6. (a) The Subcommittee's statement that significant quantities of drugs are excreted daily in the manure and urine of medicated animals is correct regarding certain compounds. For example, a steer receiving 10 mg of DES per day will excrete about 7 mg of estrogenic substances during the same period. However, in all fairness, it should be pointed out that an average dairy cow excretes approximately 30 mg of estrogenic substances per day in the urine and feces 1 and the pregnant mare excretes more than 100 mg of estrogen daily.' Manure from DES-fed steers spread on soil in the same manner as that from a dairy operation would result in less than one-third the estrogenic contamination of the latter.1 Therefore it does not appear that DES contamination of the soil and drinking water is a major problem. (b) There have been some limited studies made on the fate of DES in the soil and water. Unpublished results of the Eli Lilly Company show that DES is only sparingly soluble in water (12 ppm) and that in the presence of soil particles, over 97 percent of the DES was absorbed to the soil and no longer carried in the aqueous phase. A few studies have been designed to determine the stability of DES in the soil. It is known that estrogenic activity disappears rapidly under acid conditions.1 Experiments to determine whether DES-like compounds were taken up from the soil by plants and therefore constituted a hazard to man or animal yielded negative results; uptake of synthetic estrogens by plant roots was considered to be insufficient to be hazardous. 1 In addition to the above, it has been shown that DES has a rather short life span in the soil when exposed to ultraviolet rays, air and the effects of certain bacteria. Two genera of microorganisms have been shown to degrade DES 3 and it is likely there are others which share this capability. 7. The use of hormones as growth promotants has been banned in some countries. The Department does not feel that this practice is contradictory to its assessment of these substances for two reasons. The countries in which hormones are banned depend, almost exclusively, on an export market for their product. Many of the countries to which they ship will not accept meat raised with benefit of hormones-Italy and Sweden are examples. Therefore, to maintain these markets it has been necessary to guarantee that no hormones have been employed. In addition, none of the 21 countries mentioned practices intensive feeding of high energy diets such as is used in feed lots in the United States. In a pastureoriented production system, the growth-promoting hormones have little or no use. The answers we have given to your questions reflect our most current thinking on these matters. As new information in these areas becomes available, we shall incorporate into our programs. In all instances, the well-being of the consumer will be the major factor guiding our Departmental policies. Sincerely, 1 ARS Bulletin 44-224, Feb. 1971, p. 29. RICHARD LYNG, Goodman, L. S., and A. Bilman, The Pharmacological Basis of Therapeutics, 4th Ed. Macmillan Co., New York, 1970. 3 Zondek, B. and F. Sulman. Inactivation of estrone and diethylstilbestrol by microorganisms. Endocrin. 33:204, 1943. CHARLES C. EDWARDS, M.D., U.S. SENATE, Washington, D.C., May 1971. Commissioner of Food and Drugs, Public Health Service, Department of Health, Education, and Welfare, Rockville, Md., DEAR DR. EDWARDS: During the recent hearings on "Chemicals and the Future of Man" before the Subcommittee on Executive Reorganization and Government Research, five scientists were asked to outline the principal hazards to human health from chemicals in our food, drugs and environment. While our purpose in this phase of the inquiry was to concentrate on three potential chemical dangers-cancer, birth defects and gene damage-opening testimony very quickly extended to Federal regulation, particularly the role of the Food and Drug Administration. Claims were made in testimony that Federal scientific procedures, regulation and enforcement in several areas of food protection are inadequate and that the public faces increasing health threats as a result of these deficiencies. It therefore seems appropriate that we seek early clarification of the issues involving your agency. In order that this Subcommittee be completely informed, I would appreciate it greatly if you would reply to the attached questions as soon as possible. Thank you for your cooperation. ABE RIBICOFF, Chairman INQUIRIES SUBMITTED TO THE FDA BY THE SUBCOMMITTEE AND THE RESPONSES SUBMITTED FOR THE RECORD 1. Has the FDA promulgated any formal protocols for safety evaluation of food additives? If so, would you please list these and indicate which of these tests have been promulgated in the Federal Register? If there are no such formal guidelines, or if such protocols have not been promulgated in the Federal Register, would you please explain why not. When do you propose to institute such a requirement? A. Title 21, Code of Federal Regulations, section 121.6, is entitled "General Principles for Evaluating the Safety of Food Additives." This regulation is part of the food additive procedural regulations, promulgated