Sidney S. Mirvish

Role of N-nitroso compounds (NOC) and N-nitrosation in etiology of gastric, esophageal, nasopharyngeal and bladder cancer and contribution to cancer of known exposures to NOC

The questions of whether and how N-nitroso compounds (NOC) may be inducing cancer in humans are discussed. The principal subjects covered include nitrite-derived alkylating agents that are not NOC, reasons for the wide tissue specificity of carcinogenesis by NOC, the acute toxicity of nitrosamines in humans, mechanisms of in vivo formation of NOC by chemical and bacterial nitrosation in the stomach and via nitric oxide (NO) formation during inflammation, studies on nitrite esters, use of the nitrosoproline test to follow human gastric nitrosation, correlations of nitrate in food and water with in vivo nitrosation and the inhibition of gastric nitrosation by vitamin C and polyphenols. Evidence that specific cancers are caused by NOC is reviewed for cancer of the stomach, esophagus, nasopharynx, urinary bladder in bilharzia and colon. I review the occurrence of nitrosamines in tobacco products, nitrite-cured meat (which might be linked with childhood leukemia and brain cancer) and other foods, and in drugs and industrial situations. Finally, I discuss clues from mutations in ras and p53 genes in human tumors about whether NOC are etiologic agents and draw some general conclusions.

Ascorbate-Nitrite Reaction: Possible Means of Blocking the Formation of Carcinogenic N-Nitroso Compounds

The formation of carcinogenic N-nitroso compounds by the chemical reaction between nitrous acid and oxytetracycline, morpholine, piperazine, N-methylaniline, methylurea, and (in some experiments) dimethylamine was blocked by ascorbic acid. The extent of blocking depended on the compound nitrosated and on the experimental conditions. Urea and ammonium sulfamate were less effective as blocking agents. The possibility of in vivo formation of carcinogenic N-nitroso compounds from drugs could be lessened by the combination of such drugs with the ascorbic acid.

Effect of gastroduodenal juice and dietary fat on the development of barrett's esophagus and esophageal neoplasia: An experimental rat model

AbstractBackground: Reflux of duodenal content into the lower esophagus of rats enhances the formation of nitrosamine-induced esophageal cancer and results in the induction of adenocarcinoma. We investigated the extent of the mucosal injury that was produced when the lower esophagus of rats was exposed to the reflux of gastroduodenal juice in the presence or absence of a carcinogen and tested the hypothesis that induction of esophageal cancer in this model would be influenced by the intake of dietary fat. Methods: Esophagoduodenostomy with gastric preservation was performed in 165 Sprague-Dawley rats in order to expose the lower esophagus to the reflux of gastroduodenal juice. Postoperatively selected groups of rats were treated with the carcinogen methyl-n-amylnitrosamine (MNAN). Subsequently, rats were fed diets of differing fat and calorie content for 20 weeks until they were put to death. Results: Refluxed gastroduodenal juice, in the absence of MNAN, induced esophageal inflammatory changes (diffuse papillomatosis and hyperkeratosis) in 38 of 39 rats (97%), specialized columnar metaplasia (Barretts esophagus) in four of 39 (10%), dysplasia in three of 39 (8%), and squamous cell carcinoma in one of 39 (3%). Diet did not influence the incidence of neoplasia in the absence of MNAN treatment. In rats treated with MNAN, refluxed gastroduodenal juice induced inflammation in 110 of 111 rats (99%), columnar metaplasia in 14 of 111 (13%), and cancer in 63 of 111 (57%). Fifty-eight percent of esophageal tumors were squamous cell carcinoma and 42% were adenocarcinoma. The highest incidence of tumors was observed in rats fed the semipurified high-fat diet (24 of 29; 83%) compared with rats fed the semipurified control diet (13 of 29; 45%), semipurified, calorie-restricted diet (15 of 27; 55%), and chow diet (11 of 26; 42%), p<0.05. Conclusions: Reflux of gastroduodenal content into the lower esophagus of rats can induce both Barretts metaplasia and neoplasia. Addition of a carcinogen increases the tumor yield and results in a proportion of the lesions being adenocarcinoma. This carcinogenic process is promoted by a diet with a high fat content.

