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A risk assessment for acrylonitrile in consumer products
The Science of the Total Environment, 99 (1990) 263-279 Elsevier Science Publishers B.V., Amsterdam
263
A RISK A S S E S S M E N T FOR ACRYLONITRILE IN CONSUMER PRODUCTS*
P.K. JOHNSTON** and A.R. ROCK*
Directorate for Health Sciences, U.S. Consumer Product Safety Commission, 5401 Westbard Ave., Bethesda, MD 20207 (U.S.A.)
ABSTRACT A carcinogenic risk assessment for acrylonitrile in consumer products was prepared as part of the Second Workshop on Pragmatics of Risk Assessment, Bethesda, MD. Data from one inhalation and two oral rat bioassays served as input into several high-to-low-dose mathematical risk extrapolation models. The final unit risk estimates for humans were based on maximum likelihood estimates from the Global83 implementation of the multistage model after adjustments for surface area differences, continuous versus intermittent exposures, and the proportion of lifetime exposed. The unit risk estimates for lifetime exposure to 1 mg k g - ' d a y - 1 by inhalation and ingestion were 0.0531 and 0.2385, respectively. These risks are equivalent to risks of 3.3 × 10 -s for inhalation of l p p t in air and 3.4 × 10 9 for ingestion of 1 ng day.-
INTRODUCTION
Risk assessment at the U.S. Consumer Product Safety Commission (CPSC) is performed in several steps, including health effects assessment, dose~ response assessment, exposure assessment, and development of final risk estimates. Integral to this process is the evaluation of the limitations of the data and the uncertainties involved at each step in the process. Using the data received for use at this workshop [1-8], a risk assessment for acrylonitrile has been prepared. An explanation of the procedure used to obtain the final risk estimates and a discussion of factors which may have resulted in uncertainty in the risk estimates are presented below. HEALTH EFFECTS ASSESSMENT
A review of information on health effects of acrylonitrile highlighted two
*The views presented in this paper are solely those of the authors, who were employees of the U.S. Consumer Product Safety Commission when the paper was presented, and do not necessarily reflect the official opinions of the U.S. Consumer Product Safety Commission. Because this paper was written at the expense of the U.S. Government, it is in the public domain and may be freely copied or reprinted in accordance with 17 U.S.C. 105. **Author to whom correspondence should be addressed at the U.S. Environmental Protection Agency (ANR-445), 401 M St., S.W., Washington, DC 20460, U.S.A. t P r e s e n t address: Department of Epidemiology, UCLA School of Public Health, Los Angeles, CA, U.S.A. 0048-9697/90/$03.50
© - - 1990 Elsevier Science Publishers B.V.
264
areas of possible concern to consumers using products containing acrylonitrile. These were teratogenicity and carcinogenicity. Evidence for teratogenic effects [1] was not convincing. Although a slight increase (p = 0.06) in total malformations was seen at an inhalation dose which did not appear to be maternally toxic (80 ppm), no increases in malformations were observed at specific organ sites. Other, significant increases (p < 0.05) were seen only at a maternally toxic oral dose (65mgkg-lday-1). The authors suggested that one defect observed at the 65 mg k g - l d a y 1 dose, right-sided aortic arch, was not due to maternal toxicity since it had not been seen in over 1000 litters examined in their laboratory, even though some dams studied were under greater stress; however, further studies, perhaps using shorter dosing periods to decrease maternal toxicity, would need to be performed to confirm this effect. Data presented on the carcinogenicity of acrylonitrile were more convincing. Evidence for the carcinogenicity of acrylonitrile is presented below. These data were used to assess the carcinogenic risk of acrylonitrile by two routes: ingestion and inhalation.
Evidence for carcinogenicity Several rat bioassays, especially those of Quast et al. [2, 3] and Bio/dynamics [4~6], show significant increases in tumors at several similar sites in two different strains (Fischer 344 and Spartan Sprague-Dawley) and by two different routes of exposure (oral and inhalation). These studies strongly support the carcinogenicity of acrylonitrile in this species and are sufficient to suggest that acrylonitrile is a human carcinogen. Evidence for carcinogenicity in another mammalian species would, however, give stronger support to the classification of acrylonitrile as a carcinogen. Although human epidemiological data are available for acrylonitrile [8], they were not considered in this risk assessment because our analysis was limited to the original articles in the data package provided for the workshop [1-7]. The U.S. Environmental Protection Agency (EPA) [8], however, has reported that human evidence for carcinogenicity would be regarded as somewhere between ~'sufficient" and ~'limited" using the International Agency for Research on Cancer (IARC) classification scheme. Some, additional information which suggests oncogenic potential for acrylonitrile was also reported by EPA [8]. Included is the ability of acrylonitrile to transfo~n' golden hamster embryo cells in culture and to enhance transformation of these cells by simian adenovirus SA7. In addition, acrylonitrile and/or its metabolites have been shown to cause bacterial mutations and sisterchromatid exchange in short-term mutagenicity studies and to bind to DNA in vitro. As noted above, data from the rat bioassays alone provided sufficient evidence to suggest that acrylonitrile is a human carcinogen. Because the
265 original epidemiological studies were not provided, these bioassays were used as the basis for our risk assessment on acrylonitrile.
Pharmacokinetic factors which may affect carcinogenicity Several pharmacokinetic factors may have an impact on the carcinogenic risk assessment for acrylonitrile. Although no data were available which suggested qualitative differences in the metabolism of acrylonitrile by different species, the proportions of various metabolites formed in studies on rodents and monkeys appeared to be species-, route-, and dose-dependent [8]. However, no pharmacokinetic data from humans were available.
Evaluation of bioassay data In preparing our risk assessment, three studies were used: an inhalation study and a drinking water study by Quast et al: in Sprague-Dawley rats [2, 3], and a drinking water study by Bio/dynamics in Fischer rats [4]. Several additional bioassays were not used. Two gavage studies [5, 7] were deleted because the drinking water route was considered more appropriate to the types of exposures which consumers could receive (i.e., frequent smaller doses). An inhalation study by Maltoni et al. [7] was excluded because it did not show a significant tumor increase with exposure. A more pronounced carcinogenic response was found at similar sites in the studies used in the risk assessment, probably due to increased doses and length of exposure to acrylonitrile. A three generation reproductive study by Beliles et al., cited by the U.S. EPA [8], was not used for three reasons: a copy of the actual study was not received, the tumor incidences reported were low, and a standard bioassay format was not used. Finally, a drinking water study by Bio/dynamics using Sprague-Dawley rats [6] was removed from analysis because discrepancies were found in the data which could not be addressed without contacting outside sources. Because the dose range used in the latter study was included in the other two drinking water studies, it was felt that, for the present, those two bioassays would suffice. In a review and evaluation of the three studies used, several general factors were found which may have led to uncertainty in the risk assessment. First, decreases in body weight at the higher doses may have affected the incidence of tumor formation. Second, high mortality at high doses may have precluded an adequate period for tumor formation. Third, histopathological analyses for the studies may not have been "blind", resulting in possible bias on the part of the examiner. In addition, several factors were found which were specific to the studies by Quast et al. [2, 3]: (i) for the inhalation study, only two dose groups were used, possibly making curve-fitting for the risk assessment less accurate; (ii) animals were housed in groups, which may have resulted in increased stress or disease; (iii) for the drinking water study, not enough information was provided to
266 TABLE 1 Data used in risk assessment - Quast et al. i n h a l a t i o n study [2]a Site
Dose (ppm) 0
20
80
Nervous system Astrocytoma
0/98
4/47 b
15/98"
Tongue Papilloma or carcinoma of squamous epithelium
1/95
7/89*
Small intestine Mucinous cystadenocarcinoma
2/98
14/97"
Males
Females Nervous system Astrocytoma
0/99
4/100
16/99"
*Fisher's exact test, one-sided; p < 0.05. aExcludes data for animals dying before 6 months and/or not histopathologically examined. bValue varies in report; this value appears correct based on number of animals histopathologically examined at 6 months.
determine if acrylonitrile was volatilized from the water or if its composition changed on standing in the water; (iv) no data on histopathological analyses of individual animals were presented, making interpretation of the data more difficult; (v) a greater number of sections were taken through the nervous system and, in the drinking water study, the stomach, when grossly recognized lesions were present, which may have biased detection of tumors; and (vi) only a small number of animals was histopathologically examined at some of the major sites, and the selection criteria used (e.g., grossly recognized lesions, occurrence with another tissue to be examined) may have biased the results. To remove the latter source of uncertainty from the analysis, data for certain sites or doses where only a small percentage of the animals were histopathologically examined were not used in the risk assessment. Data used from the Quast et al. inhalation study [2] are presented in Table 1. Data have been excluded for animals dying before 6 months, because these animals did not appear to have been on test long enough to allow for a sufficient latent period. In addition, only data for animals histopathologically examined for the tissue of concern were included. Statistically significant increases in tumors at the 80 ppm dose were found for the nervous system in both sexes and the tongue and small intestine in males.