March 28, 1959. Paragraph (a) states, among other things, that the Commissioner will be guided by the principles and procedures for the safety of food additives stated in current publications of the National Academy of Sciences-National Research Council. The Food and Drug Administration's Advisory Committee on Protocols for Safety Evaluations was appointed in 1967 to provide guidance to FDA in current thinking and the type of modern testing which should be employed. Reports of Panels on Cancer Testing and on Reproduction were submitted in 1969. Copies of these are provided for the record. Finally, FDA is currently drafting for publication in the Federal Register, a replacement of section 121.6 with a text entitled "General Principles and Procedures for Evaluating the Safety of Substances Added to Food." This is expected to set forth the toxicological testing that must be done where the maximum total daily expected intake of the food additive is in one of the following three ranges: A. Up to 0.0015 mg/kg, provided that no individual food contains more than 0.05 ppm. B. Up to 0.01 mg/kg, providing that no food contains more than 1 ppm and the safety factor is at least 1000. C. In excess of 0.01 mg/kg and the safety factor falls usually in the range of 100 or less. 2. In a recent address to the Society of Toxicology, Mr. William Ruckelshaus, Administrator of the Environmental Protection Agency, urged toxicologists to evaluate chemicals for mutagenic effects. Additionally, four expert Governmental committees have recommended mandatory mutagenicity testing prior to registration of drugs and food additives. (See attachment) Do you share this view? If yes, when will the Food and Drug Administration move to include mutagenicity testing as part of a required safety testing program for food additives, drugs, and pesticides? Would you propose establishment of such protocols in the Federal Register? If not, could you please state your reason for this. A. At the time that the Advisory Committee on Protocols to the FDA deliberated on the subject of reproductive hazards which included teratogenesis and mutagenesis, it was concluded that the only test that had relevance to mammalian species was the dominant lethal test. These deliberations were attended and seconded by a number of specialists in the field. On the basis of these deliberations, the paragraph entitled "Mutagenicity," on page 282 of the article appearing in Toxicology and Applied Pharmacology, vol. 16, 1970, was included. The main thrust of this section is that the multigeneration test as designed, would allow for input in terms of dominant lethal effects. Although there was a need to develop methods for mutagenicity evaluation, this was the only relevant type of study that could be recommended at that time. Since then, the host-mediated assay has been developed and applied in several instances. However, the interpretation of test results by the recently developed mammalian systems, in terms of human hazard, must await additional experience. The value of these tests may turn out to be greater as initial screens in industrial development programs, rather than as definitive bases for human safety. In brief, the Food and Drug Administration is, in principle, in favor of testing for mutagenicity and is conducting a considerable amount of this testing both within its facilities and on contract. It is reluctant to impose mandatory requirements along this line until further experience with existing test procedures leads to more confidence in the interpretation of the results. There is at present a very considerable diversity of opinion within the scientific community in this regard, and there is no single test or battery of tests on which it would be possible to obtain general agreement that a decision of mutagenicity for man could be based. This uncertainty extends beyond the usual species specificity problem in that the test species include microorganisms or insects and, therefore, inferences from these to man are more than ordinarily dubious. We have felt that to apply in toto the philosophy of safety evaluation of food additives and pesticides to the area of drugs does not represent a sound approach, since some risk is inherent in all drugs and thus the benefit-to-risk ratio must be weighed on an individual drug and patient basis. For several years, FDA guidelines for preclinical safety testing of drugs have included reproduction studies, a segment of which takes into consideration adverse genetic effects in mammalian species. General guidelines for preclinical animal studies have been published in FDA Papers (reprint attached). More specific guidelines dealing with reproduction studies have been presented at scientific symposia (e.g., American Society of Pharmacology and Experimental Therapeutics, 1967) and published in scientific journals (e.g., J. Clin. Pharmacol, 1968). Copies of FDA guidelines have been made available to interested parties in industry and the academic community both here and abroad. It is felt that these guidelines would be disseminated more efficiently in this manner than they would by publication in the Federal Register. In this regard, it should be pointed