC ChE data from the study 10 11 Critique of the Study 12 13 The scientific validity of this study was analyzed by weighing its strengths and 14 weaknesses in order to determine if the study was sufficiently robust to be of use in the Agency's 15 risk assessment for oxamyl. 16 17 Appropriate endpoints for this study were selected. Acetyl cholinesterase inhibition in 18 the nervous system was viewed as a key molecular event in the mechanism of toxicity of 19 carbamate compounds. Measurements of acetyl cholinesterase inhibition in both RBCs and 20 brain are considered to be biomarkers of exposure for these compounds. In its policy on the se 21 of data on cholinesterase inhibition for risk assessments of organophosphorus and carbamate 22 pesticides ( USEPA, 2000), the Agency stated that it will treat cholinesterase inhibition in the 23 blood as a surrogate measure for the peripheral nervous system in animals and for the peripheral 24 and central nervous system in humans. 25 26 The animal data for oxamyl were not available to the Board to allow it to make an 27 independent judgment as to whether the methods used to determine acetyl cholinesterase 28 inhibition in the animal studies and the human study were comparable. According to EPA, 29 however, the methods used in the animal studies are comparable to that in the human study. 30 31 Previous animal data was used to predict that the starting dose 0.005 mg/ kg would have 32 no detectable effect on humans. Oral administration of the compound is the most predictive 33 mode of administration since exposure to residues would come from food consumption. There 34 was no justification for the sample size of 4 for oxamyl for each dose session. 35 36 Forty healthy males of similar age and weight were used in this study. A weakness of 37 this study is that no female subjects were used in this study and there was no indication that any 38 human female data exists from other studies. However, the Agency advised the Board that, based 39 upon its evaluation of the animal data, no sex differences would be expected. 40 41 The specific schedule for dosing was described in protocol. It was an ascending double 42 blind, placebo controlled study. Dose escalation did not occur if there were clinically significant 43 symptoms/ signs of toxicity. Stopping rules were equal to or greater than 40% inhibition of RBC 44 cholinesterase at a single time point or 25% at two consecutive time points. If there were 45 associated symptoms/ signs of carbamate toxicity, no further dose escalation would occur. 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 27 of 69 1 A repeated measures analysis of variance was used to analyze the percentage change 2 from baseline in BRC and plasma cholinesterase activity, pupillometry and saliva collection. At 3 each time point of measurements, a test for linear trend with dose was performed using a linear 4 contrast. If the test for linear trend was significant at the 5% significance level, the pairwise 5 comparisons at each timepoint were not adjusted for multiple comparisons. Otherwise, a 6 Bonferroni adjustment was applied to the pairwise test at the 0.83% significance level. 7 8 At 0.09 mg/ kg b. w., 7 12% plasma and RBC ChE inhibition was observed with three of five 9 volunteers exhibiting greater than 20% plasma ChE inhibition. Therefore, 0.06 mg/ kg b. w. was 10 considered the NOAEL. The Agency generated BMD and BMDL estimates of 0.083 mg/ kg and 11 0.069 mg/ kg, respectively, based on the RBC ChE data from the study 12 13 The Board highlighted the strengths of the study as follows: 14 15 ( a) The study design, including both the increasing dose levels and the time point of 16 observations/ measurements, was very rich in information content to allow evaluation of dose 17 response and estimation of the NOAEL and the dose escalation in a step wise fashion was 18 scientifically sound considering the uncertainties in the risk. 19 20 ( b) The study had a very clearly defined dose escalation scheme with properly justified 21 stopping rules. The schedule of observation and measurements were frequent enough to give 22 sufficient information on the time course effect of oxamyl on subjects. 23 24 ( c) The statistical methods used for analysis, including the adjustment for multiple 25 comparisons between placebo and active doses when there was no statistical significance in the 26 linear trend test, were quite adequate for the study. 27 28 The Board highlighted the weaknesses of the study as follows: 29 30 ( a) There appears to be no specific justification for the sample size of four for oxamyl for 31 each dose session and no traditional statistical test of effect size regarding hypotheses. Because 32 the study was exploratory versus confirmatory, the lack of justification was not critical. 33 Nevertheless it would have been helpful to know what effect size could be detected between the 34 placebo and the active dose groups given the sample size of the study. 35 36 ( b) Because of lack of justification for the sample size, the lack of statistical significance 37 in the observed difference between placebo and active doses could not be interpreted 38 appropriately. Specifically the statement in the Agency Data Evaluation Report ( Moyer and 39 Milznez 2001) that " RBC cholinesterase activity was statistically similar to placebo at all time 40 points in the 0.005, 0.015, 0.03, and 0.06 mg/ kg dose groups" was not justified as the study was 41 not designed to test for statistical similarity. 42 43 ( c) The Agency concluded that " there is no indication from the available rat data for 44 either oxamyl or methomyl to suggest a sex difference." The animal data were not supplied to 45 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 28 of 69 the Board and therefore the Board could not independently verify this conclusion. However the 1 Board was not uncomfortable with accepting EPA's conclusions on this matter. 2 3 The Agency also concluded that " Given the similarity between brain and RBC ChE in 4 rats, the Agency is not aware of any biological or physiological reason that human brain ChE 5 would be more sensitive than the rat brain." Again without seeing the animal data, the Board 6 could not verify this conclusion about rats, but the conclusion about human brain and RBC 7 cholinesterase is reasonable scientifically 8 9 HSRB Consensus and Rationale 10 11 Although the Board had some questions about the Agency's conclusions regarding lack 12 of sex difference and of the difference between brain and RBC ChE based on only one species, 13 the Board supported the Agency's conclusion that there were no study deficiencies identified that 14 would have affected the outcome of conclusions of this study. 15 16 Considering the high quality of the design and the conduct of the study, the Board agreed 17 that this intentional human dosing study of oxamyl was sufficiently robust to be used for 18 reducing the 10x inter species ( i. e. animal to human) uncertainty factor in the cumulative risk 19 assessment for the N methyl carbamates. 