Total N-Nitroso Compounds and Their Precursors in Hot Dogs and in the Gastrointestinal Tract and Feces of Rats and Mice: Possible Etiologic Agents for Colon Cancer

We review evidence that red and processed meat are causes of colon cancer and that processed meat is a risk factor for childhood cancer and type 2 diabetes. Associations could be due to N-nitroso compounds (NOCs) derived from nitrosation of NOC precursors (NOCPs). We review our survey of total NOC and NOCP content of foods. Only rapidly nitrosated amines, including a glycosyl amino acid, were efficiently determined as NOCPs. NOCPs in hot dogs and rat feces were partly purified by adsorption-desorption and HPLC. After nitrosation, purified hot dog fractions were directly mutagenic in Ames test. The main NOCPs in these materials may be N-glycosyl amino acids and peptides. NOC levels in rat gastrointestinal tract rose steadily from stomach to feces. NOCP levels showed similar trend but with sharp increases from stomach to duodenum. One day after Min and C57BL/6J mice were fed 4% dextran sulfate sodium to induce acute colitis, fecal NOC levels increased 1.9-fold compared with untreated mice (P < 0.05). For 7 d Swiss mice received semipurified diet, 180 g beef-pork hot dogs mixed with 820 g diet or 180 g sautéed beef mixed with 820 g diet. Fecal NOC outputs on day 7 were 3.7-5.0 (hot dog) and 2.0-2.9 (beef) times those for control groups (P < 0.002 for combined groups), perhaps reflecting higher dietary NOC intakes. Feeding a similar hot dog mixture to mice did not affect normal 7-methyldeoxyguanosine level in colonic mucosal DNA. Overall, results support the hypothesis that colonic NOCs are a cause of colon cancer.

Calcium and α-tocopherol suppress cured-meat promotion of chemically induced colon carcinogenesis in rats and reduce associated biomarkers in human volunteers

BACKGROUND Processed meat intake has been associated with increased colorectal cancer risk. We have shown that cured meat promotes carcinogen-induced preneoplastic lesions and increases specific biomarkers in the colon of rats. OBJECTIVES We investigated whether cured meat modulates biomarkers of cancer risk in human volunteers and whether specific agents can suppress cured meat-induced preneoplastic lesions in rats and associated biomarkers in rats and humans. DESIGN Six additives (calcium carbonate, inulin, rutin, carnosol, α-tocopherol, and trisodium pyrophosphate) were added to cured meat given to groups of rats for 14 d, and fecal biomarkers were measured. On the basis of these results, calcium and tocopherol were kept for the following additional experiments: cured meat, with or without calcium or tocopherol, was given to dimethylhydrazine-initiated rats (47% meat diet for 100 d) and to human volunteers in a crossover study (180 g/d for 4 d). Rat colons were scored for mucin-depleted foci, putative precancer lesions. Biomarkers of nitrosation, lipoperoxidation, and cytotoxicity were measured in the urine and feces of rats and volunteers. RESULTS Cured meat increased nitroso compounds and lipoperoxidation in human stools (both P < 0.05). Calcium normalized both biomarkers in rats and human feces, whereas tocopherol only decreased nitro compounds in rats and lipoperoxidation in feces of volunteers (all P < 0.05). Last, calcium and tocopherol reduced the number of mucin-depleted foci per colon in rats compared with nonsupplemented cured meat (P = 0.01). CONCLUSION Data suggest that the addition of calcium carbonate to the diet or α-tocopherol to cured meat may reduce colorectal cancer risk associated with cured-meat intake. This trial was registered at clinicaltrials.gov as NCT00994526.

Effect of sodium ascorbate on tumor induction in rats treated with morpholine and sodium nitrite, and with nitrosomorpholine

Groups of male MRC Wistar rats were treated for 2 years either with morpholine (10 g/kg food) together with sodium nitrite (3 g/l drinking water) or with N-nitrosomorpholine (NM, 0.15g/l drinking water). In both cases, a group of rats was given sodium ascorbate (22.7 g/kg food) in addition to these treatments. When ascorbate was present, the liver tumors induced by morpholine and nitrite showed a 1.7-fold longer induction period, a slightly lower incidence, and an absence of metastases in the lungs, indicating that ascorbate had inhibited the in vivo formation of NM. Ascorbate did not affect liver tumor induction by the performed NM. The group treated with morpholine, nitrite, and ascorbate had a 54% incidence of forestomach tumors, including an 18% incidence of squamous cell carcinomas, possibly because ascorbate promoted NM action in this organ.

N-NITROSO COMPOUNDS, NITRITE, AND NITRATE: POSSIBLE IMPLICATIONS FOR THE CAUSATION OF HUMAN CANCER

SUMMARY Cancer incidences show wide geographic variation and it is likely that chemical carcinogens are responsible for most human cancer. N -Nitroso- (NNO-) compounds cause cancer in experimental animals and could be involved in human cancer, but the level of NNO-compounds so far found in food is generally small. NNO-compounds are readily formed from nitrite and amines or amides, and the kinetics of this reaction are discussed. Tumors and other toxic effects were produced in rodents by feeding amines or amides plus nitrite (but not nitrate). Also, bacteria may catalyze NNO-compound formation. In vivo formation of NNO-compounds could occur in the stomach and at several other in vivo sites. The possibility that NNO-compounds cause human cancer of the nasopharynx, esophagus, and stomach is discussed, especially preliminary evidence that areas of high nitrate intake may show a raised incidence of gastric cancer.

