Kurt Wegener

The German Thorotrast Study: Recent Results and Assessment of Risks

The German Thorotrast study comprises 2,326 patients and 1,890 controls. Forty-eight Thorotrast patients and 239 controls are still alive and are invited for a follow-up examination every 2 years. In the deceased patients, the following neoplastic diseases with excess rates were registered (Thorotrast/controls): liver cancer (454/3); cancer of the bile ducts, including gallbladder (42/7); myeloid leukemia (40/7); myelodysplastic syndrome (30/4); plasmacytoma (10/2); non-Hodgkins lymphoma (15/5); bone sarcoma (4/1); malignant peritoneal or pleural mesothelioma (9/0). Dose calculations are based on results of whole-body counting, X-ray films, and data obtained from the hospital records on the volume of Thorotrast injected. For liver cancer, the cumulative risk estimate was calculated to be 40 per 10(4) person Sv (radiation weighting factor = 20). These figures are close to the results of the Danish study and are comparable to the results of the Life Span Study of A-bomb survivors after 40 years at risk with 18 to 48 liver cancers per 10(4) person Sv. For hematopoietic malignancies, the cumulative risk was calculated to be about 7 per 10(4) person Sv (radiation weighting factor = 20). This risk estimate is lower by a factor of 10 compared to the results of the Life Span Study.

ras gene mutations in vinyl chloride-induced liver tumours are carcinogen-specific but vary with cell type and species

Previous studies have shown that a high proportion (5/6) of human liver angiosarcomas (ASL) associated with exposure to vinyl chloride (VC) contains a GC→AT mutation at the Ki‐ras codon 13. This mutation, however, has not been found in 5 ASL or 2 hepatocellular carcinomas (HCC) induced in rats by VC. These 2 HCC did contain a mutation at codon 61 of the Ha‐ras gene. In order to extend this study and further explore the mechanisms of tumour induction, an additional 6 ASL and 6 HCC induced in rats by VC were analysed for ras gene point mutations, as well as 10 rat and 10 murine ASL induced by vinyl fluoride (VF), and 5 ASL, 6 Kupffer cell sarcomas, 4 HCC and 2 cholangiocellular carcinomas induced by Thorotrast in rats. Tumour DNA was analysed by PCR‐SSCP and direct sequencing. None of the rodent ASL contained a mutation at codon 13 of the Ki‐ras gene showing that the ras gene mutational pattern is species‐specific. The CAA→CTA mutation, previously found at codon 61 of the Ha‐ras gene in rat HCC, was observed in 5 further VC‐induced HCC but was not detected in the Thorotrast‐induced HCC, suggesting carcinogen‐specificity. This mutation was also absent in VC‐induced ASL, which supports the cell‐specificity of the ras mutational pattern in chemically induced tumours. No predominant mutation was detected in VF‐ and Thorotrast‐induced tumours. Thus, a given mutation in a tumour may be carcinogen‐specific but also depend on the species and the cell type. Int. J. Cancer 85:223–227, 2000. ©2000 Wiley‐Liss, Inc.

The effects of Thorotrast and quartz on the induction of lung tumors in rats.

In a long-term animal study, the combined and separate effects of Thorotrast (colloidal 232ThO2) and silica dust on the induction of lung tumors were investigated. Female Wistar rats were exposed for 29 d to aerosol concentrations of quartz of either 6 mg m-3, 30 mg m-3, or 0 mg m-3 (6 h d-1, 5 d wk-1). After inhalation, one-half of all exposed animals received a single intravenous injection of enriched Thorotrast (600 microL, 2960 Bq 228 Th mL-1). In all quartz-exposed groups the incidence of benign and malignant lung tumors turned out to be more than 40%. The additional Thorotrast treatment (lifelong exhalation of 220Rn) led to a marked shortening of latency times (first lung tumor was found 1 y after treatment) and to a higher total incidence in the animals exposed to 30 mg m-3 quartz (57 of 87 animals with lung tumors = 65.5%). In the group treated only with Thorotrast, three of 87 animals developed lung tumors. Statistical methods that correct for intercurrent mortality showed a significant increase of the lung tumor risk with respect to Thorotrast treatment, even for the low quartz groups with nearly similar incidences of lung tumors (in the group with ThO2, 39 out of 87 = 44.8%; in the group without ThO2, 37 out of 82 = 45.1%). The tumors were found predominantly in the peripheral regions of the lung and were preceded by proliferation and hyperplasia of the alveolar and bronchiolar epithelium. The results demonstrate a pronounced interactive effect of quartz and Thorotrast on carcinogenesis of the lung. The underlying possible mechanisms are discussed.

