Production of mouse urinary bladder carcinomas by sodium cyclamate.

Sodium cyclamate was suspended in cholesterol pellets that were surgically implanted in the urinary bladders of mice. In duplicate experiments, incidences of mouse bladder carcinomas observed in animals exposed to these pellets were 78 and 61 percent compared with incidences of 13 and 12 percent in control mice exposed to pellets of pure cholesterol. The exposure of the mouse bladder to sodium cyclamate was very brief, as the time required for 50 percent of the compound to disappear from the pellets was about 1 hour. This experimental technique was found to be highly sensitive, reproducible, and predictive of the bladder carcinogenicity of orally administered cyclamate.

Production of urinary bladder carcinomas in mice by sodium saccharin.

Pellets weighing 20 to 24 milligrams and containing 20 percent sodium saccharin suspended in cholesterol were surgically implanted into the urinary bladder lumens of female Swiss mice (60 to 90 days old) under ether anesthesia. Incidences of mouse bladder carcinomas in animals exposed to these pellets were 47 and 52 percent as compared with incidences of 13 and 12 percent in control mice exposed to pellets of pure cholesterol. The exposure of the mouse bladder to saccharin was very brief, because the time required for 50 percent of the compound to be eluted from the pellets was about 5.5 hours.

Aldosterone hypersecretion in "non-salt-losing" congenital adrenal hyperplasia.

Patients with the "non-salt-losing" form of the adrenogenital syndrome were studied before and after suppression of adrenal cortical activity with carbohydrate-active steroids. The response of aldosterone secretion to sodium deprivation was measured; in some patients response to adrenocorticotropic hormone (ACTH) was measured as well. The aldosterone secretion was normal and responded normally to sodium deprivation in all patients studied during suppression with carbohydrate-active steroids. This finding suggests that 21-hydroxylation of progesterone is normal in this syndrome. The sole abnormality in the production of aldosterone in these patients was found to be excessive secretion of aldosterone while they were not receiving suppressive doses of carbohydrate-active steroids. This finding strongly supports the view that the biogenetic pathways through which aldosterone is produced from progesterone are intact in this syndrome. No patient showed hypertension or hypokalemic alkalosis despite very high aldosterone secretion rates. This observation suggests that the hyper-aldosteronism is secondary to a tendency to sodium loss in the patient whose ACTH production is not suppressed. These studies provide additional evidence in support of the hypothesis that the salt-losing and "non-salt-losing" forms of adrenogenital syndrome are genetically and biochemically distinct.

Comparative carcinogenicity of 5-nitrothiophenes and 5-nitrofurans in rats.

We investigated the carcinogenicity of five 5-nitrothiophenes with heterocyclic substituents at the 2-position of the thiophene ring by feeding the chemicals to Sprague-Dawley rats and comparing the type and incidence of lesions with those appearing after exposure to two 5-nitrofurans. Benign and malignant mammary tumors and intestinal tract sarcomas were the most frequent lesions induced by 5-nitrothiophenes. 4-Bis(2-hydroxyethyl)amino-2-(5-nitro-2-thienyl)quinazoline caused a 100% incidence of mammary adenocarcinomas in 28 female rats at risk; it induced 3 benign and 5 malignant mammary tumors and 13 small intestine sarcomas in 20 male rats. A high incidence of similar lesions was observed in male and female rats fed the corresponding 5-nitrofuran analogue, 4-bis(2-hydroxyethyl)amino-2-(5-nitro-2-furyl)quinazoline. In marked contrast, 4 of 28 female rats receiving 4-bis(2-hydroxyethyl)amino-2-(2-thienyl)quinazoline, which lacks the nitro group at the 5-position on the thiophene ring, had solitary benign mammary tumors (P greater than 0.2). Additional 5-nitrothiophenes demonstrating significant oncogenic activity for female rats were 4-morpholino-2-(5-nitro-2-thienyl)quinazoline, 4-(2-hydroxyethylamino)-2-(5-nitro-2-thienyl)quinazoline 4-(2,3-dihydroxypropylamino)-2-(5-nitro-2-thienyl)quinazoline, and 1,2-dihydro-2-(5-nitro-2-thienyl)quinazolin-4(3H)-one. Another nitrofuran, 4,6-dimethyl-2-(5-nitro-2-furyl)-pyrimidine, provided the following types of neoplasms in 30 female rats at risk: squamous cell carcinomas of the forestomach (30), sarcomas of the intestine (21), adenocarcinomas of the kidney (2).