267 TABLE 2 Data used in risk assessment
Quast et al. drinking water study [3]a
Males Site
Dose in mg kg- ~day 1 (ppm) 0 (0)
3.42 (35)
8.53 (100)
21.18 (300)
Nervous system Astrocytoma
1/79
8/47*
19/47"
23/48*
Tongue Papilloma or carcinoma of squamous cell epithelium
1/74
5/40*
Glandular stomach/small intestine Mucinous cystadenocarcinoma
3/79
8/48*.b
Females Site
Dose in mgkg 1day 1 (ppm) 0 (0)
4.36 (35)
10.76 (100)
24.97 (300)
Nervous system Astrocytoma
0/80c
17/48*'c
22/48*
24/47*
Tongue Papilloma or carcinoma of squamous cell epithelium
0/78
Nonglandular stomach Squamous cell carcinoma
0/80
0/47
0/48
12/47"
Squamous cell carcinoma or papilloma
1/80
1/47
12/48"
30/47*
Small intestine Mucinous cystadenocarcinoma
0/80
12/44"
4/47*
*Fisher's exact test, one-sided; p < 0.05. aExcludes data for animal dying before 6 months and/or not histopathologically examined. bValue may be 47 rather than 48; number of animals histopathologically examined for glandular stomach and/or small intestine was not presented. "Values vary by one tumor in text and appendix; values used were those in the appendix. D u r i n g t h e e v a l u a t i o n o f d a t a f r o m t h e Q u a s t et al. i n h a l a t i o n s t u d y , s e v e r a l s e t s o f d a t a w e r e e x c l u d e d f r o m t h e r i s k a s s e s s m e n t . O n l y d a t a for a c t u a l t u m o r s ( a s t r o c y t o m a s ) w e r e u s e d f r o m t h e n e r v o u s s y s t e m . D a t a for g l i a l cell p r o l i f e r a t i o n suggestive of a t u m o r were n o t used. T o n g u e a n d small i n t e s t i n e
268
tumor data for males at the 20 ppm dose were excluded because only a small number of animals was histopathologically examined and may not have been randomly selected. Data for external ear gland canal tumors were deleted because the investigators stated t h a t they were histopathologically examined only if grossly enlarged, the number examined was not reported, and the tumor response was low in comparison to other sites. Data for mammary gland tumors in females were not used because only a small number of animals were histopathologically examined, the r a t strain used has a high spontaneous incidence of mammary gland tumors, and the t um or response was low in comparison to other sites. TABLE 3 Data used in risk assessment - Bio/dynamics drinking water study [4]a Males Site
Dose in mg kg- 1day- 1 (ppm)b 0 (0)
0.11 (1)
0.25 (3)
0.81 (10)
2.49 (30)
8.15 (100)
Nervous system Astrocytoma
3/180
2/90
1/89
2/90
10/89"
21189"
Ear canal Carcinomas
1/169
0/87
0/83
2/78
5/83"
9/84"
2/169
1/87
0/83
2/78
7/83*
16/84"
0/179
1/90
4/87"
4/90*
4/90*
Papillomas, adenomas, carcinomas Stomach Papillomas
Females Site
Dose in mg kg- 1day- 1 (ppm)b 0 (0)
0.12 (1)
0.36 (3)
0.82 (10)
3.65 (30)
10.89 (100)
Nervous system Astrocytoma
1/178
1/90
2/90
4/87"
6/90"
24/88"
Ear canal Carcinomas
0/173
0/84
1/81
2/80
2/84
8/76*
0/173
0/84
2/81
4/80*
5/84*
10/76"
11178
1/90
2/89
2/87
4/90"
Papillomas, adenomas, carcinomas Stomach Papillomas
*Fisher's exact test, one-sided; p < 0.05. aBased on tabulations of data for individual animals, excluding data for animals dying or sacrificed before or at 6 months and/or not histopathologically examined. bData on dose received not included in package received; values in EPA document used.
269 Data used from the Quast et al. drinking water study [3] are presented in Table 2. Nervous system tumors for both sexes were significantly increased at all treatment levels. Nonglandular stomach tumors in females were significantly increased at 100 and 300 ppm. Tumors of the tongue in both sexes, the combined glandular stomach and small intestine in males, and the small intestine in females were significantly increased at 300 ppm. In evaluating the Quast et al. drinking water study, several sets of data were excluded. Data for nervous system cell proliferation, tongue tumors at the intermediate doses, external ear gland canal tumors, and mammary gland tumors were excluded for the same reasons as for the inhalation study. Data for nonglandular stomach tumors in males were deleted because data in the text and appendix varied by 10-15 animals with carcinomas at high dose. Data for the combined glandular stomach and small intestine in males at the intermediate dose were not used because only a small number of animals was histopathologically examined for the small intestine and may not have been randomly selected. Data used in the risk assessment from the Bio/dynamics drinking water study [4] are presented in Table 3. Nervous system and ear canal tumors were significantly increased at 10, 30, and 100 ppm for females and at the two higher doses for males. Stomach papillomas were significantly increased at 3, 10, and 30 ppm for males and at 30 ppm for females. Data for stomach papillomas at the high dose were excluded from the risk assessment because the response was decreased in comparison to the other doses, possibly due to increased mortality at the high dose. In evaluating the Bio/dynamics drinking water study, lack of statistical analysis of the histopathological data made interpretation of the data more difficult. The data from the three studies, presented in Tables 1-3, served as input to the dose-response assessment for acrylonitrile. DOSE RESPONSE ASSESSMENT Two types of model are typically used to extrapolate human risks from long-term animal bioassay data. One type is that based upon a statistical distribution (i.e., one t h a t assumes the susceptibility of the population at risk follows t h a t distribution) and the other assumes a hypothetical mechanism of carcinogenic action. Both types of model can usually fit high-dose data quite well; however, it is at the lower doses t h a t the models can diverge from one another by orders of magnitude in their predicted estimates. An important consideration in the use of a particular model is whether or not the dose~ response curve is linear at low dose. Frequently, the deciding factor for whether or not there is significant variation between models is if the high-dose data are nonlinear. In carcinogenic risk assessment, it is necessary to utilize an approach that leads to or forces low-dose linearity, regardless of the nature of the high-dose data, unless there is clear evidence that the true response at low doses is
270 nonlinear. Low-dose linearity is expected if the carcinogenic mechanism of action of the chemical in question can "interact" with normal background carcinogenic activity that occurs within the body. This interaction is based upon the multistage theory of carcinogenesis t h a t assumes that cancer progresses through a series of stages which may take many years in the case of humans before a tumor is detected (i.e., a latent period). Direct reaction of a chemical with genetic material can lead to "initiation"; other mechanisms which contribute to the carcinogenic process include cell proliferation or turnover, the promotion of initiated cells to a more transformed state, and other largely unknown processes that lead to heritable genetic alterations. In addition, metabolism and biotransformation may also play a key role in tumor development. Although it is common that such background processes leading to tumor development will occur in humans, the background incidence of cancer in animals must be significantly exceeded in order t h a t the response not be masked. Therefore, in a carcinogenic bioassay, other known, outside contributors to the carcinogenic process are minimized as part of the design of the bioassay. This is a critical consideration when extrapolating from animals to humans. Thus, even though a chemical may produce positive results in a bioassay because of its ability to influence or cause multiple events, it does not necessarily follow t h a t the chemical must influence multiple events under conditions of low exposure in humans. Rather, the chemical may influence only one event at lower exposure levels, while other spontaneous, background processes are able to complete carcinogenic development. This is the concept that leads to low-dose linearity in the absence of other possible nonlinear influences, such as pharmacokinetics. Further details on risk extrapolation models, and the concepts of linearity at low dose, background processes, and the multistage theory, can be found elsewhere [9-14]. The data presented in Tables 1-3 provided the input for the high-to-low-dose mathematical risk extrapolation models including Global83, the multistage model [15]; and RISK81, the probit, logit and Weibull extreme value models [16]. The RISK81 models were only run for the major site, the nervous system, in all three data sets. This was done due to time limitations as well as the fact that consideration of the other sites would have changed ultimate estimates of risk by a factor of < 3. Global83 was run in an unrestricted mode (i.e., for the number of "stages" allowed) as opposed to a restricted mode (i.e., maximum number of "stages" allowed equals the number of dose levels minus one). The unrestricted mode allows the program to fit a maximum likelihood estimate (MLE) to the high-dose data more easily. Maximum likelihood estimates and 95% lower confidence limits (LCL) on dose for a predicted risk of 1 in 100000 (10 -5) are provided in Tables 4 and 5. Table 4 shows the results of the RISK81 models operated in the "independent" background mode, which can easily lead to nonlinear dose-response curves because of the assumption of background independence. The large risk
271 TABLE 4 Estimated risks from "independent" tolerance distribution models conditions of a bioassay to give a risk of 10 ~ Bioassay
Sex
Site/tumor
mg kg-lday 1 in rats under
Maximum likelihood estimate~ Probit
Logit
Weibull
NSb NS
0.1804 0.2924
0.0076 0.0149
0.0107 0.0107
Bio/dynamics M Oral F
NS NS
0.0210 0.0068
0.00098 0.0004
0.00062 0.00023
Quast et al. Oral
M F
NS NS
0.0033 0.18 x 10 ~
0.00001 0.56 x 10 12
0.0000004 0.74 x 10 l~
Bioassay
Sex
Site/tumor 95% lower confidence limit
Quast et al. Inhalation
Quast et al. Inhalation
M F
Probit
Logit
Weibull
M F
NS NS
0.0379 0.0683
0.0010 0.0023
0.0014 0.0016
Bio/dynamics M Oral F
NS NS
0.0059 0.0064
0.00019 0.0008
0.00011 0.0006
Quast et al. Oral
NS NS
0.0006 0.9 x 10 s
0.000001 0.16 x 10 13
0.00000004 0.16 × 10 ix
M F
aUsing RISK81 [16]. hNS, nervous system.