German Uranium Miner Study: Historical Background and Available Histopathological Material

Mining activities in the former German Democratic Republic were documented as early as 1168 in the ore mountains (Erzgebirge) of Saxony. Silver, bismuth, cobalt, nickel and tungsten were mined from then up to the end of the 19th century. After the Second World War, the Soviet Occupation Authorities reopened the old silver mines in Saxony to mine uranium for the Soviet nuclear industry. About 400, 000 workers produced a total of 220,000 tons of uranium during the years 1946 to 1990. After the reunification of Germany, the archive of the Institute of Pathology of the mining area was opened for research. It contains protocols of 28,975 autopsy cases and about 400,000 slides collected from 1957 to 1992, about 66,000 tissue blocks, and 238 whole lungs. From the autopsy cases, 17,466 could be identified as workers of the uranium mining company. The remainder of the cases were in the population of the mining area. A comparison of the frequencies of malignancies of male workers older than 15 years with those of the population of the mining area for the years 1957 to 1989 demonstrates a significantly higher percentage of lung cancer among the uranium miners. There was no significant difference for other solid cancers and leukemias.

German Uranium Miner Study-Pathological and Molecular Genetic Findings

Uranium miners of the former Wismut company in Germany form the largest cohort of workers exposed to (222)Rn and dust in the world. The German Uranium Miner Study, Research Group Pathology, is evaluating the central pathology archive of the Wismut company. The main tasks of our study are pathological-anatomical and molecular genetic investigations of 28,975 autopsy cases and the evaluation of mining pollutants in the lungs by neutron activation analysis. As part of an observer agreement study, lung tumors are classified according to the WHO/IASLC classification and nontumorigenic lung disorders are registered. Lung tumors were analyzed for the presence of a proposed radon-specific mutation in the TP53 gene (formerly known as p53). Interim results are: (a) In the years 1957 to 1965, a high rate (69%) of small cell carcinomas was found which had declined to 34% by 1990. (b) The percentage of the deceased who suffered from silicosis is not higher in the group of lung tumors than in other tumor groups or the nontumor group. (c) The hypothesis of a radon-characteristic hotspot mutation in the TP53 tumor suppressor gene is not supported by our investigations. (d) Neutron activation analysis demonstrates that uranium, arsenic, chromium, cobalt and antimony can be found in tissue samples from the miners even when they had stopped working more than 20 years before death.

Recent Results of the German Thorotrast Study-statistical Evaluation of Animal Experiments With Regard to the Nonradiation Effects in Human Thorotrastosis

Our first long-term animal experiment made use of 1920 female Wistar rats divided into 20 groups of 96 animals each. These were injected at 12 weeks of age with different volumes and different dosages of Thorotrast which was enriched with 230Th to enhance the alpha-energy emission rate. The purpose of the study was to evaluate the effects due to the colloidal substance and the radiation. In the main experiment, 12 groups of rats were injected intravenously with 60, 120 and 300 microliters Thorotrast. 230Th was added to some Thorotrast preparations so that the total alpha-energy emission rate varied by factors of 1, 2, 5 and 10 relative to normal Thorotrast. Two groups were injected with 12 and 60 microliters of 50-fold enriched Thorotrast. One group was given 600 microliters of normal Thorotrast. In addition, we had 5 control groups, 1 NaCl and 4 Dextrin groups. The latter were injected with 60, 120, 300 or 600 microliters of Dextrin. The first animals died 8 months after injection, and the last 11 animals were killed 41 months after starting the experiment. The number of animals that developed a hepatic or splenic tumor increased by a factor of 10 in the highest dose-rate groups compared to controls. Our results demonstrated a linear correlation between the dose-rate and the number of primary hepatic and splenic tumors. It appeared that the volume of injected Thorotrast, by itself, had little influence on the number of tumors. However, at a constant dose-rate of 10, a 50-fold increase in the volume of Thorotrast (12-600 microliters) decreased the minimal tumor-appearance time by about 250 days.

