Disodium 2,2'-([1,1'-biphenyl]-4,4'-diyldivinylene)bis(benzenesulphonate)

Specific investigations: other studies

Description of key information

The test substance shows stimulatory effects on the rat exocrine pancreas. 
The test substance is not phototoxic.

Additional information

The first study (Bouis 1998) was designed to identify possible acute stimulatory effects on the rat exocrine pancreas by means of two experimental designs:

(1) Animals were fed overnight with the test substance admixed to the diet at 50000 ppm. Additional animals were fed overnight with cholestyramine, a bile salt binding resin known to exert a trophic effect on the rat pancreas, admixed to the diet at 60000 ppm. Following treatment, the animals were sacrificed and a possible stimulation of the exocrine pancreas was investigated by analysis of alpha amylase and lipase activities in pancreas homogenates.

(2) Animals were treated by single oral administration of the substance at 1000 mg/kg body weight and sacrificed after 90 min or 6 h. Additional animals were treated with camostat, a known trypsin inhibitor and stimulator of exocrine pancreatic growth (Goeke et al. 1986; endogenous CCK release and pancreatic growth in rats feed a preotease inhibitor (camostate), Pancreas 1; pp. 509 – 515), at 200 mg/kg body weight and also sacrificed after 90 min or 6 h. A possible stimulation of the exocrine pancreas was investigated by analysis of pancreatic alpha amylase and lipase activities in pancreas homogenates of animals sacrificed 90 min after treatment as well as by the determination of pancreatic ornithine decarboxylase (ODC) activity and polyamine content in animals sacrificed 6 h after treatment.

Overnight feeding or single oral administration of the test substance 5 resulted in a statistically significant (p≤0.05) decrease of pancreatic alpha amylase activity to 56 and 69% of control, respectively, as well as in a statistically significant (p≤0.05) decrease of pancreatic lipase activity to 78 and 62% of control, respectively. Similarly, overnight feeding with cholestyramine or a single oral administration of camostat statistically significantly (p≤0.05) decreased pancreatic hydrolytic enzyme activities to 57 and 37% of control for alpha amylase, and to 80 and 45% of control for lipase, respectively.

Upon single oral administration of the test substance, pancreatic ODC activity and pancreatic putrescine, spermidine and spermine contents were similar to control. In contrast, a single oral administration of camostat caused a strong, 150-fold statistically significant increase in pancreatic ODC activity, as well as a 7-fold increase in pancreatic putrescine content.

In conclusion, the results show that the substance, like cholestyramine and camostat, stimulated pancreatic enzyme release but was without an effect on pancreatic ODC activity and polyamine content.

A second study (Weber 1998) was designed to investigate an stimulating effect of the substance on the exocrine pancreas. For that purpose, male rats were treated with the substance admixed to the diet at 10000 or 50000 ppm for different time periods up to 28 days. Additional animals were treated by oral intubation with camostat at 200 mg/kg/day for the same time periods. The rats used were Tif:RAIf (SPF), from the same strain and supplier as in the lifetime feeding study (Basler 1990).

Treatment at 10000 or 50000 ppm resulted in an overall mean daily test article intake of 810 or 4306 mg/kg body weight, respectively. Treatment with the substance or the reference compound camostat was well tolerated and no treatment-related deaths or clinical signs were observed except for grayish/softened/sticky feces of animals treated with 50000 ppm.

Treatment with FWA-5 resulted in a statistically significant (p≤0.05) dose-related decrease in body weight. Since the food consumption, except for the first two days with a clear reduction at 50000 ppm, was similar in control and both treated groups, the food consumption ratio was increased in a dose-related manner after treatment with 10000 or 50000 ppm. This may indicate a reduced ability of food utilization or may at least partially be due to the

lower caloric content of food, admixed with 1% or 5% of the test substance. Treatment with camostat had no significant effect on body weight, food consumption or food consumption ratio.

In the absence of a significant effect on the absolute pancreas weight, treatment at 50000 ppm resulted in significantly (p≤0.05) increased relative pancreas weights at days 7 and 28. This weight effect is attributed to the observed hypertrophy and increased proliferation of pancreatic acinar cells. The acinar cell hypertrophy, observed from day 7 on, was determined by histopathology and by morphometry of acinar cell number per unit area. The observed hypertrophy was also reflected by moderately increased pancreatic protein content after 14 days of treatment with 50000 ppm. An increased proliferation of acinar cells, likewise contributing to the increased organ weight, was found at day 7. Treatment with 10000 ppm had no significant effects on pancreatic weight, protein content, histopathology and cell proliferation. Treatment with the reference compound camostat at 200 mg/kg/day induced strong hypertrophy and hyperplasia and consequently a strong increase in pancreatic weight and some increase in pancreatic protein content.

In addition to measurements on pancreatic acinar cell size and number, the contents of the pancreatic polyamines putrescine, spermidine and spermine were determined. These naturally occurring polyamines are known to play an important role in cell proliferation and differentia-tion. Increased pancreatic contents of polyamines putrescine and spermidine have also been reported in camostat stimulated pancreatic tissue (Loeser et al. 1989, Role or ornithine decarboxylase and polyamines in camostate (FOY-305)-induced pancreatic growth in rats. Digestion 43, pp 98 - 112). Statistically significant (p≤0.05) increased levels of putrescine (day 3) and spermidine (day 28) were found after treatment with the test substance or camostat (days 14 and 28). Although the absence of a clear time -and dose-relationship with the substance and camostat was not apparent in this relatively short study, the biological significance of these findings may suggest some adaptive response to the treatments or insufficient duration of the test. However, it can be concluded that this study indicates a statistically significant increase in pancreatic polyamines and stimulation in growth of the exocrine pancreas of male rats treated with 50000 ppm for up to 28 days.

Other investigations indicate enhanced pancreatic growth can promote non-genotoxic pancreatic tumor formation (Woutersen et al. 1990; Early indicators of exocrine pancreas carcinogenesis produced by non-genotoxic agents. Mutation Research, 248 pp 291 - 302). The initial indications of this pancreatic stimulation were observed with the substance after 28-days of dosing at 50000 ppm, which over a life-time of exposure could provide sufficient time to follow the classic course to malignancy: hyperplasia and adenoma leading to pancreatic carcinoma, as was seen in the 2-year study in rats (Ciba 1990e).

As reported above, the substance is not mutagenic and not genotoxic in standard assays. According to the concept of non-genotoxic carcinogenesis, thresholds exist below which no subchronic changes are observed and consequently no tumor development occurs (Purchase I 1994, Current knowledge of mechanisms of carcinogenicity: genotoxins versus non-genotoxins; Human Experimental Toxicology 13; pp 17 - 28). From the data presented, 5000 ppm and 10000 ppm are considered below the threshold for subchronic effects and, accordingly, no tumorigenicity was observed in the life-time carcinogenicity study at these doses (Ciba 1990e). For the substance it is concluded that the pancreatic tumorigenicity is not a genetic effect and may be viewed as a threshold event in light of the genotoxicity data, the low incidence of carcinomas, and the additional dose-response mechanism studies summarized here.

Phototoxicity: There were two studies done to determine wether the test substances could produce an augmented acute response of skin to a single ultraviolet light exposure. The results of pretreatment with the test substance were compared with pretreatment by a known phototoxic agent, 8 -methoxypsoralen (8 -MOP) or the vehicle using the skin of hairless mice, miniature pigs, and man. The test substance was not phototoxic in any species.