20 21 3.2. Ethical considerations 22 23 Charge to the Board 24 25 a. The Agency requests that the Board provide comment on the following: 26 27   Whether inclusion in the protocol submitted to the ethics committee of a factually 28 inaccurate statement regarding unavailability of data on accidental or incidental 29 exposure to oxamyl should be considered significantly deficient relative to the 30 ethical standards prevailing when the study was conducted; 31 32   Whether the absence from the protocol of any discussion of the potential risks to 33 subjects or benefits to society of conducting the proposed research ( as required by 34 the Declaration of Helsinki, Principle # 5) should be considered significantly 35 deficient relative to the ethical standards prevailing when the study was 36 conducted; and 37 38 b. The Agency asks that the Board provide comment on the following, taking into 39 account all that is known about the ethical conduct of [ this/ each] study: 40 41   OPP's conclusion that there is not clear and convincing evidence that the conduct 42 of the research was fundamentally unethical. 43 44 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 29 of 69   Whether there is clear and convincing evidence that the conduct of the study was 1 significantly deficient relative to the ethical standards prevailing when the study 2 was conducted. 3 4 Board Response to the Charge 5 6 Study Overview 7 8 The study was performed by a contract research organization, Inveresk Clinical Research 9 of Edinburgh, Scotland and sponsored by the Haskell Laboratory for Toxicology and Industrial 10 Medicine, E. I. du Pont de Nemours and Company, of Newark, Delaware. The protocol 11 documents specifically state that the research was conducted in compliance with the Principles of 12 Good Clinical Practice promulgated by the International Conference on Harmonization 13 ( CMPM/ ICH/ 135/ 95), and the 1996 version of the Declaration of Helsinki. Comments provided 14 by du Pont Crop Protection ( Dupont 2006), also assert compliance with the 1996 version of the 15 Declaration of Helsinki. 16 17 Critique of Study 18 19 The Board concurred with the factual observations of the strengths and weaknesses of the 20 study, as detailed in Carley 2006b). However, further comments are warranted regarding: 1) 21 whether the exclusion of data on accidental or incidental exposure to oxamyl meets the legal 22 definition of research misconduct prevalent at the time the study was conducted; 2) whether the 23 escalating dose oral exposure protocol used was designed to minimize risks to study 24 participants; and 3) whether the documentation and process of study subject enrollment was 25 sufficient to meet prevailing standards of voluntary informed consent. 26 27 1) Exclusion of Available Data on Accidental or Incidental Exposure 28 29 Page 15 of the original study protocol provided to the independent ethics review 30 committee established by Inveresk Clinical Research included the following statement: " No data 31 on accidental or incidental exposure of people to oxamyl is available" ( McFarlane and Freestone 32 1999). This statement is false. Prior to the start of the Randomised Double Blind Ascending Oral 33 Dose Study with Oxamyl in late 1998, Du Pont submitted eleven adverse event reports involving 34 exposure of humans to oxamyl for inclusion in the EPA's Office of Pesticide Programs ( OPP) 35 Incident Data System ( IDS). Between 1982 and 1994, the State of California Department of 36 Food and Agriculture's Pesticide Illness Surveillance Program recorded an additional 61 37 incidents, including 30 cases involving occupational exposure to oxamyl alone. In comments 38 provided to the EPA on March 31, 2006, Du Pont Crop Protection stated that " most of the 39 accidents or incidents reported in these databases lack confirmation of the product involved or 40 the amount of product involved in the exposure. Also, some reports lack information on 41 symptoms or details of the exposure. Therefore, none of this data was useful for designing the 42 study." ( DuPont 2006). This latter statement may be true, but the discounting of the relevance of 43 adverse event data from accidental or incidental exposures is ill advised and borders on the 44 unprofessional. This may have been an appropriate decision but the data, evaluation and 45 assessment should have been discussed in the protocol and the consent document. Furthermore, 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 30 of 69 the statement in the original study protocol remains untrue. The question before the Board, 1 therefore, was whether or not this misstatement met the legal definition of research misconduct. 2 3 The Agency has accepted the federal policy on research misconduct ( Classification No.: 4 3120.5; Approval Date: 03/ 18/ 2003; Review Date: 03/ 18/ 2006), as defined in 42 CFR 93.103: 5 6 Research misconduct means fabrication, falsification, or plagiarism in proposing, 7 performing, or reviewing research, or in reporting research results. 8 9 ( a) Fabrication is making up data or results and recording or reporting them. 10 11 ( b) Falsification is manipulating research materials, equipment, or processes, or changing 12 or omitting data or results such that the research is not accurately represented in the research 13 record. 14 15 ( c) Plagiarism is the appropriation of another person's ideas, processes, results, or words 16 without giving appropriate credit. 17 18 ( d) Research misconduct does not include honest error or differences of opinion. 19 Although the failure to include the incidental exposure data may meet this definition of 20 falsification, a finding of research misconduct made under 42 CFR 93.104 requires that: 1) there 21 be a significant departure from accepted practices of the relevant research community; 2) the 22 misconduct be committed intentionally, knowingly, or recklessly; and 3) the allegation be proven 23 by a preponderance of the evidence. Under the standards outlined in 42 CFR 93.103 104, given 24 the limitations in the studies cited by Du Pont, and lack of evidence that the absence of 25 information was intentional, knowing, or reckless, it is the opinion of the Board that this 26 omission does not meet the evidentiary threshold necessary for a finding of research misconduct 27 based on falsification. 28 29 2) Minimization of Risks to Study Participants 30 31 Reasonable exposure levels appear to have been chosen using available animal data. 32 Although human exposure data from the EPA OPP IDS and California's Pesticide Illness 33 Surveillance Program also may have been useful in designing the study so as to minimize risk to 34 research subjects, there is no clear and convincing evidence of willful or blatant disregard for 35 participant safety. Adherence to a strict dose escalation protocol, with a stopping rule based on 36 40% inhibition of RBC cholinesterase, reduced the likelihood that study subjects would 37 experience unanticipated adverse clinical events. Based on the knowledge available to 38 investigators at the time the study was conducted, this double blind ascending dose oral 39 exposure trial appears to have been designed to adequately minimize potential risks to study 40 participants. 41 42 3) Voluntary Informed Consent 43 44 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 31 of 69 It is clear that the written documentation for informed consent failed to meet the 1 standards outlined in Principles of Good Clinical Practice ( CMPM/ ICH/ 135/ 95), and the 1996 2 version of the Declaration of Helsinki. 