e s t i m a t e s r e s u l t i n g f r o m t h e i n p u t o f t h e Q u a s t et al. o r a l f e m a l e n e r v o u s s y s t e m d a t a a r e d u e to t h e i n i t i a l s t e e p n e s s of t h e d a t a a n d t h e f a c t t h a t t h e r e w e r e zero c o n t r o l a n i m a l s r e s p o n d i n g . S u c h a n o m a l i e s o f t e n o c c u r w i t h t h e RISK81 models. T a b l e 5 d i s p l a y s t h e r e s u l t s of t h e R I S K 8 1 m o d e l s o p e r a t e d i n t h e " a d d i t i v e " b a c k g r o u n d m o d e a n d G l o b a l 8 3 . T h e m a x i m u m l i k e l i h o o d e s t i m a t e s for t h e s e m o d e l s a r e d i s p l a y e d a t t h e t o p of t h e t a b l e a n d t h e 95% l o w e r c o n f i d e n c e l i m i t s a r e s h o w n a t t h e b o t t o m . T h e r e is r e l a t i v e l y l i t t l e v a r i a t i o n i n t h e m a x i m u m likelihood risk estimates in the "additive" models that were applied. The e x c e p t i o n s to t h i s s i m i l a r i t y a r e t h e p r o b i t m o d e l r i s k e s t i m a t e s i n t h e Q u a s t et al. i n h a l a t i o n s t u d y a t t h e s i t e o f t h e n e r v o u s s y s t e m a n d a l l m o d e l s for t h e Q u a s t e t al. o r a l f e m a l e n e r v o u s s y s t e m t u m o r s . T h e a d d i t i v e a n d t h e i n d e p e n d e n t R I S K 8 1 m o d e l s a r e s i m i l a r b e c a u s e t h e r e is n o b a c k g r o u n d i n t h e d a t a . T h e s e r i s k e s t i m a t e s a r e p r e s e n t e d for c o m p a r i s o n p u r p o s e s a n d w i l l n o t be c o n s i d e r e d f u r t h e r . T h i s is b e c a u s e , if t h e d a t a a r e l i n e a r , t h e e s t i m a t e s a r e s i m i l a r to t h e m u l t i s t a g e m o d e l e s t i m a t e s , a n d if t h e d a t a a r e n o n l i n e a r , t h e
272
%
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t-, ¢D O
O
O
O
o o
i
I
¢9 O a0 o
O
O
o.o. O
O
~D ¢9
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O X
c~ O
c5o
o
o
c5o
~9
o
~3 t~ .<
~o
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273
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x
x
r~
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274 TABLE 6 Risk from 1 mg kg-~day 1 to rats under conditions of the bioassay Bioassay
Sex
Site/tumor
Maximum likelihood estimate
95% lower confidence limit
Quast et al. Inhalation
M F
NS ~ NS
0.0017 0.0015
0.0024 0.0022
Bio/dynamics Oral
M F
NS NS
0.0323 0.0213
0.0435 0.0345
Quast et al. Oral
M F
NS NS
0.0500 0.0500
0.0500 0.0500
aNS, nervous system.
multistage model estimates would be used in any event because it would force linearity at low dose. The risks predicted by the multistage model served as the basis for subsequent risk analysis. The majority of the risk estimates predicted by the other models is very similar to the predictions of Global83. Patterns observed for the 95% lower confidence limits are similar to those for the MLEs. Again, the risk estimates for the probit model in the Quast et al. inhalation study at the site of the female nervous system deviate from the others, as do all the models for the Quast et al. oral female nervous system tumors. The surface area correction [14] was applied before risk estimates based upon a specific sex and site were calculated. Only the data from the Quast et al. inhalation study had to be converted, from ppm to m g k g lday 1, as data
TABLE 7 Risk from 1 mg k g - l d a y -1 to humans using the multistage model {risk × [70 000/animal weight (g)]i~3} Bioassay
Sex
Site/tumor
Maximum likelihood estimate
95% lower confidence limit
Quast et al. Inhalation
M F
NS ~ NS
0.0528 0.0534
0.0745 0.0783
Quast et al. Oral
M F
NS NS
0.2809 0.3251
0.2809 0.3251
Bio/dynamics Oral
M F
NS NS
0.2097 0.1385
0.2823 0.2242
a NS, nervous system.
275 from the other bioassays were presented in mg kg ~day 1. For the Quast et al. inhalation study, it was calculated t hat the dose for male rats continuously inhaling l p p m was 1.23mgkg-~day ~ and for female rats, 1.41mgkg 1day 1. Since the animals in the Quast et al. inhalation study inhaled acrylonitrile for 6 hours per day, 5 days per week, the risks from 1 mg kg 1day- ~ were adjusted by the factors 24/6 and 7/5. The risk from l m g k g - l d a y 1 to animals is shown in Table 6. The highest risk estimates can be seen in the Quast et al. oral study and the lowest risk estimates result from the Quast et al. inhalation bioassay data. It should be noted th at the risk estimates from 1 mg kg-~day 1 in the two drinking water studies are very similar. Table 7 shows the m g k g 1day 1 estimates for humans adjusted by the surface area correction factor, i.e. the factor [70000/animal wt(g)]lJ3; and the factor 26/24, the proportion of lifetime correction to give lifetime carcinogenic estimated risk from lifetime continuous exposure to 1.0mgkg ~day 1. The estimates were adjusted by the factor 26/24 to account for the animals being ~ 2 months old at the start of the bioassay. This is the simplest correction that can be made and there is no data to indicate t hat any other adjustment is appropriate. The surface area correction is used, as opposed to a m g k g 1day 1 correction, because, although both are not incompatible with existing empirical data, erring on the side of safety results in a "best" estimate of risk. Again, the risk estimates in the oral studies are very similar and are several times higher than the risk estimates based upon the inhalation study. The final ~'best" human risk estimates will be based on the maximum likelihood estimates, since they are linear at low dose as output t hrough use of the multistage model. That is, there is no need to force linearity by using the upper confidence limits on risk. If the route of exposure to this chemical were via inhalation, then the "best" risk estimate would be an average of the two inhalation values for both sexes of rats. If the route of exposure to this chemical were via the oral route, or ingestion, then the "best" risk estimate would be an average of the four oral estimates presented in Table 7. Therefore, a lifetime inhalation exposure of 1 mg kg ~day 1 would lead to an estimated unit risk of 0.0531, and a lifetime oral exposure of l m g k g ~day i would lead to an estimated risk of 0.2385. Site-specific risks have been provided; tumor incidences at independent sites were not combined before modeling. After modeling, risk estimates for different sites can be added together to present a total cancer risk, if desired. There are many uncertainties i nhe r ent in carcinogenic risk assessment. This particular example is an especially difficult one, as it is not typical of most chemicals due to the fact t ha t there are multiple studies and numerous sites responding. In sum, the major uncertainties of this process, species to species extrapolation and high- to low-dose extrapolation, must be borne in mind when reaching conclusions based upon these estimates.