The Combined and Separate Action of Neutron Radiation and Zirconium Dioxide on the Liver of Rats

To simulate the chronic alpha radiation of Thorotrast, the liver of female Wistar rats was exposed to fractionated neutron irradiation at 14-d intervals (0.2 Gy per fraction) over 2 y to a total dose of 10.0 Gy. Prior to the start of irradiation, one-half of the animals received 120 microL of non-radioactive Zirconotrast (ZrO2), which is comparable to Thorotrast with regard to all other physical and chemical properties. One year after beginning irradiation, the first liver tumor was detected. At the end of the life-span study, the incidence of irradiated animals with liver tumors was about 40%. In the animals treated additionally with ZrO2, the incidence, time of onset, and overall number of liver tumors was nearly equal, indicating that the fractionated neutron irradiation was the exclusive cause of tumor development. The lifelong-deposited ZrO2 colloid had no stimulating effect. Compared to earlier animal studies dealing with Thorotrast, the same histological types of benign and malignant liver tumors were found.

Preneoplastic foci of altered hepatocytes induced in rats by irradiation with α-particles of Thorotrast and neutrons

Prestages of hepatocellular neoplasms induced in rats by continuous internal alpha-radiation of Thorotrast or by fractionated external radiation with neutrons were studied by cytomorphological, cytochemical and morphometric methods. Irradiation with both Thorotrast and neutrons resulted in a significant increase in the number and volume fraction of foci of altered hepatocytes (FAH), the occurrence of which at 14 months correlated well with the previously reported increased incidence of hepatocellular neoplasms appearing after long lag periods. The morphological and biochemical phenotypes of radiation-induced FAH were similar to those of preneoplastic lesions described earlier in hepatocarcinogenesis elicited by chemicals or viruses.

Recent results of the German Thorotrast study--pathoanatomical changes in animal experiments and comparison to human thorotrastosis.

A survival experiment is described in which 1920 Wistar rats were used. These rats were injected intravenously with different quantities and different alpha-doses of Thorotrast. The following observations were made: The distribution of Thorotrast in the liver of the experimental animals is similar to that in human livers. Liver fibrosis and liver cirrhosis are rarely seen in experimental animals. The liver cell carcinomas, intrahepatic bile duct carcinomas and haemangiosarcomas that developed in the liver of the rats showed an identical biology and morphology with those seen in corresponding Thorotrast tumours in human patients. One particular tumour type that occurred in the liver of the rats probably represents a Kupffer cell sarcoma: the tumour cells show a positive peroxidase reaction and the metastases contained Thorotrast. Unlike human Thorotrast liver tumours, rat liver tumours include benign tumours such as liver cell adenomas and intrahepatic bile duct adenomas. The animals of the control group did not develop these benign liver tumours. The total frequency of the liver and spleen tumours in the trial groups receiving 230Th enriched Thorotrast was dependent on the dose given. The relationship between dose and effect was almost linear. The volume of the injected Thorotrast quantity, given a constant dose rate, seems to have only a slight influence on the number of tumours.

Translocation of Thorotrast in the body.

To investigate whether there is a permanent translocation of Thorotrast in the body, the liver of male Lewis rats was removed 4 weeks after injection of Thorotrast (300 microliters) and replaced by a donor liver. In half of the animals the spleen was removed as well. Measuring the 232Th content in the donor liver at different times after implantation demonstrated a permanent transport of 232Th into the surrogate organ. After 231 days a 232Th depot of about 1.1 mg was found, representing about 3% of the total body burden. The additional removal of the spleen resulted in a significantly lower transport of 232Th into the implanted liver. Histological examinations of the donated livers revealed increasing local concentration of Thorotrast granules, leading to the development of conglomerates. A comparable translocation of Thorotrast was verified in two humans who required liver transplantation more than 40 years after Thorotrast injection.