3 4 The information sheet that was given to research subjects as an appendix to the signed 5 informed consent document lacked adequate discussion of the potential risks of participating in 6 the research study. Several examples of what Board members consider inadequate discussion of 7 potential risks are illustrated below: 8 9 Data on incidents involving accidental oxamyl exposure were available through both the 10 EPA's OPP IDS and the California Pesticide Illness Surveillance Program, as previously 11 discussed. These data included reports of adverse clinical events, including detailed reports of 12 systemic effects, eye effects and dermatological effects resulting from exposure to oxamyl. 13 These data were not provided to study participants. 14 15 The background information provided to study participants enrolling in both the original 16 and amended study included the statement that " results from this study would further confirm 17 that the use of oxamyl for crop protection would not pose an unreasonable risk to human health." 18 ( McFarlane and Freestone 1999). Although the informed consent document clearly states that 19 this is the first study involving intentional exposure of human subjects, particularly given the 20 incident data discussed above, this statement seems to be written in order to downplay any fears 21 that study participants might have with regard to participation. 22 23 Study participants enrolling in the amended study, designed to look for clinical or 24 biological outcomes associated with exposure to 0.15 mg/ kg body weight of oxamyl, were 25 informed that " no significant side effects [ were] noticed in any of [ the previous lower] dose 26 levels." ( McFarlane and Freestone 1999). Previous trial participants receiving 0.09 mg/ kg body 27 weight of oxamyl exhibited 4 24% and 12 22% RBC and plasma cholinesterase inhibition. The 28 study investigators concluded that these observed levels of cholinesterase inhibition did not 29 qualify as adverse events because of the variability in the data, but they also state that the level of 30 inhibition observed in study participants receiving 0.09 mg/ kg body weight of oxamyl achieved 31 statistical significance. Because the consent form described cholinesterase inhibition as a side 32 effect of oxamyl exposure, it can be argued that this information should have been provided to 33 subjects participating in the 0.15 mg/ kg dose study. The information sheet provided to 34 volunteers, however, specifically describes cholinesterase inhibition as one of the side effects of 35 oxamyl exposure. 36 37 The letter to the subjects' general practitioner contained no information about the actual 38 study and the product being administered other than the name of the compound. It thus 39 minimized the extent to which the opinion of the general practitioner could protect the research 40 subject. 41 42 In hindsight, it is clear that the documentation provided failed to rigorously meet the 43 standards of voluntary informed consent applicable to studies conducted in Scotland in 1998 44 1999. However, there was no clear and convincing evidence that these deficiencies knowingly 45 and seriously impaired the informed consent process. 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 32 of 69 1 HSRB Consensus and Rationale 2 3 The Board concluded that: 4 5 a) The oxamyl human toxicity study failed to fully meet the specific ethical standards 6 prevalent at the time the research was conducted. 7 b) There was no clear or convincing evidence that the research was fundamentally 8 unethical intended to seriously harm participants or that informed consent was not obtained. 9 c) There was no clear and convincing evidence of significant deficiencies in the ethical 10 procedures that could have resulted in serious harm ( based on the knowledge available at the 11 time the study was conducted) nor that information provided to participants seriously impaired 12 their informed consent. 13 14 Although the study failed to meet the specific ethical standards prevalent at the time the 15 research was conducted, including those cited in the study protocol itself, the Board concurred 16 with the assessment of the Agency that there was no clear and convincing evidence that the 17 conduct of the research was fundamentally unethical in that it was not intended to harm subjects 18 nor failed entirely to obtain informed consent. 19 20 It is believed that potential risks to study participants were inadequately addressed in the 21 documents provided to research participants at enrollment, thus impairing the informed consent 22 process. Nevertheless, the Board believed that there was no clear and convincing evidence to 23 suggest that the study investigators failed to obtain voluntary informed consent from all research 24 participants. 25 26 The Board was of the opinion that the deficiencies noted above are unlikely to have 27 resulted in serious harm to study participants, based on the knowledge available to the 28 investigators at the time. 29 30 The Board concluded that there is no obvious reason why the Agency cannot rely on the 31 results of this study, as appropriate under current pesticide laws, given the absence of clear and 32 convincing evidence that the research was fundamentally unethical or significantly deficient 33 relative to the ethical standards prevailing when the study was conducted. 34 35 4. Azinphos Methyl 36 37 Charge to the Board 38 39 Azinphos methyl ( AZM) is an organophosphate pesticide ( OP). Consistent with other 40 OPs, AZM elicits neurotoxicity through the inhibition of the enzyme, acetylcholinesterase, via 41 phosphorylation of the active site. At sufficiently high doses, exposure to AZM can lead to a 42 variety of clinical signs. The Agency is developing an assessment to estimate risk to workers 43 from exposure to AZM. In addition, AZM is a member of the OP common mechanism group 44 and is thus included in the cumulative risk assessment for the OPs. 45 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 33 of 69 Scientific considerations 1 2 The Agency's WOE document and DER for AZM describe the study design and results 3 of the AZM repeat dose, oral, human toxicity study. The WOE document also discusses the 4 Agency's conclusions regarding the usefulness of the human study in the worker risk assessment 5 and in the cumulative risk assessment for the OPs. For AZM, the Agency has concluded that the 6 human toxicity study is appropriate for developing a point of departure for extrapolation of risk 7 to workers exposed to AZM via the dermal and inhalation routes. For the cumulative risk 8 assessment, the Agency has determined that because no cholinesterase inhibition was seen in the 9 human toxicity study, it is not possible to evaluate whether steady state had been reached in 10 humans at 28 days of exposure. Thus, the Agency has concluded that the AZM repeat dose, oral, 11 toxicity study is not sufficiently robust for informing the inter species factor in the cumulative 12 risk assessment of the OPs. 13 14 Please comment on the scientific evidence that supports the conclusions for: 15 16 a. the use of the human toxicity study to develop a point of departure for extrapolation of 17 risk to workers in the worker risk assessment and 18 19 b. the determination that the human toxicity study cannot be used to inform the inter 20 species factor in the cumulative risk assessment. 21 22 Board Response to the Charge 23 24 Brief Overview of the Study 25 26 In a repeat dose oral human toxicity study, conducted in 1999 by the Inveresk Laboratory 27 ( McFarlane & Freestone, 1999b), four males were dosed with a placebo ( lactose) and eight 28 males were dosed with 0.25 mg/ kg/ day of AZM for 28 consecutive days. The sole dose of 0.25 29 mg/ kg was chosen based on the results of subchronic rat study and an earlier study with human 30 subjects that indicated this would be a NOAEL for inhibition of plasma and RBC cholinesterase. 