276 EXPOSURE ASSESSMENT
Although exposure assessment is an integral part of risk assessment, an exposure assessment for acrylonitrile was not prepared. Very minimal exposure to this chemical would be expected from consumer products, since it is used as a chemical intermediate, residual levels are low, and the rate of leaching appears minimal [8]. Possible exposure scenarios from use of consumer products containing acrylonitrile include: (i) inhalation exposure due to the migration of acrylonitrile to indoor air from apparel, carpeting, and household furnishings; (ii) dermal exposure due to leaching from apparel worn by consumers; and (iii) oral exposure for infants from sucking on apparel or plastic toys. FINAL RISK ESTIMATES
Because an exposure assessment for acrylonitrile was not performed, final risk estimates based on specific exposure scenarios cannot be made. However, more general final risk estimates have been calculated for perspective using the estimated unit risks developed earlier for inhalation and oral exposures. For instance, assuming a 70 kg adult inhales 20 m ~ of air a day, containing 1 ppt of acrylonitrile, the carcinogenic risk would be 3.3 x 10 s. Assuming exposure through ingestion of 1 ng of acrylonitrile a day, the carcinogenic risk would be 3.4 x 10 9. ACKNOWLEDGEMENT
The authors t h a n k Dr Murray Cohn for his review of the manuscript. REFERENCES 1
2
3
4
5
F.J. Murray, B.A. Schwetz, K.D. Nitschke, J.A. John, J.M. Narris and P.J. Gehring, Teratogenicity of acrylonitrile given to rats by gavage or by inhalation, Food Cosmet. Toxicol., 16 (1978) 547-551. J.F. Quast, D.J. Schuetz, M.F. Balmer, T.S. Gushow, C.N. Park and M.J. McKenna, A two-year toxicity and oncogenicity study with acrylonitrile following inhalation exposure of rats. Prepared by the Toxicology Research Laboratory, Health and Environmental Sciences, Dow Chemical U.S.A., Midland, MI, for the Chemical Manufacturing Association, Washington, DC, 1980. J.F. Quast, C.E. Wade, C.G. Humiston, R.M. Carreon, E.A. Hermann, C.N. Park and B.A. Schwetz, A two-year toxicity and oncogenicity study with acrylonitrile incorporated into the drinking water of rats. Prepared by the Toxicology Research Laboratory, Health and Environmental Sciences, Dow Chemical U.S.A., Midland, MI, for the Chemical Manufacturing Association, Washington, DC, 1980. Bio/dynamics Inc., A twenty-four month oral toxicity/carcinogenicity study of acrylonitrile administered in the drinking water to Fischer 344 rats. Prepared by Bio/dynamics Inc., East Millstone, NJ, for Monsanto Company, St. Louis, MO, Project No. 77 1744, 1980. Bio/dynamics Inc., A twenty-four month oral toxicity/carcinogenicity study of acrylonitrile administered by intubation to Spartan rats. Prepared by Bio/dynamics Inc., East Millstone, NJ, for Monsanto Company, St. Louis, MO, Project No. 77-1746, 1980.
277 6
7 8
9 10
11 12 13 14 15 16
Bio/dynamics Inc., A twenty-four month oral toxicity/carcinogenicity study of acrylonitrile administered in the drinking water to Spartan rats. Prepared by Bio/dynamics Inc., East Millstone, NJ, for Monsanto Company, St. Louis, MO, Project No. 77 1745, 1980. C. Maltoni, A. Ciliberti and V. DiMaio, Carcinogenicity bioassays on rats of acrylonitrile administered by i n h a l a t i o n and by ingestion, Med. Lav., 68 (1977) 401411. U.S. Environmental Protection Agency, Health Assessment document for acrylonitrile. Office of Research and Development, U.S. Environmental Protection Agency, Research Triangle Park, NC, EPA-600/8-82-007F, 1983. K.S. Crump, D.G. Hoel, C.H. Langley and R. Peto, Fundamental carcinogemc processes and their implications for low dose risk assessment, Cancer Res., 36 (1976) 2973 2979. K.S. Crump, H.A. Guess and K.L. Deal, Confidence intervals and test of hypothesis concerning dose response relations inferred from animal carcinogenicity data, Biometrics, 33 (1977) 437~451. H.A. Guess and K.S. Crump, Low-dose-rate extrapolation of data from animal carcinogenicity experiments analysis of a new statistical technique, Math. Biosci., 32 (1976) 15~36. D. Hoel, Incorporation of background in dose-response models, Fed. Proc., 39 (1980) 73 75. R. Peto, Carcinogenic effects of chronic exposure to very low levels of toxic substances, Environ. Health Perspect., 22 (1978) 15~159. U.S. Interagency Staff Group on Carcinogens, Chemical carcinogens: a review of the science and its associated principles, Environ. Health Perspect., 67 (1986) 201-282. R.B. Howe and K.S. Crump, Global83 (computer program), Research Systems, Inc., Ruston, LA, 1983. J. Kovar and D. Krewski, RISK81: A computer program for low dose extrapolation of quantal response toxicity data, Health and Welfare Canada, 1981.
OPEN DISCUSSION
QUESTION FOR MS ROCK FROM A MEMBER OF T H E AUDIENCE: When you, as the CPSC, select a single endpoint for your risk extrapolation do you just leave out all of the ot her animals with all the other tumor sites? Or, in o t h e r words, how do you a c c ount for the possibility t hat the animals h a v e n ' t other tumor sites and died of them? I'm sure t ha t you know that EPA combines all the animals with tumors. Let me say that again. When you pick a single tumor site to do a quantitative risk assessment, r a t h e r than all of the animals with tumors, how do you correct for the fact that, in picking a single tumor site, you eliminated the possibility t ha t the animals t hat died of other tumors, had they not gotten the other tumors, might also have died of this particular tumor? MS ROCK: We always present all the risk estimates for all of the sites, but we are required to come up with a single best estimate of risk. Because CPSC statutes require th at we do cost benefit analysis of hazards from carcinogens and seeing as we need one estimate t ha t is what we use.
QUESTION FOR DR JOHNSTON FROM DR BOORMAN: I t hought that you suggested th at the Maltoni Study was not used because the tumor incidence is low. I t h o u g h t th at I understood you to say that. My question to you is, if the Consumer P r o d u ct Safety Commission was faced with several studies would they discard one of those studies if the tumor response was weak or low?
278 DR JOHNSTON: It was not low. There were no significant increases in the inhalation study of Maltoni et al. DR BOORMAN: Let me rephrase it. If the Consumer Product Safety Commission was faced with three studies and you had reason to believe t hat they were all done with similar quality, and one was moderately positive, one was weakly positive and one was negative, would CPSC only use the strong positive to draw its conclusions? DR JOHNSTON: I believe t ha t we would only use a study t hat showed a significant statistical increase in tumors. We would not use a study t hat did not show any significant increase. DR BOORMAN: I would understand it if you are probably discarding some data, I would think t h a t perhaps all data can be used. I am not in risk assessment so I am probably on dangerous grounds, but I think t hat it seems unusual not to use three studies t ha t you believe are well done. QUESTION FOR DR J O H N S T O N FROM A M E MBE R OF THE AUDIENCE: You made some statement about discarding some tumors because of the gross examinations and sampling of lesions and I did not understand that. DR JOHNSTON: The authors stated t ha t they only examined the tissue if there were grossly recognized lesions. T hat may lead to some bias. If they examine all of the tissues they have a more representative statistical sample and they can determine if there were small tumors in some of the tissues, etc. We only use data where histopathological examinations were performed. In cases where there were biases we would not use the data. DR BOORMAN: I think t ha t in some cases t hat is true and in some cases t hat is not true. Mammary tumors in rats, for example, are rarely detected by histopathology, but are always detected grossly. So t h a t concept should be applied carefully. In the findings you said something about blind pathology (i.e. having the pathologist view the microscopic slides without knowing the group or the test animals from whence they came, Ed.). This is not a question, this is just a statement; I am strongly against blind pathology for the initial pathology evaluation. I think t hat it would be a giant leap backwards if you insisted on blind pathology. It would take a short presentation to explain all of the reasons, but I think t ha t the best way is to have the initial pathology evaluation done by a person who has all of the data at hand and can compare high dose with the controls. There may be equivocal lesions or you may have difficult lesions or lesions t hat are graded. You may take those lesions and look at them in a blind fashion. But always insisting on blind pathology in the beginning I think would be a step backwards.