out that methods for evaluating drugs for safety are continuously undergoing reappraisal. Although general principles should be followed, it would be a mistake to think of methodology in absolute terms as if there were some standard which, once discovered, would provide a blue print for all future investigations. The suitability of additional screening methods for mutagenic potential is being explored through contract proposals and a series of workshop sessions sponsored in part by this Administration. FDA also cosponsored a symposium on mutagenicity in Washington in November, 1970. As a result of this meeting, plans have been drawn up for a workshop to be conducted at Brown University in July of this year under the sponsorship of the Environmental Mutagen Society and the National Academy of Sciences. Indications are that FDA will also be asked to support this endeavor to bring together interested scientists from government, industry, and the academic community. GUIDELINES FOR REPRODUCTION STUDIES FOR SAFETY EVALUATION OF DRUGS FOR HUMAN USE, JANUARY 1966 For the past few years, we have been recommending the two-litter rat reproduction study as a routine screen for the appraisal of safety of new drugs for use during pregnancy and in women of childbearing potential. In view of recent advances in knowledge and technology, and with experience gained from this test, both in the laboratory and from the evaluations of submitted reports, it is our opinion that now more effective test designs should be employed for routine use. The two-litter test encompassed the many parameters of the reproductive process, among which we might list fertility in both the male and female; effects on the zygote, its transport and implantation; embryo-toxicity and fetal death; effects on parturition and the newborn, on lactation and care of the young; and especially, the teratogenic potential of the drug. We feel that the action of a drug on these same parameters can be investigated more effectively by the use of the experimental design described below. This design consists of the division of the studies into three segments each of which would pertain to a specific phase of the reproductive process. These segments are: I. Study of Fertility and General Reproductive Performance; II. Teratological Study; and III. Perinatal and Postnatal Study. An outline of these segments follows as our general recommendation. A. Basic protocol I. Study of Fertility and General Reproductive Performance. In this segment emphasis is placed on the effects of a given drug on gonadal function, estrous cycles, mating behavior, conception rates, and the early stages of gestation. Studies in this segment provide information of drug action on these functions in a relatively short time. Secondarily, this segment also provides for an overall pilot screening of the agent on the entire reproductive process including teratogenesis, late stages of gestation, parturition, and lactation. The results obtained from these studies should serve as a guide for subsequent studies in depth. II. Teratological Study.-Within this segment the effort is concentrated on determining whether a drug has a potential for embryotoxicity and/or teratogenic effects. For this purpose, drug administration is restricted to the period of organogenesis. It is possible to give high dosages of the drug for short periods of time which should yield results amenable to a more exacting interpretation of teratogenic potential. III. Perinatal and Postnatal Study. The purpose of this segment is to study the effects of a drug administered during the last third of pregnancy and the period of lactation. Studies should delineate the effects of the drug on late fetal development, labor and delivery, lactation, neonatal viability, and growth of the newborn. B. Experimental procedures I. Study of Fertility and General Reproductive Performance.-For an adequate study of fertility, both males and females should be studied. The rat has been the animal used most for this type of investigation. In the study of the drug effect on male fertility, rats of that sex should have attained a minimum age of 40 days before drug adininistration begins. They should be treated for 60 to 80 days prior to mating to assure the absence of a potential effect on spermatogenesis. Male animals from subacute or chronic drug toxicity studies may be used. These pretreated males can be mated with either treated or non-treated females. As a minimum, 10 male animals should be mated with 20 females. For the study of drug action on female fertility, adult or at least sexually mature animals should be used. It is advisable to establish estrous cycles by daily vaginal smears. After 14 days of drug administration, the females are exposed to males. The occurrence of copulation must be established by daily vaginal inspections for sperm or the copulatory plug. This finding is considered day 0 of pregnancy. Dosing of the animals should be continued daily. To determine whether conception has occurred, it would be desirable to check for new estrous periods or for the placental sign. (a) One-half of the females under test should be sacrificed on day 13 of their respective