31 Previous human studies were conducted in 1972 and 1974 and regulatory agencies reluctant to 32 rely on the data " due to deficiencies in documentation, specifically the availability of raw data 33 and the lack of GLP compliance." 34 35 Dosing was orally by capsule and the subjects were housed in a clinic throughout the 36 dosing period. The subjects were monitored for reactions and samples of blood were taken at 37 predose ( eight times) and daily throughout the dosing period. On some days, a blood sample was 38 taken both prior to administering the dose and four hours later. Both plasma cholinesterase ( ChE) 39 and RBC acetylcholinesterase ( AChE) were measured. In addition, hematology, clinical 40 chemistry, and urinalysis were evaluated as well as blood pressure and electrocardiogram ( ECG). 41 There were no reactions to treatment and blood pressure and ECG were not affected. Eight of 42 the 12 volunteers were noted to have symptoms of a viral infection and some were treated with 43 paracetamol. The postdosing group mean data were compared three ways in an effort to see if 44 AZM caused inhibition of either plasma ChE or RBC AChE. None of the comparisons indicated 45 inhibition of either measure. Also, samples taken four hours after dosing did not indicate 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 34 of 69 inhibition. It was concluded that a dose of 0.25 mg/ kg/ day for 28 days did not result in either 1 clinical symptoms or inhibition of either plasma ChE or RBC AChE in human males, so that this 2 dose could be considered a NOAEL. 3 4 Critique of the Study 5 6 One strength of the study was its design as a randomized, double blind study including a 7 placebo group. The clear inclusion and exclusion criteria for subject selection appeared to have 8 been chosen to reduce the likelihood of confounding. The subjects were in good health, were not 9 using prescription or illicit drugs, were non smokers, and were not agricultural workers or 10 pesticide applicators who might have experienced OP exposure within the preceding month. 11 Subjects resided in the clinic from the day before study start to 24 hours after the last dose was 12 administered, received a standardized diet, and were under medical and nursing supervision 13 throughout. 14 15 The assessment of critical outcomes was comprehensive, involving a general clinical 16 exam, measurement of systolic and diastolic blood pressure, the administration of an ECG, and 17 various blood and urine studies. Multiple measurements were made of plasma ChE and RBC 18 AChE, namely on 8 pre dose days then daily 4 hours after dosing. Moreover, on 12 post dosing 19 days, two blood samples were taken, one before the dose and one 4 hours afterwards. 20 21 Possible group differences in plasma ChE and RBC AChE were evaluated in several 22 ways, both in grouped and individual data) 23 a) Comparison of the day to day ChE activities within the AZM group to the same day 24 values in the control group 25 b) Comparison of same day predosing ChE activities with inhibition 4 hours after dosing 26 c) Comparison of the day to day ChE activities values within the AZM group to 27 predosing baseline value. 28 29 Several weaknesses were identified in the study. First, the subjects were all male, 30 however there have been no consistent sex differences in sensitivity observed in animal models. 31 The sample size was not large, involving only 12 adult males ( four in the placebo group, eight in 32 the AZM group), raising a concern that the statistical power might have been inadequate to 33 detect a modest degree of inhibition in the AZM group. 34 35 The standard deviations of the ChE levels were large, as great as 20% for plasma ChE 36 and 15% for RBC ChE, reducing confidence in the consistency of these endpoints. 37 38 A variety of symptoms were reported over the course of the study, with the majority 39 occurring in subjects administered AZM. The conclusion drawn in the Agency's Data 40 Evaluation Record ( Doherty 1999) that none of the symptoms likely reflected effects of AZM is 41 not verifiable, nor is it credible. 42 43 Certain aspects of the statistical analyses are problematic. For the analyses of percentage 44 change each day from baseline ( predosing) plasma and RBC cholinesterase activity in the 45 subjects receiving AZM, the choice of repeated measures ANOVA as the statistical approach 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 35 of 69 was appropriate given the structure of the data. This would have allowed each subject to be used 1 as his own control, but this does not appear to be the way the analyses were done. The pooled 2 baseline RBC and plasma ChE values were used as covariates rather than individual subject 3 values. This essentially ignores the value of the individual levels in increasing statistical power 4 by taking account of pre dosing inter individual differences in ChE levels. The other two types 5 of analyses conducted were not limited by this deficiency, but the analysis of within subject 6 change from baseline is potentially the most sensitive analysis in this study. 7 8 In proposing to use the results of this study for dermal and inhalation risk assessment, the 9 Agency applied a dermal absorption factor of 42% based on a rat study and an inhalation 10 absorption factor of 100%. Bayer Crop Science commented that data are available supporting a 11 lower estimate of human dermal absorption ( 21.5%) but these data were not available for 12 evaluation by the Board. Another concern was whether the effects of a single oral dose 13 administered daily provide a reasonable estimate of the effects of the continuous dosing via all 14 routes of exposure that would be experienced by agricultural workers. 15 16 The major concern regarding this human study was it was designed to demonstrate a 17 NOAEL. No LOAEL could be established to validate the NOAEL because only one dose was 18 used. The fact that inhibition was not observed at the single dose used raised concern about 19 sensitivity of the methods used. The Agency was reassured by the laboratory's observation of 20 plasma and RBC cholinesterase inhibition in other studies of carbamates, including aldicarb, 21 methomyl, and oxamyl. Nevertheless, it is not possible to be sure if the absence of an effect in 22 this study was because the methods used were, for some reason, not up to the standards of those 23 used in the other studies, and therefore less sensitive. 24 25 HSRB Consensus and Rationale 26 27 The Board concluded that: 28 29 a. Data from the 28 day repeat oral dose study of AZM should not be used in developing 30 a point of departure for extrapolation of risk to workers exposed to AZM via the dermal and 31 inhalation routes. 32 33 b. Data from the 28 day repeat oral dose study of AZM cannot be used to inform the 34 inter species factor in the cumulative risk assessment. 35 36 Despite the study's strengths, the HSRB had only limited confidence in the inferences 37 that can be drawn from this study, in large part because only one AZM dose was used. In order 38 to use these data, one must assume that the lab would have detected inhibition had it been present 39 at the single dose administered. The conclusions drawn could be correct, but this cannot be 40 determined with certainty. The conclusion that this study is not appropriate for developing a 41 point of departure for either the worker risk assessment or the cumulative risk assessment is 42 consistent with the recommendation of the NAS committee ( NAS 2001) that the Agency should 43 not consider data from NOEL only studies. 