First draft prepared by P.J. Abbott
Food Science and Safety Section, National Food Authority, Canberra,
Alitame was considered by the Committee at its forty-fourth
meeting (Annex 1, reference 116). At that meeting, the Committee
evaluated a large number of studies of toxicity, and specifications
and a toxicological monograph were prepared. The Committee concluded,
however, that an ADI could not be allocated because of concerns about
the deficiencies of the studies of carcinogenicity. Specifically, in a
study with Long-Evans rats, poor survival at 24 months and an
increased incidence of hepatocellular adenomas in females at the
highest dose were noted. While it was considered unlikely that the
adenomas would have progressed to carcinomas, the data were
inconclusive in this regard. In a study with Sprague-Dawley rats, low
survival rates were seen at 22 and 24 months, and the study was
considered inadequate for drawing conclusions about the potential
carcinogenicity of alitame.
At its present meeting, the Committee reconsidered the adequacy
of the carcinogenicity studies in rats in the light of general
considerations about the survival of animals in contemporary long-term
studies (Annex 1, reference 122) and on the basis of a further
statistical analysis of the data from the two studies, which
specifically assessed the power of the study in Sprague-Dawley rats to
detect hepatic lesions.
2. BIOLOGICAL DATA: Carcinogenicity
Two two-year studies were performed in rats with alitame, one in
Long-Evans rats and the other in the Sprague-Dawley strain. The
details of the design of these studies were described previously
(Annex 1, reference 117). Further information has been provided that
allows comparison of the two studies (Pfizer, 1996).
Table 1 shows the numbers of surviving animals in the two studies
at 18, 22, and 24 months. The initial numbers of animals were 50 per
group in the study with Long-Evans rats and 70 per group in the study
with Sprague-Dawley rats. At 24 months, the number of surviving
Sprague-Dawley rats was only slightly smaller than that of Long-Evans
rats, despite the difference in survival rates.
In the original report, data on hepatic lesions were analysed by
the statistical model of Peto et al. (1980). In the more recent
report, the data on the incidence of hepatocellular adenomas and
eosinophilic foci in the study on Long-Evans rats were re-examined
using the model of Dinse & Haseman (1986), which allows estimation of
the predictive importance of time on study and dose. The model
developed for the data from the study of Long-Evans rats was used to
analyse the survival data for the Sprague-Dawley rats, and the
Table 1. Numbers of surviving animals in studies with Long-Evans and
Sex Dose At 18 months At 22 months At 24 months
LE SD LE SD LE SD
Male High 35 49 19 24 14 18
Mid 41 44 23 11 15 6
Low 35 43 23 15 17 7
Controlsa 37 85 24 24 20 10
Female High 42 51 25 27 17 12
Mid 45 49 25 21 21 17
Low 40 56 28 32 20 22
Controlsa 46 111 32 60 23 42
LE, Long-Evans; SD, Sprague-Dawley
a Average of two control groups in the study of Sprague-Dawley rats
probability of developing an adenoma or eosinophilic foci was
determined. This predicted incidence was then compared with the
observed incidences. The model predictions and the observed outcomes
are shown in Table 2.
The author concluded that (i) the study with Sprague-Dawley rats
was of adequate length to assess the incidence of hepatic lesions, and
(ii) the process of hepatocellular adenoma formation in Long-Evans
rats is specific to that strain.
The Committee noted that while the survival of Long-Evans rats at
24 months was low, it was comparable to that seen in contemporary
studies using this strain, and no hepatocellular carcinomas were seen.
It also noted that the survival of Sprague-Dawley rats was comparable
to that in contemporary studies in rats of this strain and that
because there were more animals per group initially (70) than in the
study using Long-Evans rats (50), the number of surviving animals was
comparable between the two strains. On the basis of a logistic
regression model developed using data from the study with Long-Evans
rats, an increased probability of developing hepatocellular adenomas
and eosinophilic foci would have been expected in the Sprague-Dawley
rats, given the doses and time on study, but no increase was observed.
On the basis of these considerations the two studies were considered
adequate to assess carcinogenicity.
Table 2. Observed and predicted incidences of hepatocellular adenomas
and eosinophilic foci in studies of Long-Evans and
Lesion Strain Dose Observed Predicted
Hepatocellular Long-Evans Control 0 0.32
adenoma Low 0 0.43
High 12 12.25
Sprague-Dawley Controla 0 0.60
Low 0 0.49
High 0 12.72
Eosinophilic foci Long-Evans Control 2 3.98
Low 3 4.33
Mid 11 6.47
High 18 19.23
Sprague-Dawley Controla 8 8.22
Low 4 5.41
Mid 4 6.43
High 5 22.13
a Average of two control groups
The Committee concluded that there was no evidence that alitame
is carcinogenic. The Committee therefore allocated an ADI of 0-1 mg/kg
bw, on the basis of the NOAEL of 100 mg/kg bw per day identified at
the forty-fourth meeting in the 18-month study in dogs, using a safety
factor of 100.
The Committee noted that, although not specifically requested, a
further study of tolerance to repeated doses of alitame in diabetic
subjects is under way, and, as is customary, the results of this study
should be forwarded to the Committee when available.
Dinse, G.E. & Haseman, J.K. (1986) Logistic regression analysis of
incidental tumor data from animal carcinogenicity experiments.
Fundam. Appl. Toxicol., 6, 44-52.
Peto, R., Pike, M.C., Day, N.E., Gray, R.G., Lee, P.N., Parish, S.,
Peto, J., Richards, S. & Wahrendorf, J. (1980) Guidelines for simple,
sensitive significance tests for carcinogenic effects in long-term
animal experiments. In: IARC Monographs on the Evaluation of the
Carcinogenic Risk of Chemicals to Humans, Suppl. 2, Long-term and
Short-term Screening Assays for Carcinogens: A Critical Appraisal,
Lyon, International Agency for Research on Cancer, pp. 311-426.
Pfizer (1996) Alitame 2 year chronic studies with in-utero exposure
in Long-Evans and Sprague-Dawley rats: Comparative discussion of
findings related to survival. Unpublished report submitted to WHO by
Central Research Division, Pfizer Inc., Groton, Connecticut, USA.