279 QUESTION FOR DR JOHNSTON FROM DR THAKUR: My question refers to different models of testing. DR JOHNSTON: CPSC generally uses the multistage model because it forces linearity at low dose, especially with respect to the lower confidence limit. In this case the estimates came out very similar to many of the other models. So it was appropriate to use that.
263
A RISK A S S E S S M E N T FOR ACRYLONITRILE IN CONSUMER PRODUCTS*
P.K. JOHNSTON** and A.R. ROCK*
Directorate for Health Sciences, U.S. Consumer Product Safety Commission, 5401 Westbard Ave., Bethesda, MD 20207 (U.S.A.)
ABSTRACT A carcinogenic risk assessment for acrylonitrile in consumer products was prepared as part of the Second Workshop on Pragmatics of Risk Assessment, Bethesda, MD. Data from one inhalation and two oral rat bioassays served as input into several high-to-low-dose mathematical risk extrapolation models. The final unit risk estimates for humans were based on maximum likelihood estimates from the Global83 implementation of the multistage model after adjustments for surface area differences, continuous versus intermittent exposures, and the proportion of lifetime exposed. The unit risk estimates for lifetime exposure to 1 mg k g - ' d a y - 1 by inhalation and ingestion were 0.0531 and 0.2385, respectively. These risks are equivalent to risks of 3.3 × 10 -s for inhalation of l p p t in air and 3.4 × 10 9 for ingestion of 1 ng day.-
INTRODUCTION
Risk assessment at the U.S. Consumer Product Safety Commission (CPSC) is performed in several steps, including health effects assessment, dose~ response assessment, exposure assessment, and development of final risk estimates. Integral to this process is the evaluation of the limitations of the data and the uncertainties involved at each step in the process. Using the data received for use at this workshop [1-8], a risk assessment for acrylonitrile has been prepared. An explanation of the procedure used to obtain the final risk estimates and a discussion of factors which may have resulted in uncertainty in the risk estimates are presented below. HEALTH EFFECTS ASSESSMENT
A review of information on health effects of acrylonitrile highlighted two
*The views presented in this paper are solely those of the authors, who were employees of the U.S. Consumer Product Safety Commission when the paper was presented, and do not necessarily reflect the official opinions of the U.S. Consumer Product Safety Commission. Because this paper was written at the expense of the U.S. Government, it is in the public domain and may be freely copied or reprinted in accordance with 17 U.S.C. 105. **Author to whom correspondence should be addressed at the U.S. Environmental Protection Agency (ANR-445), 401 M St., S.W., Washington, DC 20460, U.S.A. t P r e s e n t address: Department of Epidemiology, UCLA School of Public Health, Los Angeles, CA, U.S.A. 0048-9697/90/$03.50
© - - 1990 Elsevier Science Publishers B.V.
264
areas of possible concern to consumers using products containing acrylonitrile. These were teratogenicity and carcinogenicity. Evidence for teratogenic effects [1] was not convincing. Although a slight increase (p = 0.06) in total malformations was seen at an inhalation dose which did not appear to be maternally toxic (80 ppm), no increases in malformations were observed at specific organ sites. Other, significant increases (p < 0.05) were seen only at a maternally toxic oral dose (65mgkg-lday-1). The authors suggested that one defect observed at the 65 mg k g - l d a y 1 dose, right-sided aortic arch, was not due to maternal toxicity since it had not been seen in over 1000 litters examined in their laboratory, even though some dams studied were under greater stress; however, further studies, perhaps using shorter dosing periods to decrease maternal toxicity, would need to be performed to confirm this effect. Data presented on the carcinogenicity of acrylonitrile were more convincing. Evidence for the carcinogenicity of acrylonitrile is presented below. These data were used to assess the carcinogenic risk of acrylonitrile by two routes: ingestion and inhalation.
Evidence for carcinogenicity Several rat bioassays, especially those of Quast et al. [2, 3] and Bio/dynamics [4~6], show significant increases in tumors at several similar sites in two different strains (Fischer 344 and Spartan Sprague-Dawley) and by two different routes of exposure (oral and inhalation). These studies strongly support the carcinogenicity of acrylonitrile in this species and are sufficient to suggest that acrylonitrile is a human carcinogen. Evidence for carcinogenicity in another mammalian species would, however, give stronger support to the classification of acrylonitrile as a carcinogen. Although human epidemiological data are available for acrylonitrile [8], they were not considered in this risk assessment because our analysis was limited to the original articles in the data package provided for the workshop [1-7]. The U.S. Environmental Protection Agency (EPA) [8], however, has reported that human evidence for carcinogenicity would be regarded as somewhere between ~'sufficient" and ~'limited" using the International Agency for Research on Cancer (IARC) classification scheme. Some, additional information which suggests oncogenic potential for acrylonitrile was also reported by EPA [8]. Included is the ability of acrylonitrile to transfo~n' golden hamster embryo cells in culture and to enhance transformation of these cells by simian adenovirus SA7. In addition, acrylonitrile and/or its metabolites have been shown to cause bacterial mutations and sisterchromatid exchange in short-term mutagenicity studies and to bind to DNA in vitro. As noted above, data from the rat bioassays alone provided sufficient evidence to suggest that acrylonitrile is a human carcinogen. Because the
265 original epidemiological studies were not provided, these bioassays were used as the basis for our risk assessment on acrylonitrile.
Pharmacokinetic factors which may affect carcinogenicity Several pharmacokinetic factors may have an impact on the carcinogenic risk assessment for acrylonitrile. Although no data were available which suggested qualitative differences in the metabolism of acrylonitrile by different species, the proportions of various metabolites formed in studies on rodents and monkeys appeared to be species-, route-, and dose-dependent [8]. However, no pharmacokinetic data from humans were available.
Evaluation of bioassay data In preparing our risk assessment, three studies were used: an inhalation study and a drinking water study by Quast et al: in Sprague-Dawley rats [2, 3], and a drinking water study by Bio/dynamics in Fischer rats [4]. Several additional bioassays were not used. Two gavage studies [5, 7] were deleted because the drinking water route was considered more appropriate to the types of exposures which consumers could receive (i.e., frequent smaller doses). An inhalation study by Maltoni et al. [7] was excluded because it did not show a significant tumor increase with exposure. A more pronounced carcinogenic response was found at similar sites in the studies used in the risk assessment, probably due to increased doses and length of exposure to acrylonitrile. A three generation reproductive study by Beliles et al., cited by the U.S. EPA [8], was not used for three reasons: a copy of the actual study was not received, the tumor incidences reported were low, and a standard bioassay format was not used. Finally, a drinking water study by Bio/dynamics using Sprague-Dawley rats [6] was removed from analysis because discrepancies were found in the data which could not be addressed without contacting outside sources. Because the dose range used in the latter study was included in the other two drinking water studies, it was felt that, for the present, those two bioassays would suffice. In a review and evaluation of the three studies used, several general factors were found which may have led to uncertainty in the risk assessment. First, decreases in body weight at the higher doses may have affected the incidence of tumor formation. Second, high mortality at high doses may have precluded an adequate period for tumor formation. Third, histopathological analyses for the studies may not have been "blind", resulting in possible bias on the part of the examiner. In addition, several factors were found which were specific to the studies by Quast et al. [2, 3]: (i) for the inhalation study, only two dose groups were used, possibly making curve-fitting for the risk assessment less accurate; (ii) animals were housed in groups, which may have resulted in increased stress or disease; (iii) for the drinking water study, not enough information was provided to
266 TABLE 1 Data used in risk assessment - Quast et al. i n h a l a t i o n study [2]a Site
Dose (ppm) 0
20
80
Nervous system Astrocytoma
0/98
4/47 b
15/98"
Tongue Papilloma or carcinoma of squamous epithelium
1/95
7/89*
Small intestine Mucinous cystadenocarcinoma
2/98
14/97"
Males
Females Nervous system Astrocytoma
0/99
4/100
16/99"
*Fisher's exact test, one-sided; p < 0.05. aExcludes data for animals dying before 6 months and/or not histopathologically examined. bValue varies in report; this value appears correct based on number of animals histopathologically examined at 6 months.
determine if acrylonitrile was volatilized from the water or if its composition changed on standing in the water; (iv) no data on histopathological analyses of individual animals were presented, making interpretation of the data more difficult; (v) a greater number of sections were taken through the nervous system and, in the drinking water study, the stomach, when grossly recognized lesions were present, which may have biased detection of tumors; and (vi) only a small number of animals was histopathologically examined at some of the major sites, and the selection criteria used (e.g., grossly recognized lesions, occurrence with another tissue to be examined) may have biased the results. To remove the latter source of uncertainty from the analysis, data for certain sites or doses where only a small percentage of the animals were histopathologically examined were not used in the risk assessment. Data used from the Quast et al. inhalation study [2] are presented in Table 1. Data have been excluded for animals dying before 6 months, because these animals did not appear to have been on test long enough to allow for a sufficient latent period. In addition, only data for animals histopathologically examined for the tissue of concern were included. Statistically significant increases in tumors at the 80 ppm dose were found for the nervous system in both sexes and the tongue and small intestine in males.