44 45 Ethical considerations 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 36 of 69 1 Charge to the Board 2 3 a. The Agency requests that the Board provide comment on the following: 4 5 Whether the informed consent materials   which refer to " the company" and " supervising 6 doctor" without further identification, and contain no discussion of who would benefit from the 7 research   should be considered significantly deficient relative to the ethical standards prevailing 8 when the study was conducted; and, 9 10 Whether the absence from the protocol of any discussion of the potential risks to subjects 11 or benefits to society of conducting the proposed research ( as required by the 1996 Declaration 12 of Helsinki, Principle # 5, with which the research asserted compliance) should be considered 13 significantly deficient relative to the ethical standards prevailing when the study was conducted; 14 and 15 16 b. The Agency asks that the Board provide comment on the following, taking into 17 account all that is known about the ethical conduct of [ this/ each] study: 18 19 OPP's conclusion that there is not clear and convincing evidence that the conduct of the 20 research was fundamentally unethical. 21 22 Whether there is clear and convincing evidence that the conduct of the study was 23 significantly deficient relative to the ethical standards prevailing when the study was conducted. 24 25 Brief Overview of the Study 26 27 As noted above, the study was conducted in 1999 by a contract research organization, 28 Inveresk Clinical Research of Edinburgh, Scotland, at its Elphinstone Research Centre in 29 Tranent. The study sponsor was the Bayer Corporation, Agricultural Division, of Stilwell, 30 Kansas. The protocol documents specifically state that the research was conducted in compliance 31 with the guideline for Good Clinical Practice, and in accordance with the guidelines of the 32 Declaration of Helsinki, 1964, as amended through 1996. 33 34 Critique of Study 35 36 The Board concurred with the factual observations of the strengths and weaknesses of the 37 study, as detailed in Carley ( 2006d). However, further comments are warranted with regard to 38 whether the documentation and process of study subject enrollment was sufficient to meet 39 prevailing standards of voluntary informed consent. 40 41 It is clear that the written documentation for informed consent failed to meet fully the 42 standards contained in the 1996 version of the Declaration of Helsinki. 43 44 In particular, the information sheet that was given to research subjects as an appendix to 45 the signed informed consent document contained the following statement under the subheading 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 37 of 69 Aim: " The aim of the study is to establish a recommended daily intake following long term 1 dietary exposure and to determine the most appropriate way of assessing workers exposure." By 2 using the phrase " recommended daily intake," this statement would appear to suggest that it is 3 actually desirable to consume a daily minimum amount of the study compound. This sentence is 4 thus misleading. However, other portions of the information sheet made it clear that the study 5 compound was a pesticide, and that consuming it is not good for people, thus leading the Board 6 to conclude that this one sentence by itself does not provide clear and convincing evidence that 7 the consent process as a whole was significantly deficient relative to the ethical standards 8 prevailing at the time the study was conducted. 9 10 HSRB Consensus and Rationale 11 12 The Board concluded that: 13 14 a) The azinphos methyl human toxicity study failed to fully meet the specific ethical 15 standards prevalent at the time the research was conducted. 16 17 b) There was no clear and convincing evidence that the research was fundamentally 18 unethical intended to seriously harm participants or that informed consent was not obtained. 19 20 c) There was no clear and convincing evidence of significant deficiencies in the ethical 21 procedures that could have resulted in serious harm ( based on the knowledge available at the 22 time the study was conducted) nor that information provided to participants seriously impaired 23 their informed consent. 24 25 The Board determined that the informed consent materials   which refer to " the 26 company" and " supervising doctor", without further identification, and which contain no 27 discussion of who would benefit from the research   did not fully comply with the ethical 28 standards at the time. However, the Board did not find clear and convincing evidence that the 29 level of non compliance should be considered significantly deficient relative to those then 30 prevailing ethical standards. With regard to the use of the terms " the company" and " supervising 31 doctor," this type of language, though highly undesirable, is unfortunately not uncommon, even 32 at the present. With regard to the benefit issue, the Board determined that the consent materials, 33 while far from ideal, did in fact somewhat address this issue. They indicate, for example, that the 34 study may lead to further information about the risk that this compound poses to workers and 35 consumers. 36 37 The Board determined that the absence from the protocol of discussion of the potential 38 risks to subjects or benefits to society of conducting the proposed research ( as required by the 39 Declaration of Helsinki, Principle # 5) should not be considered significantly deficient relative to 40 the ethical standards prevailing when the study was conducted. The potential risks and benefits 41 are described in the research subject information sheet/ informed consent document. While not in 42 the protocol itself, they are nonetheless included in the materials provided to the ethics 43 committee ( and to the potential research subject). Thus the ethics committee and the potential 44 research subjects had the opportunity to weigh the risks and benefits. 45 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 38 of 69 The Board concluded that there was no obvious reason why the Agency cannot rely on 1 the results of this study, as appropriate under current pesticide laws, given the absence of clear 2 and convincing evidence that the research was fundamentally unethical or significantly deficient 3 relative to the ethical standards prevailing when the study was conducted. 