Donald Nagel

Current knowledge of pancreatic carcinogenesis in the hamster and its relevance to the human disease

Syrian hamsters present a unique species for induction of pancreatic tumors that in many aspects resemble human pancreatic cancer. The specific response of Syrian hamsters, in contrast to many other rodents, for development of pancreatic ductal (ductular) tumors is not yet known. All pancreatic carcinogens thus far tested show certain common features. They are all nitrosamines that possess or can be metabolized to compounds with 2‐oxopropyl‐ or 2‐hydroxypropyl substituents. All but one, N‐nitrosomethyl(2‐oxopropyl)amine, occur or metabolize to nitrosamines with the ability to cyclize and form structures resembling glucose. Hence it is suggested that this cyclic structure may be responsible for the pancreatic carcinogenicity of these nitrosamines, as has been proposed for the pancreatotropic effect of streptozotocin. It is also of further interest that one pancreatic ductal (ductular) carcinogen, N‐nitroso‐2‐methoxy‐2,6‐dimethylmorpholine, which possesses a totally cyclic structure, acts, like streptozotocin, as β‐cell cytotoxic and diabetogenic when given in a high single dose. Modification of pancreatic tumor induction has been demonstrated by specific procedures. A high fat diet significantly increases both the incidence and number of induced cancers. Methods for early diagnosis and therapy are being developed and their significance and applicabilities for clinical use will be of major importance. Compared with the other most common types of human cancer, pancreatic cancer has extraordinary characteristics, which make the disease one of the most mysterious of maladies. Consequently, pancreatic cancer represents a serious international problem and requires urgent resolution, especially with regard to its etiology, early diagnosis, prevention, and therapy.

Characterization of bifunctional adducts produced in DNA by trans-diamminedichloroplatinum(II)

The cancer chemotherapeutic drug cis-diamminedichloroplatinum(II) (cis-DDP) produces bifunctional reactions with DNA which appear critical to its toxic action. The relative inefficacy of the isomer trans-DDP results from its production of predominantly monofunctional adducts in DNA. However, trans-DDP is also toxic and this is presumed to result from bifunctional reaction. These reactions have been characterized by platinating pure DNA followed by enzyme digestion, HPLC separation and analysis by atomic absorption and nuclear magnetic resonance (NMR). Bifunctional adducts occur between deoxyguanosine (dG) and either deoxyadenosine (dA), deoxycytidine (dC) or another dG. Although dG-Pt-dG occurs in both double-stranded (approximately 40% of total adducts) and single-stranded DNA (approximately 60%) there is a marked preference for formation of dG-Pt-dC in double-stranded DNA (approximately 50%) and dG-Pt-dA in single-stranded DNA (approximately 35%). Only dG-Pt-dG forms rapidly; the other adducts derive from rapid formation of a monofunctional dG-Pt and further reaction with dA or dC over many hours.

Methylation of hamster DNA by the carcinogen N-nitroso-bis (2-oxopropyl)amine

The alkylation of hamster liver, lung and pancreas DNA by [1-14C]- and [2,3-14C]N-nitrosobis (2-oxopropyl) amine (BOP) has been examined. The specific activity of pancreas DNA after [2,3-14C]BOP administration was only 2% of that when [1-14C]BOP was given. 7-Methylguanine, but not O-6-methylguanine, was found in hydrolysates of liver and pancreas DNA. Nearly equal amount of alkylation were produced in the liver when [1-14C]- and [2,3-14C]BOP were given. At least one-half of the radioactivity in the liver was associated with N-alkylated purines, whereas only 20% was in this form in the pancreas.

Occurrence, stability and decomposition of β-n[γ-l( + )-glutamyl]-4-hydroxymethylphenylhydrazine (agaritine) from the mushroom Agaricus bisporus

A Chromatographic technique was developed that could clearly separate β-N[γ-l(+)-glutamyl]-4-hydroxymethylphenylhydrazine (agaritine) from all other components in 10–500-μl samples of mushroom extracts. Locally purchased mushrooms were found to contain mean levels of 0.4–0.7 mg agaritine/g. The agaritine content of the mushrooms had decreased by 2–47% after I wk of storage in a domestic refrigerator and by 36–76% after 2 wk of such storage. Canned mushroom soup and canned mushrooms did not contain detectable agaritine; a sample of frozen mushrooms contained a mean level of 0.33 mg/g and a batch of fresh mushrooms lost about 32% of their agaritine content on cooking. In mice given 3 mg agaritine by gavage, agaritine was detected in all parts of the gastro-intestinal tract 15 min after dosing, but none was detectable in the gut after 3 hr. The enzyme γ-glutamyltranspeptidase derived from pigs kidney was found to be capable of decomposing agaritine to glutamic acid and 4-(hydroxymethyl)phenylhydrazine, and to have nine times such activity as an enzyme isolated from mushrooms.

Removal of toxic metals and nonmetals from contaminated water

The effects of the application of potassium ferrate to remove possible toxic compounds are presented. Potassium ferrate (K2FeO4) is shown in this work to be an effective means to remove toxic metals and nonmetals from aqueous solution. The toxic material present in water is precipitated from aqueous solution and readily removed. Potassium ferrate removes itself from solution. Discolored contaminated water may be made clear by utilizing potassium ferrate. In addition, turbidities of solutions induced by dissolved substances are eliminated by the action of potassium ferrate. The efficacy of potassium ferrate in cleaning contaminated water shows great potential in application to municipal and industrial waste water.

Effect of pH and ions on the electronic structure of saccharin.