pregnancies. The dams should be examined for number and distribution of embroyos in each uterine horn, presence of empty implantation sites, and embryos undergoing resorption. In addition, any abnormal condition in the uterus that may have contributed to embryonic death should be noted. (b) The remaining dams should be continued on drug and allowed to litter normally. The duration of gestation should be calculated and the litters examined as soon as possible after delivery for litter size, stillborn and live born, and gross anomalies. The pups should be weighed individually at delivery. Any dead pups should be preserved for a study of skeletal anomalies. The pups should be weighed and counted again on day 4 and day 21. Efforts should be made to determine the cause of any adverse effects observed. At this point it may be decided whether a second litter should be initiated, or in special cases (e.g., sex steroids) whether the reproductive performance of the offspring should be studied (F, matings). II. Teratological Study. In tests for teratogenic effects, at least two species should be employed. The species used most frequently have been the mouse, rat, and rabbit. Drug treatment should be started early enough and continued long enough to cover the period of organ formulation for the particular species used. Thus, the treatment period for the mouse and rat would be day 6 through day 15 of pregnancy (day 0 being the day sperm were found). In the rabbit the corresponding period would be day 6 through 18. Untreated males should be used to produce these pregnancies. Fetuses should be delivered by cesarean section one or two days prior to the anticipated date of parturition. The number of fetuses, their placement in the uterine horn, correlation with the number of corpora lutea, live and dead fetuses, and early and late resorptions should all be determined. Fetuses should be weighed individually and each one should be carefully examined immediately for any external anomalies. It is important that internal defects be sought even though no external anomalies are seen. Rat fetuses can be randomized into groups, i.e., onethird for dissection or alternatively the slicing method of Wilson, to discover visceral anomalies, and two-thirds for clearing and bone staining with alizarin. In a teratology study using rabbits, it might be desirable to incubate one-half of the fetuses for a period of 24 hours, with frequent observations for survival during the first 6 hours. The fetuses should then be preserved by appropriate means and processed for a study of visceral and skeletal anomalies. In the case of rabbits all fetuses should be examined for external, viceral, and skeletal anomalies. III. Perinatal and Postnatal Study.-The period of drug administration to the dam should cover the final one-third of gestation and continue throughout lactation to weaning. Particular emphasis should be placed on observation of labor and delivery. Dystocia, prolonged labor, and delayed labor are all possibilities. The duration of gestation should be calculated. The observations on litter size, pup weight, etc., mentioned in segment I-b should be carried out. Continuous administration through the nursing period will allow for detection of adverse effects on lactation, nursing instinct, as well as a toxic action of the drug or its metabolities on the newborn by secretion in the milk. Some litters could be exchanged between between control and high-dose dams to elucidate causes of poor survival, if such is observed. Because some drugs may produce effects not detectable in early life, some of the offspring may have to be raised to adulthood. As examples, some drugs may cause poor maze learning in later life or sterility of the F1 generation. C. Specific considerations I. Drug Route.-While several routes of drug administration may be employed, it would be preferable to use the same one planned for clinical administration. For an oral preparation administration should be by stomach tube or by capsule when possible, rather than by incorporation in the diet. When clinical administration is to be by the dermal route and lack of absorption systemically can be established, reproduction studies need not be performed. However, if this cannot be shown, the drug may be given topically under an occlusive covering or it may be given orally if it is absorbed by that route. Further modifications might be needed for special situations, when the mode of application (e.g., vaginal) of itself is apt to cause an adverse action on the dam. In such cases it might be appropriate to modify the schedule of treatment by dividing time of administration for a teratological study into shorter sub-periods, for separate groups of animals, the sums of these sub-periods to cover the entire suggested treatment period. II. Drug Dosage.-At least two dosage levels should be used. The high dosage should be sub-toxic, i.e., the maximum tolerated dosage, so as not to adversely affect the dam by anorexia, sedation, or other exaggerated pharmacological effects. The maximum tolerated dosage may not be the same for all 3 segments. The lower dosage should take into account the proposed therapeutic dose and |