4 5 5. DDVP 6 7 Charge to the Board 8 9 Like AZM, DDVP is an organophosphate pesticide ( OP) which elicits neurotoxicity 10 through the inhibition of acetylcholinesterase, via phosphorylation of the active site. The 11 Agency is conducting an aggregate ( single chemical, multi route, multi duration) risk assessment 12 of DDVP. In addition, DDVP is a member of the OP common mechanism group and is thus 13 included in the cumulative ( multi chemical, multi route) risk assessment for the OPs. 14 15 5.1. Scientific considerations 16 17 a. The Agency's WOE document and DER for DDVP describe the study design and results 18 of the DDVP repeat dose oral human study. The WOE document also discusses the Agency's 19 conclusions regarding the usefulness of this study in the aggregate risk assessment and in the 20 cumulative risk assessment for the OPs. For the single chemical risk assessment, the Agency has 21 concluded that the human study is sufficiently robust for developing a point of departure for 22 estimating dermal, incidental oral, and inhalation risk from exposure to DDVP in the single 23 chemical risk assessment. For the cumulative risk assessment, the Agency has determined that 24 results of the DDVP multi dose human toxicity study do not support reducing the default 10X 25 inter species factor in the cumulative risk assessment of the OPs. 26 27 Please comment on the scientific evidence that supports the conclusions for: 28 29 a) the Agency's conclusions for use of the human study for developing a point of 30 departure for estimating risk in the single chemical, aggregate risk assessment and 31 32 b) the Agency's determination that the human study cannot be used to reduce the 33 interspecies factor in the cumulative risk assessment. 34 35 Board Response to the Charge 36 37 Study Overview 38 39 A single blind, randomized placebo controlled oral study was conducted with DDVP in 40 which six healthy male volunteers were administered a daily dose of DDVP at approximately 41 0.1mg/ kg/ day and three volunteers were administered a placebo ( Gledhill 1997). DDVP was 42 dissolved in corn oil and administered daily in a gelatin capsule to fasted subjects ( subjects did 43 not eat after midnight prior to each morning dose) for a total of 21 days. Blood samples were 44 collected for analysis of red blood cell ( RBC) cholinesterase activity. For each subject, the 45 baseline level of RBC cholinesterase activity was established from seven pre dose samples, and a 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 39 of 69 single blood sample was collected approximately 24 hours after dosing ( just prior to dosing on 1 the following day) on days 1, 2, 4, 7, 9, 11, 14, 16 and 18 of the study. No blood samples were 2 collected on the last three days of the study ( days 19 21). After cessation of dosing, subjects 3 returned approximately 1 week later ( on or about day 25) for a follow up analysis of RBC 4 cholinesterase activity. The criterion for withdrawal from the study was a inhibition of RBC 5 cholinesterase activity of 20% or more on a given day's analysis followed by a further decrease 6 in activity in the next successive sample. A linear mixed model was fitted to the cholinesterase 7 activity and significant differences were found within subjects and also between groups 8 ( treatment versus placebo subjects) for pre dose and post dose cholinesterase activity. 9 10 Critique of Study 11 12 The scientific validity of the repeated dose study for DDVP was determined by the 13 relative strengths and weakness of the design and conduct of the study. 14 15 The strengths of the study were considered to be as follows: 16   The repeat dosing paradigm affords the opportunity to more critically evaluate the 17 sustained nature of RBC cholinesterase inhibition associated with OPs. 18   The analysis of RBC cholinesterase activity is fairly robust, given the assessment of 19 predose baseline values on 7 separate occasions. The baseline values were fairly 20 consistent both within and between subjects ( 16,000   21,000 IU). 21   A low, but statistically significant effect was observed in RBC cholinesterase inhibition. 22 23 There are, however, numerous weaknesses of the study that are relevant to the utility of these 24 data in establishing a point of departure for use in single chemical, aggregate and cumulative risk 25 assessments for DDVP. These weaknesses included: 26 27   A single dosage was used, preventing establishment of dose response relationships. 28 29   The sample size was small and included only male volunteers. In this regard, it is not 30 clear whether the study was properly powered, given that no sample size calculations 31 seem to have been used in order to arrive at the number of volunteers used in the study. 32 33   RBC cholinesterase inhibition was determined only at 24 hours post dosing. Although 34 organophosphates are known to show prolonged inhibition of cholinesterase activity, it 35 was not clear whether the peak of RBC cholinesterase was established. 36 37   The sponsor did not include the analysis of plasma cholinesterase measurements in the 38 study. The reason for this omission is not clear, and it was noted that virtually every 39 toxicological and exposure assessment study that has focused on cholinesterase inhibition 40 due to organphosphorus insecticides includes measurement of both plasma ( serum) 41 cholinesterase and RBC cholinesterase. The combination of these measurements has been 42 used routinely for risk assessments by numerous regulatory agencies, including the EPA, 43 as well as by clinicians who diagnose pesticide poisonings. Current worker surveillance 44 programs in some states ( California and Washington) require measurement of both 45 plasma and erythrocyte cholinesterase, and both measurements are used in combination 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 40 of 69 to initiate workplace investigations and remove workers from pesticide related job 1 activities until enzyme activities return to baseline. 2 3   Sampling was incomplete because the last day of analysis of enzyme inhibition was day 4 18 but dosing continued until day 21. It is not clear whether any additional decrease in 5 enzyme activity would have been observed on days 19 21. In the sampling schedule 6 outlined above, the maximum time gap between samples during the dosing period ( days 7 1 21) was three days ( between days 11 and 14). Logically then, an additional sample 8 would have been taken on day 21 or 22, the time in the study when maximum effects 9 might be observed. In this regard, two subjects presented with inhibition values of 22 10 and 23%, exceeding the 20% cut off for subsequent analysis and consideration for 11 withdrawal on day 18 of the study. The lack of additional measurements in these subjects 12 during the remainder of the dosing period is scientifically inadequate and ethically 13 troublesome. 14 15   It is neither clear nor convincing that steady state inhibition of RBC cholinesterase 16 activity was achieved in the study. The data suggest that the level of enzyme inhibition 17 was still increasing, particularly in the time period from 11 18 days, with a difference of 18 16% in activity on Day 18 compared to pre dose values. Dosing was stopped on Day 21, 19 with no further analysis until approximately 4 days after cessation of dosing. Although 20 the difference in activity between day 18 post dose and the average of the pre dose 21 measurements was deemed to be " not biologically significant" by the study sponsor, the 22 possibility that RBC cholinesterase inhibition may have reached or exceeded 20% by the 23 end of the study cannot be excluded. 24 25   There was a lack of appropriate follow up in all subjects in this study. As noted earlier, 26 two subjects presented with inhibition values of 22 and 23%, exceeding the 20% cut off 27 for subsequent analysis and consideration for withdrawal on Day 18. However, there 28 were no additional analyses of RBC cholinesterase activity in these individuals on days 29 19 21 of dosing. Additionally, a blood sample was collected within 4 to 10 days 30 following completion of the 21 day dosing period for all subjects. The mean inhibition 31 of the enzyme observed at seven days following cessation of dosing was 83%, suggesting 32 little if any recovery of enzyme at this time, but these data were not analyzed for 33 statistical significance. Furthermore, one subject ended his participation in the study ( day 34 25) with RBC cholinesterase inhibition greater than 20%. In all cases, there was no 35 additional follow up of the subjects after the single post dose analysis of cholinesterase 36 activity. 