Supercritical fluid technology based large porous celecoxib–PLGA microparticles do not induce pulmonary fibrosis and sustain drug delivery and efficacy for several weeks following a single dose

Although pulmonary dosing of large porous particles has been shown to sustain drug delivery for a few days, there are no reports on safety or long term delivery. In this study we prepared large porous poly(lactide-co-glycolide) (PLGA) microparticles of celecoxib using supercritical fluid pressure-quench technology and demonstrated 4.8-, 15.7-, and 2.1-fold greater drug levels in lung, bronchoalveolar lavage fluid (BAL), and plasma compared to conventional microparticles on day 21 after a single intratracheal dosing of dry powders in A/J mice. Porous particle based delivery was 50.2-, 95.5-, and 7.7-fold higher compared to plain drug in the lung, BAL, and plasma, respectively. Toxicity of the formulations was assessed on day 21 following a fibrosis assessment protocol in A/J mice. There was no significant change in lactate dehydrogenase (LDH), total protein, and total cell counts in the BAL, and soluble collagen levels in the lung tissue following particle or drug treatments. Lung histology indicated no significant hyperplasia, granuloma, or collagen deposition in the treated groups. Chemopreventive potential of celecoxib porous particles was assessed in a benzo[a]pyrene (B[a]P) induced lung cancer model in A/J mice, on day 60 following a single intratracheal dose with or without single intravenous paclitaxel/carboplatin treatment. The combination group was more effective than individual groups, with the inhibition of tumor multiplicity and reduction of vascular endothelial growth factor in the BAL being 70 and 58%, respectively. Thus, large porous celecoxib-PLGA microparticles prepared using supercritical fluid technology exhibited sustained drug delivery and anti-tumor efficacy, without causing any significant toxicity.

Inhibition by allyl sulfides and phenethyl isothiocyanate of methyl-n-pentylnitrosamine depentylation by rat esophageal microsomes, human and rat CYP2E1, and rat CYP2A3

Garlic and Cruciferae are associated with reduced risks of several human cancers, and some of their constituents are anticarcinogenic in animals. Here we studied inhibition of in vitro metabolism of the rat esophageal carcinogen methyl-n-pentylnitrosamine (MPN) by garlic-derived allyl sulfides and by Cruciferae-derived phenethyl isothiocyanate (PEITC) and sulforaphane. The test inhibitors were incubated with [3H]-MPN, NADPH-generating system and rat esophageal microsomes (REM) or a cytochrome P450 (CYP). [3H]-MPN activation by depentylation was assayed by HPLC with radiometric determination of [3H]-pentaldehyde 2,4-dinitrophenylhydrazone. IC50 for depentylation of 40 μM MPN by rat CYP2E1 was 5-12 μM for diallyl sulfide (DAS), diallyl disulfide (DADS), and PEITC and 10-20 μM for diallyl sulfone, allyl mercaptan, and diallyl trisulfide. Maximum inhibition required preincubation of rat CYP2E1 with DAS for 15 min and with DADS for 30 min. Using these preincubation times, Ki for MPN depentylation by REM, rat and human CYP2E1, and rat CYP2A3 was 0.6-1.6 μM for inhibition by DAS and 1.7-70 μM for inhibition by DADS. With PEITC, Ki for MPN depentylation by REM, rat CYP2E1, and rat CYP2A3 was 0.4-4.6 μM. These low Ki and IC50 values may help explain how garlic and Cruciferae inhibit carcinogenesis.

Effect of duodenal components of the refluxate on development of esophageal neoplasia in rats

When duodenal content is allowed to reflux into the esophagus of nitrosamine-treated rats, esophageal cancer is induced more rapidly and at higher frequency than after carcinogen treatment alone. The purpose of the present study was to identify the components of the duodenal content that are responsible for enhancing esophageal carcinogenesis. Eight-week-old Sprague-Dawley rats underwent one of four operations as follows: diversion of bile alone, pancreatic juice alone, both bile and pancreatic juice into the esophagus, or a control operation with no induced reflux. Two weeks after surgery, rats were treated with the esophageal carcinogen 2,6-dimethylnitrosomorpholine (48 mg/kg [0.1 of LDs0] intraperitoneally weekly for 20 weeks). The rats were killed at age 30 weeks. The esophagus was removed and full-length strips were examined under a microscope; separate segments were taken for flow cytometric evaluation. The prevalence of DNA aneuploidy and histologic esophageal papillomas or squamous cancer was increased in carcinogen-treated rats with pancreatic juice reflux (P <0.05 vs. control) and the combination of pancreatic and bile reflux (P <0.05 vs. control) but not in rats with bile reflux alone. We conclude that pancreatic juice is the most potent component of the duodenal refluxate in the promotion of esophageal carcinogenesis in rats.