Correlation between the carcinogenicities of nitrofuran derivatives and their destructive actions on sebaceous glands of mouse skin.

The effects of six nitrofuran derivatives (including a formerly used food preservative) on mouse skin sebaceous glands were investigated. A close correlation was found between the carcinogenicities and destructive activities of nitrofuran derivatives on the sebaceous glands. 5-Nitro-2-furaldehyde semicarbazone and 4-methyl-1-[(5-nitrofurfurylidene)amino]-2-imidazolidinone, which are carcinogenic, caused marked destruction of the glands at a dose of 1-5 mg/mouse. 2-(5-Nitro-2-furfurylidene)-aminoethanol almost completely destroyed the glands at a dose of 5 mg/mouse; its carcinogenicity has not yet been investigated. 1-[(5-Nitrofurfurylidene)amino]-carcinogenic, did not affect the glands, even at a dose of 5 mg/mouse. 2-(2-Furyl)-3-(5-nitro-2-furyl)acrylamide, which is a potent mutagen but not carcinogenic, had no effect on the glands at a dose of 5 mg/mouse. Under similar conditions, the potent carcinogen 7,12-dimethylbenz(alpha)athracene almost completely destroyed the sebaceous glands at a dose of 0.05 mg/mouse, but dimethyl sulfoxide (used as solvent for the test compounds) had no effect.

Mutagenicity for Salmonella typhimurium of urine obtained from humans receiving nitrofurantoin.

Urine samples from 12 humans receiving oral therapeutic doses of nitrofurantoin were mutagenic for Salmonella typhimurium strain TA 100 and nonmutagenic for strain TA 100-FR1. Mutagenic activity of the urine was not increased by treatment with beta-glucuronidase. Spot mutation assay of the chromatogram of urine revealed that the mutagenic activity of the urine was mainly due to unmetabolized nitrofurantoin.

Carcinogenicity of tannin and tannin-free extracts of bracken fern (Pteridium aquilinum) in rats.

F344 inbred and Sprague-Dawley noninbred rats were fed a basic diet (groups 1 and 7) or a basic diet supplemented with 0.1% (later, 0.2 and 0.4%) tannin (group 2) isolated from bracken fern (Pteridium aquilinum) (BF), 33% BF (groups 3 and 6), 2% chloroform fraction of BF (group 4), or 4% tannin-free fraction of BF (group 5). The following incidences of intestinal or bladder tumors were observed: group 1, intestinal and bladder, 0/16; group 2, 0/21; group 7, 0/16; groups 4 and 5, intestinal, 7/15, bladder, 0/15; group 3, intestinal, 19/20, bladder, 12/20; and group 6, intestinal, 22/30, bladder, 15/30. The chloroform-methanol fraction prepared from urine of rats fed BF, chloroform fraction of BF, or tannin-free fraction of BF demonstrated mutagenicity for Salmonella typhimurium TA 100 but not for TA 98. No mutagenicity was detected in other prepared fractions. F344 rats in group 8 received weekly sc injections of tannin solution (0.1 g/kg body wt) for 38 weeks, and 16/20 developed palpable tumors that were malignant fibrous histiocytomas at the injection site. No tumor was present in any rat of control group 9.

Identification of carcinogenic tannin isolated from Bracken fern (Pteridium aquilinum).