The sodium salt of saccharin is biologically more active as a urothelial cell mitogen in vivo, when fed to male rats, than are the potassium or calcium salts or the acid form, despite similar concentrations of saccharin excreted in the urine. The differences in bladder-mitogenic activity between sodium saccharin and the other salts of saccharin may be the result of known differences in the ionic composition of the urine of rats receiving these various forms of saccharin. These changes in the rat urine following administration of the different salts of saccharin could be responsible for the observed mitogenic responses to oral saccharin; alternatively the differences in the ionic composition of the urine could result in changes in the electronic structure of the saccharin molecule itself, allowing it to be more active in certain ionic environments. Since the pKa of saccharin is 1.8, essentially all of the saccharin in urine (pH greater than 5) will exist in the ionized form. We have used 17O, 15N, 13C and two-dimensional nuclear magnetic resonance (NMR) spectroscopy to explore the electronic structure of the saccharin molecule in aqueous solution. By observing the NMR spectra of the saccharinate ion in the presence of varying concentrations of hydrogen, potassium, sodium, calcium, magnesium, bicarbonate and urate, we have demonstrated that at physiological levels none of these ions significantly alters the electronic structure of the saccharin molecule. Hence the differences in the mitogenic response to the different saccharin salts cannot be explained by alterations in the structure of the saccharin molecule.

Common metabolites of N-nitroso-2,6-dimethylmorpholine and N-nitroso-bis(2-oxopropyl)amine in the Syrian hamster

N-Nitroso-2,6-demethylmorpholine (NDMM) administered intraperitoneally to Syrian hamsters was metabolized to N-nitroso-bis(2-hydroxypropy)(2-oxopropyl)amine(HPOP) and N-nitroso-bis(2-hydroxpropyl)amine (BHP) which were identified in blood and urine. The potent pancreatic carcinogen N-nitroso-bis(2-oxoproply)amine was previously shown to form the same metabolites. Preliminary results indicate that NDMM also induces pancreatic and other tumors in Syrian hamsters, similar to those found following BHP treatment.

Identification of cholesterol as a mouse skin lipid that reacts with nitrogen dioxide to yield a nitrosating agent, and of cholesteryl nitrite as the nitrosating agent produced in a chemical system from cholesterol.

The skin lipids of mice exposed to nitrogen dioxide (NO2) and mouse skin lipids exposed in vitro to NO2 contain nitrosating agents (NSAs), that react with amines to produce nitrosamines. This situation represents a potential hazard of exposure to NO2. A principal NSA precursor in mouse skin lipids was purified by thin-layer and high-performance liquid chromatography. Each fraction was assayed by bubbling in NO2 and determining NSA. The precursor was identified as cholesterol on the basis of its chromatographic behavior and spectral properties. In a chemical system, cholesterol reacted with NO2 to give 13% yields of an NSA, which was identified from its spectral properties as the previously known compound, cholesteryl-3-beta-nitrite. These findings and the chromatographic behavior of a major NSA in the skin lipids of NO2-exposed mice suggested that this NSA was cholesteryl nitrite.

Removal of Nitrosamines from Waste Water by Potassium Ferrate Oxidation

Potassium ferrate (K2FeO4) is useful in the advanced treatment of waste water. Additional evidence of this capability is presented in this study. Potassium ferrate is a very strong oxidant and is highly soluble in water. The nitrosamine studied in this work was toxic and was a potent pancreatic tumorigen in laboratory animals. Nitrosamines, which are potent carcinogens, are widespread throughout the environment and can be eliminated from waste water effluent by the action of potassium ferrate. Potassium ferrate and the nitrosamine was placed in aqueous solution and allowed to react to completion. Analysis by photospectroscopy revealed that the nitrosamine was completely degraded. This result suggests that potassium ferrate is useful for decontamination of some waste water collections.

Immunomodulatory action of levamisole--I. Structural analysis and immunomodulating activity of levamisole degradation products.

In our laboratory we observed that solutions of levamisole (LMS) stored at 4 degrees C consistently enhanced the lymphocyte proliferation response to concanavalin A (Con A) more than freshly prepared solutions did. To determine if the increased immunopotentiation observed with the stored solutions of LMS was due to products formed from LMS, we assessed the stability of LMS when stored at 4 or 37 degrees C at pH 6, 7, 7.5 and 8. Analysis of the various solutions by high pressure liquid chromatography demonstrated that LMS decomposes during storage in neutral and alkaline conditions to form three products. The formation of the products was accelerated by increasing the temperature from 4 to 37 degrees C. The three degradation products were purified by preparative high pressure liquid chromatography and their structures determined by mass spectrometry, infrared spectrometry and homo- and heteronuclear two dimensional nuclear magnetic resonance spectroscopy. The degradation products, denoted as No. 1, No. 2 and No. 3, based on their high pressure liquid chromatography retention times, were identified as: No. 1, 3-(2-mercaptoethyl)-5-phenylimidazolidine-2-one; No. 2, 6-phenyl-2,3-dihydroimidazo (2,1-b) thiazole and No. 3, bis [3-(2-oxo-5-phenylimidazolidin-1-yl) ethyl] disulfide. Product 2 significantly enhanced murine lymphocyte proliferation responses to concanavalin A (Con A) at concentrations between 0.5 and 10.0 micrograms/ml (whereas the optimum concentration of LMS is 10-100 fold higher (50-100 micrograms/ml)). Products 1, 2 and 3 significantly inhibited the lymphocyte proliferative response at concentrations greater than 2.2, 10.0 and 10.0 micrograms/ml, respectively. These studies indicate that under relatively mild conditions, including physiological conditions, LMS may decompose to products which inhibit or enhance lymphocyte responses to Con A.