37 38 HSRB Consensus and Rationale 39 40 The Board concluded that: 41 42 a) The DDVP repeat dose human toxicity study was sufficiently robust for developing a point 43 of departure for estimating dermal, incidental oral, and inhalation risk from exposure to DDVP in 44 the single chemical risk assessment. 45 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 41 of 69 In specific response to the Agency's charge question, the HSRB concluded that there are 1 numerous technical limitations to the data obtained in the study. In addition to the weaknesses of 2 the study design and execution outlined above, the HSRB concluded that the evaluation of only a 3 single dose level of DDVP along with the omission of plasma cholinesterase measurements 4 greatly limit value of this study in terms of producing new knowledge that can be used in the 5 regulatory process and in medical diagnosis. Furthermore, study investigators have an obligation 6 to design a study that provides appropriate oversight of subjects until indications of the effects of 7 the administered dose are no longer present, and the Board considered that the design of this 8 study which continued intentional dosing without collection of blood samples for cholinesterase 9 analysis was not defensible scientifically. Following considerable discussion, the HSRB 10 concluded that the dosage evaluated in the repeat dosing human study can be used as a LOAEL 11 for the single chemical aggregate risk assessment for DDVP. Although a study using a single 12 dose level is not ideal for establishing a LOAEL, there was general consensus that RBC 13 cholinesterase is a well characterized endpoint for compounds that inhibit acetylcholinesterase 14 activity and therefore, because the decreased activity in RBC cholinesterase activity observed in 15 this study was at or near the limit of what could be distinguished from baseline values, it was 16 unlikely that a lower dose would produce a measurable effect in RBC cholinesterase activity. 17 However, the HSRB strongly recommended that, in most cases, studies in human subjects 18 designed to define the NOAEL or LOAEL for a compound should include more than a single 19 dose level. 20 21 b) Results of the DDVP repeat dose human toxicity study should not be used to support 22 reducing the default 10X inter species factor in the cumulative risk assessment of the 23 organophosphate pesticides. 24 25 Although the data obtained in this study can be used to establish a point of departure for the 26 single chemical aggregate risk assessment, the consensus of the HSRB was that the scientific 27 limitations of the study design do not justify its use in the cumulative risk assessment. In 28 particular, the lack of sample collection through to the completion of dosing, the lack of a clear 29 demonstration that steady state inhibition of RBC cholinesterase inhibition had been achieved, 30 and the lack of any dose response data limit the overall utility of the human data. Accordingly, 31 the HSRB recommended that the default interspecies uncertainty factor should be applied for the 32 cumulative risk assessment for DDVP. 33 34 Charge to the Board 35 36 b. The Agency has concluded that other human studies made available to the Board do not 37 provide sufficient scientifically sound information to warrant any reduction in the 10X inter 38 species uncertainty factor used to derive reference dose values for DDVP based on animal 39 toxicity endpoints. 40 41 Please comment on the scientific evidence that supports these conclusions. 42 43 Board Response to the Charge 44 45 HSRB Consensus and Rationale 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 42 of 69 1 The HSRB concluded that the other DDVP human toxicity studies available for the 2 Board's consideration should not be used for determining a reduction in the 10X uncertainty 3 factor to derive reference dose values for DDVP based on animal toxicity endpoints. 4 5 The Board's deliberations for this question focused on the acceptability or not of these 6 human studies for the Agency's intended purposes, rather than on the evaluation of the 7 magnitude of the interspecies uncertainty factor. Since the results of animal studies or 8 benchmark dose modeling were not provided to the Board, the review was restricted to the 9 evaluation of the acceptability of the human studies for the intended purposes. The Board 10 concluded these other human studies did not warrant use in determining the inter species 11 uncertainty factor because of policy considerations ( e. g., exposures involving infants, children, 12 pregnant women) or due to uncertainty associated with the study itself ( e. g., inappropriate 13 duration, questionable route, exposure data of limited utility, lack of time course data on critical 14 effect). Board concerns regarding these other human studies are provided in Table 1. 15 16 While the studies reviewed are not useable either because they are unacceptable due to 17 policy considerations or due to uncertainty associated with the study itself, the Board also 18 suggested that the Agency should take a critical hard look at those studies that are ethically 19 acceptable ( e. g., the ones that did not include exposures of children and pregnant women), to the 20 extent to which they can be informative about the magnitude of the interspecies uncertainty 21 factor. In making such evaluations, the Agency should consider taking into account: ( 1) 22 applicability of Haber's law, ( 2) nature of the toxic moiety, and ( 3) the relative importance of 23 interspecies differences in pharmacokinetics and sensitivity. Further, consideration should be 24 given to the manner in which the data from the different species are pooled and analyzed ( e. g., 25 continuous response versus maximal response). 26 27 28 29 30 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 43 of 69 1 Table 1. Evaluation of Specific DDVP Human Studies Provided to the HSRB 2 3 4 Reference Observations/ Concerns Glendhill Acute toxicity study Time course measurements of cholinesterase Cavagna ( 1970) Conducted in newborns Slomka ( 1981) Exposure through plastic bead ( polyvinyl resin) formulation; rate of absorption not known; data provided as ranges Cervoni ( 1969) Efficacy trial; measurement of cholinesterase inhibition not the main focus; no assurance of consistent methods of measurement of inhibition Leary ( 1974) Exposure via insecticide strips; possibility children were involved, but composition of families not given; inhalation route   caveat of dose calculations; no information on consent of subjects Smith ( 1972) Performed in altitude chamber; short term inhalation study; no information on consent of subjects Cavagna ( 1969) Exposure via insecticide strips included pregnant women, infants and children in a hospital Pena Chavarria ( 1969) Exposure of patients ( most of them anemic, treated for parasite infection); not designed to identify NOAEL or LOAEL Stein ( 1966) Insufficient information on the time course of cholinesterase inhibition; doses not quantitated Funckes ( 1963) Study population included children, some