267 TABLE 2 Data used in risk assessment
Quast et al. drinking water study [3]a
Males Site
Dose in mg kg- ~day 1 (ppm) 0 (0)
3.42 (35)
8.53 (100)
21.18 (300)
Nervous system Astrocytoma
1/79
8/47*
19/47"
23/48*
Tongue Papilloma or carcinoma of squamous cell epithelium
1/74
5/40*
Glandular stomach/small intestine Mucinous cystadenocarcinoma
3/79
8/48*.b
Females Site
Dose in mgkg 1day 1 (ppm) 0 (0)
4.36 (35)
10.76 (100)
24.97 (300)
Nervous system Astrocytoma
0/80c
17/48*'c
22/48*
24/47*
Tongue Papilloma or carcinoma of squamous cell epithelium
0/78
Nonglandular stomach Squamous cell carcinoma
0/80
0/47
0/48
12/47"
Squamous cell carcinoma or papilloma
1/80
1/47
12/48"
30/47*
Small intestine Mucinous cystadenocarcinoma
0/80
12/44"
4/47*
*Fisher's exact test, one-sided; p < 0.05. aExcludes data for animal dying before 6 months and/or not histopathologically examined. bValue may be 47 rather than 48; number of animals histopathologically examined for glandular stomach and/or small intestine was not presented. "Values vary by one tumor in text and appendix; values used were those in the appendix. D u r i n g t h e e v a l u a t i o n o f d a t a f r o m t h e Q u a s t et al. i n h a l a t i o n s t u d y , s e v e r a l s e t s o f d a t a w e r e e x c l u d e d f r o m t h e r i s k a s s e s s m e n t . O n l y d a t a for a c t u a l t u m o r s ( a s t r o c y t o m a s ) w e r e u s e d f r o m t h e n e r v o u s s y s t e m . D a t a for g l i a l cell p r o l i f e r a t i o n suggestive of a t u m o r were n o t used. T o n g u e a n d small i n t e s t i n e
268
tumor data for males at the 20 ppm dose were excluded because only a small number of animals was histopathologically examined and may not have been randomly selected. Data for external ear gland canal tumors were deleted because the investigators stated t h a t they were histopathologically examined only if grossly enlarged, the number examined was not reported, and the tumor response was low in comparison to other sites. Data for mammary gland tumors in females were not used because only a small number of animals were histopathologically examined, the r a t strain used has a high spontaneous incidence of mammary gland tumors, and the t um or response was low in comparison to other sites. TABLE 3 Data used in risk assessment - Bio/dynamics drinking water study [4]a Males Site
Dose in mg kg- 1day- 1 (ppm)b 0 (0)
0.11 (1)
0.25 (3)
0.81 (10)
2.49 (30)
8.15 (100)
Nervous system Astrocytoma
3/180
2/90
1/89
2/90
10/89"
21189"
Ear canal Carcinomas
1/169
0/87
0/83
2/78
5/83"
9/84"
2/169
1/87
0/83
2/78
7/83*
16/84"
0/179
1/90
4/87"
4/90*
4/90*
Papillomas, adenomas, carcinomas Stomach Papillomas
Females Site
Dose in mg kg- 1day- 1 (ppm)b 0 (0)
0.12 (1)
0.36 (3)
0.82 (10)
3.65 (30)
10.89 (100)
Nervous system Astrocytoma
1/178
1/90
2/90
4/87"
6/90"
24/88"
Ear canal Carcinomas
0/173
0/84
1/81
2/80
2/84
8/76*
0/173
0/84
2/81
4/80*
5/84*
10/76"
11178
1/90
2/89
2/87
4/90"
Papillomas, adenomas, carcinomas Stomach Papillomas
*Fisher's exact test, one-sided; p < 0.05. aBased on tabulations of data for individual animals, excluding data for animals dying or sacrificed before or at 6 months and/or not histopathologically examined. bData on dose received not included in package received; values in EPA document used.
269 Data used from the Quast et al. drinking water study [3] are presented in Table 2. Nervous system tumors for both sexes were significantly increased at all treatment levels. Nonglandular stomach tumors in females were significantly increased at 100 and 300 ppm. Tumors of the tongue in both sexes, the combined glandular stomach and small intestine in males, and the small intestine in females were significantly increased at 300 ppm. In evaluating the Quast et al. drinking water study, several sets of data were excluded. Data for nervous system cell proliferation, tongue tumors at the intermediate doses, external ear gland canal tumors, and mammary gland tumors were excluded for the same reasons as for the inhalation study. Data for nonglandular stomach tumors in males were deleted because data in the text and appendix varied by 10-15 animals with carcinomas at high dose. Data for the combined glandular stomach and small intestine in males at the intermediate dose were not used because only a small number of animals was histopathologically examined for the small intestine and may not have been randomly selected. Data used in the risk assessment from the Bio/dynamics drinking water study [4] are presented in Table 3. Nervous system and ear canal tumors were significantly increased at 10, 30, and 100 ppm for females and at the two higher doses for males. Stomach papillomas were significantly increased at 3, 10, and 30 ppm for males and at 30 ppm for females. Data for stomach papillomas at the high dose were excluded from the risk assessment because the response was decreased in comparison to the other doses, possibly due to increased mortality at the high dose. In evaluating the Bio/dynamics drinking water study, lack of statistical analysis of the histopathological data made interpretation of the data more difficult. The data from the three studies, presented in Tables 1-3, served as input to the dose-response assessment for acrylonitrile. DOSE RESPONSE ASSESSMENT Two types of model are typically used to extrapolate human risks from long-term animal bioassay data. One type is that based upon a statistical distribution (i.e., one t h a t assumes the susceptibility of the population at risk follows t h a t distribution) and the other assumes a hypothetical mechanism of carcinogenic action. Both types of model can usually fit high-dose data quite well; however, it is at the lower doses t h a t the models can diverge from one another by orders of magnitude in their predicted estimates. An important consideration in the use of a particular model is whether or not the dose~ response curve is linear at low dose. Frequently, the deciding factor for whether or not there is significant variation between models is if the high-dose data are nonlinear. In carcinogenic risk assessment, it is necessary to utilize an approach that leads to or forces low-dose linearity, regardless of the nature of the high-dose data, unless there is clear evidence that the true response at low doses is
270 nonlinear. Low-dose linearity is expected if the carcinogenic mechanism of action of the chemical in question can "interact" with normal background carcinogenic activity that occurs within the body. This interaction is based upon the multistage theory of carcinogenesis t h a t assumes that cancer progresses through a series of stages which may take many years in the case of humans before a tumor is detected (i.e., a latent period). Direct reaction of a chemical with genetic material can lead to "initiation"; other mechanisms which contribute to the carcinogenic process include cell proliferation or turnover, the promotion of initiated cells to a more transformed state, and other largely unknown processes that lead to heritable genetic alterations. In addition, metabolism and biotransformation may also play a key role in tumor development. Although it is common that such background processes leading to tumor development will occur in humans, the background incidence of cancer in animals must be significantly exceeded in order t h a t the response not be masked. Therefore, in a carcinogenic bioassay, other known, outside contributors to the carcinogenic process are minimized as part of the design of the bioassay. This is a critical consideration when extrapolating from animals to humans. Thus, even though a chemical may produce positive results in a bioassay because of its ability to influence or cause multiple events, it does not necessarily follow t h a t the chemical must influence multiple events under conditions of low exposure in humans. Rather, the chemical may influence only one event at lower exposure levels, while other spontaneous, background processes are able to complete carcinogenic development. This is the concept that leads to low-dose linearity in the absence of other possible nonlinear influences, such as pharmacokinetics. Further details on risk extrapolation models, and the concepts of linearity at low dose, background processes, and the multistage theory, can be found elsewhere [9-14]. The data presented in Tables 1-3 provided the input for the high-to-low-dose mathematical risk extrapolation models including Global83, the multistage model [15]; and RISK81, the probit, logit and Weibull extreme value models [16]. The RISK81 models were only run for the major site, the nervous system, in all three data sets. This was done due to time limitations as well as the fact that consideration of the other sites would have changed ultimate estimates of risk by a factor of < 3. Global83 was run in an unrestricted mode (i.e., for the number of "stages" allowed) as opposed to a restricted mode (i.e., maximum number of "stages" allowed equals the number of dose levels minus one). The unrestricted mode allows the program to fit a maximum likelihood estimate (MLE) to the high-dose data more easily. Maximum likelihood estimates and 95% lower confidence limits (LCL) on dose for a predicted risk of 1 in 100000 (10 -5) are provided in Tables 4 and 5. Table 4 shows the results of the RISK81 models operated in the "independent" background mode, which can easily lead to nonlinear dose-response curves because of the assumption of background independence. The large risk
271 TABLE 4 Estimated risks from "independent" tolerance distribution models conditions of a bioassay to give a risk of 10 ~ Bioassay
Sex
Site/tumor
mg kg-lday 1 in rats under
Maximum likelihood estimate~ Probit
Logit
Weibull
NSb NS
0.1804 0.2924
0.0076 0.0149
0.0107 0.0107
Bio/dynamics M Oral F
NS NS
0.0210 0.0068
0.00098 0.0004
0.00062 0.00023
Quast et al. Oral
M F
NS NS
0.0033 0.18 x 10 ~
0.00001 0.56 x 10 12
0.0000004 0.74 x 10 l~
Bioassay
Sex
Site/tumor 95% lower confidence limit
Quast et al. Inhalation
Quast et al. Inhalation
M F
Probit
Logit
Weibull
M F
NS NS
0.0379 0.0683
0.0010 0.0023
0.0014 0.0016
Bio/dynamics M Oral F
NS NS
0.0059 0.0064
0.00019 0.0008
0.00011 0.0006
Quast et al. Oral
NS NS
0.0006 0.9 x 10 s
0.000001 0.16 x 10 13
0.00000004 0.16 × 10 ix
M F
aUsing RISK81 [16]. hNS, nervous system.