We attempted to isolate a carcinogenic substance from bracken fern (Pteridium aquilinum), a naturally occurring toxicant responsible for the production of chronic enzootic hematuria and urinary bladder cancer of cattle and carcinogenic for various target organs of several species. Hot methanol extracts of bracken fern were solubilized in water and extracted with chloroform followed by a mixture of n-butanol-butanone (1:1). That fraction was dried and triturated with ether-methanol (4:1), n-butanol, and finally absolute ethanol. The insoluble residue was dissolved in 10% aqueous methanol and passed through Dowex 1 OH-, Dowex 50 H+, or Dowex 1 OH- and then Dowex 50 H+ ion exchange resins. A condensed tannin, isolated from one ot the fractions, was identical to that isolated from bracken fern by the caffeine procedure used for the separation of tannins from other plant constituents. Three systems were used for bioassay; induction of bladder carcinoma by implantation of cholesterol pellets containing bracken fern fractions into the bladder lumens of mice; acute toxicity by ip injection of brachen fern fraction into mice; and growth inhibition of Escherichia coli. The following fractions induced significantly greater incidences of bladder carcinoma than did cholesterol pellets only: tannin, Dowex 50 H+, residue, n-butanol, and methanol. Tiliroside, a component of bracken fern fractions into the bladder lumens of mice; acute genic acid, and quercetin were not carcinogenic. Tannin was the most toxic (mean lethal dose: 0.16 mg/g) and carcinogenic. None of the carcinogenic fractions inhibited growth of E. coli.

Carcinogenicity of the antineoplastic agent, 5-(3,3-dimethyl-1-triazeno)-imidazole-4-carboxamide, and its metabolites in rats.

Chronic oral administration of the antineoplastic agent, 5-(3,3-dimethyl-1-triazeno)imidazole-4-carboxamide (NSC-45388, DTIC), induced predominantly thymic and mammary tumors as demonstrated previously. Male and female Sprague-Dawley and female Buffalo rats were susceptible to the carcinogenicity of DTIC. A 50% incidence of mammary adenocarcinomas was induced in males within 18 weeks. Type of tumor and tumor incidence were dose dependent. Single and multiple intraperitoneal injections of DTIC did not alter organ specificity. DTIC-induced thymic lymphosarcomas and mammary adenocarcinomas were transplantable. Tissue distribution studies revealed no correlation between uptake of DTIC by a given tissue and its susceptibility to carcinogenicity. Metabolites of DTIC were tested for carcinogenic activity. Animals administered 5-diazoimidazole-4-carboxamide orally, intraperitoneally, or intragastrically developed low incidences of thymic, stomach, bladder, or mammary tumors. A low incidence of mammary tumors developed in rats fed 2-azahypoxanthine. A variety of tumors, including several ependymoblastomas, were induced in rats that received 5-aminoimidazole-4-carboxamide orally. 5-(3-Methyl-1-triazeno)imidazole-4-carboxamide (MTIC), when fed or given in single or multiple intraperitoneal injections, induced a high incidence of mammary adenofibromas and a low incidence of uterine leiomyosarcomas. Control rats had low incidences of mammary adenocarcinomas and adenofibromas after 52 weeks. These data show that the carcinogenic properties of DTIC resemble those of carcinogenic N-nitroso compounds, hydrazine, azo, and azoxy-alkanes and aryltriazenes and thus suggest similar mechanism(s) of action. These data also indicate that MTIC is involved in the induction of mammary adenofibromas and uterine leiomyosarcomas by DTIC.

Effect of age, sex, and intestinal flora on the induction of colon tumors in rats.

Germfree and conventional Sprague-Dawley rats were assessed for their susceptibility to intrarectally injected N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) or N-methyl-N-nitrosourea (MNU). Adenocarcinoma of the colon was induced in germfree and conventional rats by both MNNG and MNU. The colons of germfree rats were more susceptible to the direct-acting carcinogens, as manifested by earlier morbidity and development of colon tumors (50% tumors within 30-35 wk), than were those of conventional rats (50% colon tumors within 48-50 wk). Germfree and conventional male rats were more susceptible to the carconogens than were their female germfree and conventional counterparts. Young (30 days old at the start of the experiment) germfree rats developed colon tumors more quickly (15-20 wk) than did older (60 days) germfree rats after intrarectal injections of MNNG. No colon tumors were observed in germfree and conventional rats after 75 weekly intrarectal injections with a buffer. Transplantation of an adenocarcinoma induced with MNU in a female rat to germfree and conventional rats showed that it was easily transplantable, required no immunosuppression, and had essentially the same morphologic characteristics as did the primary tumor.