below six years; time course evaluation on inhibition insufficient Witter ( 1961) Simulated aircraft cabin; one hour exposures; time course information lacking Gratz ( 1963 Inadequate information to evaluate dose and NOAEL; inadequate information about how many people exposed at each concentration; single application based exposures 5 5.2. Ethical considerations 6 7 Charge to the Board 8 9 a. The Agency requests that the Board provide comment on the following: 10 11 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 44 of 69 Whether references to the test material as a drug and other statements that could indicate 1 the study constituted medical research, that appear in the materials used to obtain informed 2 consent should be considered significantly deficient relative to the ethical standards prevailing 3 when the study was conducted; 4 5 Whether the administration of the test material for three additional days without 6 monitoring subjects' cholinesterase levels following the detection of cholinesterase inhibition 7 greater than 20 % in some subjects should be considered significantly deficient relative to the 8 ethical standards prevailing when the study was conducted; and 9 10 Whether the lack of medical surveillance of subjects, following the termination of dosing, 11 to establish the subjects' cholinesterase levels returned to normal should be considered 12 significantly deficient relative to the ethical standards prevailing when the study was conducted; 13 and 14 15 b. The Agency asks that the Board provide comment on the following, taking into account all 16 that is known about the ethical conduct of the Gledhill repeated dose study: 17 18 OPP's conclusion that there is not clear and convincing evidence that the conduct of the 19 research was fundamentally unethical; and 20 21 Whether there is clear and convincing evidence that the conduct of the Gledhill repeat 22 dose study was significantly deficient relative to the ethical standards prevailing when the study 23 was conducted. 24 25 Board Response to the Charge 26 27 Brief Overview of the Study 28 29 As noted above, the study was conducted in 1996 by the Zeneca Central Toxicology 30 Laboratory, Cheshire, United Kingdom. The study sponsor was the Amvac Chemical 31 Corporation. The protocol documents specifically state that the research was conducted in 32 compliance with the guideline of the Declaration of Helsinki, 1964, as amended through 1989. 33 34 Critique of Study 35 36 The Board concurred with the Agency's factual observations of the strengths and 37 weaknesses of the study, as detailed in Carley ( 2006e). However, further comments are 38 warranted with regard to: 1) whether the provisions in the protocol for monitoring the subjects 39 adequately protected them, and 2) whether the documentation and process of study subject 40 enrollment was sufficient to meet prevailing standards of voluntary informed consent. 41 42 1. Monitoring of Subjects While Receiving Study Compound 43 44 The protocol permitted subjects to continue to be administered the study compound for 45 three days following the detection of RBC cholinesterase inhibition of greater than 20%. Given 46 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 45 of 69 that study subjects were not monitored during most of the study ( and were not full time residents 1 at the study laboratory), it was possible that they might develop clinically significant symptoms 2 during this period and no one would be available to detect that and to treat them. The Board 3 concluded that this aspect of the study design was inadequate. However, given that there had 4 been a prior similar study at triple the dose used in this study, and that study had demonstrated 5 minimal symptoms, the Board could not conclude that there was clear and convincing evidence 6 that this element of the study design could have resulted in serious harm based on the knowledge 7 available to the investigators at the time. 8 9 2. Monitoring of Subjects at End of Study 10 11 The protocol also provided that, at the end of the study, inhibition levels would not be 12 retested for periods as long as seven days even though some subjects had RBC cholinesterase 13 inhibition of greater than 20%. During this period of time, due to the long acting nature of the 14 study compound, subjects' cholinesterase inhibition levels could have been increasing, and thus 15 symptoms might have developed and, theoretically have become serious at a time when they 16 were being infrequently monitored. The Board determined that this aspect of the study design 17 was inadequate. However, as noted above, given that there had been a prior similar study at 18 triple the dose used in this study, and that study had demonstrated minimal symptoms, the Board 19 could not conclude that there was clear and convincing evidence that this element of the study 20 design could have resulted in serious harm based on the knowledge available to the investigators 21 at the time. 22 23 The inadequate monitoring during and at the end of the study undermines the scientific 24 value of the study. The fact that the research subjects were unsupervised during the study, and 25 the ascertainment of adverse event data was through self report, renders the data establishing a 26 NOAEL suspect. The risk of a " false negative" conclusion is quite high. In addition, the failure 27 to achieve a steady state in the inhibition of acetyl cholinesterase activity may render these data 28 inadequate as well, and of limited utility. This fact, also noted in the Board's scientific review, 29 can negatively influence the prospective benefit to risk ratio. 30 31 3. Informed Consent 32 33 It is clear that the written documentation for informed consent failed to fully meet the 34 standards outlined of the 1989 version of the Declaration of Helsinki. 35 36 In particular, the information sheet that was given to research subjects as an appendix to 37 the signed informed consent document under the subheading Possible Adverse Events noted that 38 if " any symptoms possibly due to enzyme blocking did occur, they could rapidly be reversed by 39 a specific antidote." Although that statement is a correct statement of the scientific issue, it failed 40 to take account the fact that this study did not involve subjects remaining under 24 hour 41 supervision. For most of the time, the subjects would not be under any supervision. Given that 42 enzyme inhibition could build up over time due to the long acting nature of the compound being 43 studied, symptoms could likely occur at a time when there was no one available to administer the 44 antidote. Telling the subjects that an antidote could rapidly reverse symptoms without explaining 45 Proposed Final Draft V. 1. Dated May 16, 2006; Do Not Cite or Quote 46 of 69 that there were circumstances in which that antidote would not be readily available was not an 1 adequate way to inform subjects of the true nature of this risk. 2 3 In hindsight, it is clear that the documentation provided fails to rigorously meet the 4 standards of voluntary informed consent applicable to studies conducted in the United Kingdom 5 in 1996. However, there was not clear and convincing evidence that these deficiencies 6 knowingly and seriously impaired the informed consent process. 7 8 HSRB Consensus and Rationale 9 10 The Board concluded that: 11 12 a) The DDVP repeat dose oral human toxicity study failed to fully meet the specific 13 ethical standards prevalent