e s t i m a t e s r e s u l t i n g f r o m t h e i n p u t o f t h e Q u a s t et al. o r a l f e m a l e n e r v o u s s y s t e m d a t a a r e d u e to t h e i n i t i a l s t e e p n e s s of t h e d a t a a n d t h e f a c t t h a t t h e r e w e r e zero c o n t r o l a n i m a l s r e s p o n d i n g . S u c h a n o m a l i e s o f t e n o c c u r w i t h t h e RISK81 models. T a b l e 5 d i s p l a y s t h e r e s u l t s of t h e R I S K 8 1 m o d e l s o p e r a t e d i n t h e " a d d i t i v e " b a c k g r o u n d m o d e a n d G l o b a l 8 3 . T h e m a x i m u m l i k e l i h o o d e s t i m a t e s for t h e s e m o d e l s a r e d i s p l a y e d a t t h e t o p of t h e t a b l e a n d t h e 95% l o w e r c o n f i d e n c e l i m i t s a r e s h o w n a t t h e b o t t o m . T h e r e is r e l a t i v e l y l i t t l e v a r i a t i o n i n t h e m a x i m u m likelihood risk estimates in the "additive" models that were applied. The e x c e p t i o n s to t h i s s i m i l a r i t y a r e t h e p r o b i t m o d e l r i s k e s t i m a t e s i n t h e Q u a s t et al. i n h a l a t i o n s t u d y a t t h e s i t e o f t h e n e r v o u s s y s t e m a n d a l l m o d e l s for t h e Q u a s t e t al. o r a l f e m a l e n e r v o u s s y s t e m t u m o r s . T h e a d d i t i v e a n d t h e i n d e p e n d e n t R I S K 8 1 m o d e l s a r e s i m i l a r b e c a u s e t h e r e is n o b a c k g r o u n d i n t h e d a t a . T h e s e r i s k e s t i m a t e s a r e p r e s e n t e d for c o m p a r i s o n p u r p o s e s a n d w i l l n o t be c o n s i d e r e d f u r t h e r . T h i s is b e c a u s e , if t h e d a t a a r e l i n e a r , t h e e s t i m a t e s a r e s i m i l a r to t h e m u l t i s t a g e m o d e l e s t i m a t e s , a n d if t h e d a t a a r e n o n l i n e a r , t h e
272
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274 TABLE 6 Risk from 1 mg kg-~day 1 to rats under conditions of the bioassay Bioassay
Sex
Site/tumor
Maximum likelihood estimate
95% lower confidence limit
Quast et al. Inhalation
M F
NS ~ NS
0.0017 0.0015
0.0024 0.0022
Bio/dynamics Oral
M F
NS NS
0.0323 0.0213
0.0435 0.0345
Quast et al. Oral
M F
NS NS
0.0500 0.0500
0.0500 0.0500
aNS, nervous system.
multistage model estimates would be used in any event because it would force linearity at low dose. The risks predicted by the multistage model served as the basis for subsequent risk analysis. The majority of the risk estimates predicted by the other models is very similar to the predictions of Global83. Patterns observed for the 95% lower confidence limits are similar to those for the MLEs. Again, the risk estimates for the probit model in the Quast et al. inhalation study at the site of the female nervous system deviate from the others, as do all the models for the Quast et al. oral female nervous system tumors. The surface area correction [14] was applied before risk estimates based upon a specific sex and site were calculated. Only the data from the Quast et al. inhalation study had to be converted, from ppm to m g k g lday 1, as data
TABLE 7 Risk from 1 mg k g - l d a y -1 to humans using the multistage model {risk × [70 000/animal weight (g)]i~3} Bioassay
Sex
Site/tumor
Maximum likelihood estimate
95% lower confidence limit
Quast et al. Inhalation
M F
NS ~ NS
0.0528 0.0534
0.0745 0.0783
Quast et al. Oral
M F
NS NS
0.2809 0.3251
0.2809 0.3251
Bio/dynamics Oral
M F
NS NS
0.2097 0.1385
0.2823 0.2242
a NS, nervous system.
275 from the other bioassays were presented in mg kg ~day 1. For the Quast et al. inhalation study, it was calculated t hat the dose for male rats continuously inhaling l p p m was 1.23mgkg-~day ~ and for female rats, 1.41mgkg 1day 1. Since the animals in the Quast et al. inhalation study inhaled acrylonitrile for 6 hours per day, 5 days per week, the risks from 1 mg kg 1day- ~ were adjusted by the factors 24/6 and 7/5. The risk from l m g k g - l d a y 1 to animals is shown in Table 6. The highest risk estimates can be seen in the Quast et al. oral study and the lowest risk estimates result from the Quast et al. inhalation bioassay data. It should be noted th at the risk estimates from 1 mg kg-~day 1 in the two drinking water studies are very similar. Table 7 shows the m g k g 1day 1 estimates for humans adjusted by the surface area correction factor, i.e. the factor [70000/animal wt(g)]lJ3; and the factor 26/24, the proportion of lifetime correction to give lifetime carcinogenic estimated risk from lifetime continuous exposure to 1.0mgkg ~day 1. The estimates were adjusted by the factor 26/24 to account for the animals being ~ 2 months old at the start of the bioassay. This is the simplest correction that can be made and there is no data to indicate t hat any other adjustment is appropriate. The surface area correction is used, as opposed to a m g k g 1day 1 correction, because, although both are not incompatible with existing empirical data, erring on the side of safety results in a "best" estimate of risk. Again, the risk estimates in the oral studies are very similar and are several times higher than the risk estimates based upon the inhalation study. The final ~'best" human risk estimates will be based on the maximum likelihood estimates, since they are linear at low dose as output t hrough use of the multistage model. That is, there is no need to force linearity by using the upper confidence limits on risk. If the route of exposure to this chemical were via inhalation, then the "best" risk estimate would be an average of the two inhalation values for both sexes of rats. If the route of exposure to this chemical were via the oral route, or ingestion, then the "best" risk estimate would be an average of the four oral estimates presented in Table 7. Therefore, a lifetime inhalation exposure of 1 mg kg ~day 1 would lead to an estimated unit risk of 0.0531, and a lifetime oral exposure of l m g k g ~day i would lead to an estimated risk of 0.2385. Site-specific risks have been provided; tumor incidences at independent sites were not combined before modeling. After modeling, risk estimates for different sites can be added together to present a total cancer risk, if desired. There are many uncertainties i nhe r ent in carcinogenic risk assessment. This particular example is an especially difficult one, as it is not typical of most chemicals due to the fact t ha t there are multiple studies and numerous sites responding. In sum, the major uncertainties of this process, species to species extrapolation and high- to low-dose extrapolation, must be borne in mind when reaching conclusions based upon these estimates.
276 EXPOSURE ASSESSMENT
Although exposure assessment is an integral part of risk assessment, an exposure assessment for acrylonitrile was not prepared. Very minimal exposure to this chemical would be expected from consumer products, since it is used as a chemical intermediate, residual levels are low, and the rate of leaching appears minimal [8]. Possible exposure scenarios from use of consumer products containing acrylonitrile include: (i) inhalation exposure due to the migration of acrylonitrile to indoor air from apparel, carpeting, and household furnishings; (ii) dermal exposure due to leaching from apparel worn by consumers; and (iii) oral exposure for infants from sucking on apparel or plastic toys. FINAL RISK ESTIMATES
Because an exposure assessment for acrylonitrile was not performed, final risk estimates based on specific exposure scenarios cannot be made. However, more general final risk estimates have been calculated for perspective using the estimated unit risks developed earlier for inhalation and oral exposures. For instance, assuming a 70 kg adult inhales 20 m ~ of air a day, containing 1 ppt of acrylonitrile, the carcinogenic risk would be 3.3 x 10 s. Assuming exposure through ingestion of 1 ng of acrylonitrile a day, the carcinogenic risk would be 3.4 x 10 9. ACKNOWLEDGEMENT
The authors t h a n k Dr Murray Cohn for his review of the manuscript. REFERENCES 1
2
3
4
5
F.J. Murray, B.A. Schwetz, K.D. Nitschke, J.A. John, J.M. Narris and P.J. Gehring, Teratogenicity of acrylonitrile given to rats by gavage or by inhalation, Food Cosmet. Toxicol., 16 (1978) 547-551. J.F. Quast, D.J. Schuetz, M.F. Balmer, T.S. Gushow, C.N. Park and M.J. McKenna, A two-year toxicity and oncogenicity study with acrylonitrile following inhalation exposure of rats. Prepared by the Toxicology Research Laboratory, Health and Environmental Sciences, Dow Chemical U.S.A., Midland, MI, for the Chemical Manufacturing Association, Washington, DC, 1980. J.F. Quast, C.E. Wade, C.G. Humiston, R.M. Carreon, E.A. Hermann, C.N. Park and B.A. Schwetz, A two-year toxicity and oncogenicity study with acrylonitrile incorporated into the drinking water of rats. Prepared by the Toxicology Research Laboratory, Health and Environmental Sciences, Dow Chemical U.S.A., Midland, MI, for the Chemical Manufacturing Association, Washington, DC, 1980. Bio/dynamics Inc., A twenty-four month oral toxicity/carcinogenicity study of acrylonitrile administered in the drinking water to Fischer 344 rats. Prepared by Bio/dynamics Inc., East Millstone, NJ, for Monsanto Company, St. Louis, MO, Project No. 77 1744, 1980. Bio/dynamics Inc., A twenty-four month oral toxicity/carcinogenicity study of acrylonitrile administered by intubation to Spartan rats. Prepared by Bio/dynamics Inc., East Millstone, NJ, for Monsanto Company, St. Louis, MO, Project No. 77-1746, 1980.
277 6
7 8
9 10
11 12 13 14 15 16
Bio/dynamics Inc., A twenty-four month oral toxicity/carcinogenicity study of acrylonitrile administered in the drinking water to Spartan rats. Prepared by Bio/dynamics Inc., East Millstone, NJ, for Monsanto Company, St. Louis, MO, Project No. 77 1745, 1980. C. Maltoni, A. Ciliberti and V. DiMaio, Carcinogenicity bioassays on rats of acrylonitrile administered by i n h a l a t i o n and by ingestion, Med. Lav., 68 (1977) 401411. U.S. Environmental Protection Agency, Health Assessment document for acrylonitrile. Office of Research and Development, U.S. Environmental Protection Agency, Research Triangle Park, NC, EPA-600/8-82-007F, 1983. K.S. Crump, D.G. Hoel, C.H. Langley and R. Peto, Fundamental carcinogemc processes and their implications for low dose risk assessment, Cancer Res., 36 (1976) 2973 2979. K.S. Crump, H.A. Guess and K.L. Deal, Confidence intervals and test of hypothesis concerning dose response relations inferred from animal carcinogenicity data, Biometrics, 33 (1977) 437~451. H.A. Guess and K.S. Crump, Low-dose-rate extrapolation of data from animal carcinogenicity experiments analysis of a new statistical technique, Math. Biosci., 32 (1976) 15~36. D. Hoel, Incorporation of background in dose-response models, Fed. Proc., 39 (1980) 73 75. R. Peto, Carcinogenic effects of chronic exposure to very low levels of toxic substances, Environ. Health Perspect., 22 (1978) 15~159. U.S. Interagency Staff Group on Carcinogens, Chemical carcinogens: a review of the science and its associated principles, Environ. Health Perspect., 67 (1986) 201-282. R.B. Howe and K.S. Crump, Global83 (computer program), Research Systems, Inc., Ruston, LA, 1983. J. Kovar and D. Krewski, RISK81: A computer program for low dose extrapolation of quantal response toxicity data, Health and Welfare Canada, 1981.
OPEN DISCUSSION
QUESTION FOR MS ROCK FROM A MEMBER OF T H E AUDIENCE: When you, as the CPSC, select a single endpoint for your risk extrapolation do you just leave out all of the ot her animals with all the other tumor sites? Or, in o t h e r words, how do you a c c ount for the possibility t hat the animals h a v e n ' t other tumor sites and died of them? I'm sure t ha t you know that EPA combines all the animals with tumors. Let me say that again. When you pick a single tumor site to do a quantitative risk assessment, r a t h e r than all of the animals with tumors, how do you correct for the fact that, in picking a single tumor site, you eliminated the possibility t ha t the animals t hat died of other tumors, had they not gotten the other tumors, might also have died of this particular tumor? MS ROCK: We always present all the risk estimates for all of the sites, but we are required to come up with a single best estimate of risk. Because CPSC statutes require th at we do cost benefit analysis of hazards from carcinogens and seeing as we need one estimate t ha t is what we use.
QUESTION FOR DR JOHNSTON FROM DR BOORMAN: I t hought that you suggested th at the Maltoni Study was not used because the tumor incidence is low. I t h o u g h t th at I understood you to say that. My question to you is, if the Consumer P r o d u ct Safety Commission was faced with several studies would they discard one of those studies if the tumor response was weak or low?
278 DR JOHNSTON: It was not low. There were no significant increases in the inhalation study of Maltoni et al. DR BOORMAN: Let me rephrase it. If the Consumer Product Safety Commission was faced with three studies and you had reason to believe t hat they were all done with similar quality, and one was moderately positive, one was weakly positive and one was negative, would CPSC only use the strong positive to draw its conclusions? DR JOHNSTON: I believe t ha t we would only use a study t hat showed a significant statistical increase in tumors. We would not use a study t hat did not show any significant increase. DR BOORMAN: I would understand it if you are probably discarding some data, I would think t h a t perhaps all data can be used. I am not in risk assessment so I am probably on dangerous grounds, but I think t hat it seems unusual not to use three studies t ha t you believe are well done. QUESTION FOR DR J O H N S T O N FROM A M E MBE R OF THE AUDIENCE: You made some statement about discarding some tumors because of the gross examinations and sampling of lesions and I did not understand that. DR JOHNSTON: The authors stated t ha t they only examined the tissue if there were grossly recognized lesions. T hat may lead to some bias. If they examine all of the tissues they have a more representative statistical sample and they can determine if there were small tumors in some of the tissues, etc. We only use data where histopathological examinations were performed. In cases where there were biases we would not use the data. DR BOORMAN: I think t ha t in some cases t hat is true and in some cases t hat is not true. Mammary tumors in rats, for example, are rarely detected by histopathology, but are always detected grossly. So t h a t concept should be applied carefully. In the findings you said something about blind pathology (i.e. having the pathologist view the microscopic slides without knowing the group or the test animals from whence they came, Ed.). This is not a question, this is just a statement; I am strongly against blind pathology for the initial pathology evaluation. I think t hat it would be a giant leap backwards if you insisted on blind pathology. It would take a short presentation to explain all of the reasons, but I think t ha t the best way is to have the initial pathology evaluation done by a person who has all of the data at hand and can compare high dose with the controls. There may be equivocal lesions or you may have difficult lesions or lesions t hat are graded. You may take those lesions and look at them in a blind fashion. But always insisting on blind pathology in the beginning I think would be a step backwards.
279 QUESTION FOR DR JOHNSTON FROM DR THAKUR: My question refers to different models of testing. DR JOHNSTON: CPSC generally uses the multistage model because it forces linearity at low dose, especially with respect to the lower confidence limit. In this case the estimates came out very similar to many of the other models. So it was appropriate to use that.