The pathogenesis of experimental bladder cancer.

The pathogenesis of signal morphological lesions of the urinary bladder induced in several species following administration of N-butyl-N-(4-hydroxybutyl)nitrosamine, bracken fern, or N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide is presented. Incidences of bladder neoplasia exceeding 80% were generated in the rat by each compound. Bladder neoplasia was induced in the following species by each substance: by N-butyl-N-(4-hydroxybutyl)nitrosamine in the mouse, hamster, guinea pig, and dog; by bracken fern in the guinea pig, mouse, and cow; and by N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide in mouse, hamster, and dog. The guinea pig appeared resistant to the bladder oncogenicity of N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide. Different species displayed a gradient of bladder neoplastic responsiveness. Hyperplasia was a consistent early lesion and was usually focal. Early hyperplastic lesions regressed following removal of the carcinogenic stimulus, but later lesions appeared to be irreversible. These animal systems appear useful in providing opportunities for investigations relevant to human bladder cancer.

Effect of p-hydroxyacetanilide, sodium sulfate, and L-methionine on the leukemogenicity of N-[4-(5-nitro-2-furyl)-2-thiazolyl]acetamide.

Dietary administration of N-[4-(5-nitro-2-furyl)-2-thiazolyl]acetamide to mice for 14 weeks followed by 16 weeks of control diet resulted in a high incidence of lymphocytic leukemia and a low incidence of forestomach squamous cell papillomas. The coadministration of p-hydroxyacetanilide at a dose of 1.0% with either 250 or 500 ppm of N-[4-(5-nitro-2-furyl)-2-thiazolyl]acetamide resulted in inhibition of leukemogenesis, whereas when p-hydroxyacetanilide was coadministered with 1000 ppm of N-[4-(5-nitro-2-furyl)-2-thiazolyl]acetamide the leukemia incidence was not significantly reduced, but the latent period was prolonged. When sodium sulfate was administered with p-hydroxyacetanilide and N-[4-(5-nitro-2-furyl)-2-thiazolyl]acetamide, leukemogenesis was partially restored. L-Methionine, fed in place of sodium sulfate, unblocked leukemogenicity inhibition by p-hydroxyacetanilide. None of these chemicals, p-hydroxyacetanilide, sodium sulfate, or L-methionine, significantly affected the incidence of forestomach papillomas induced by N-[4-(5-nitro-2-furyl)-2-thiazolyl]acetamide, although tumor incidences in all groups were low. p-Hydroxyacetanilide and sodium sulfate had no significant effect on the high incidence of stomach tumors induced by formic acid 2-[4-(5-nitro-2-furyl)-2-thiazolyl]hydrazide or bladder tumors induced by N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide.

Biotransformation of the bladder carcinogen N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide in mice.

The biotransformation of N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide (FANFT), a potent urinary bladder carcinogen, was studied in mice. About 82% of radioactivity was excreted as 14CO2 within 36 hr after intragastric administration of N-[4-(5-nitro-2-furyl)-2-thiazolyl]-[14C]formamide, suggesting its deformylation to 2-amino-4-(5-nitro-2-furyl)thiazole ( ANFT ). The latter was formed in vitro as a product following incubation of FANFT with mouse liver homogenates. Chromatographic analysis of mouse urine obtained 24 hr after the i.p. administration of N-[4-(5-nitro-2-furyl)-[2-14C]thiazolyl]formamide revealed excretion of ANFT and unmetabolized FANFT, suggesting the prevalence of the deformylation reaction in vivo. In addition, at least two more metabolites were present in urine. One of these metabolites exhibited chromatographic properties similar to those exhibited by a compound derived from the in vitro nitroreduction of ANFT . This metabolite was isolated from urine of FANFT-fed animals and from in vitro enzymatic reduction of ANFT with mouse liver homogenates. The isolated products had chromatographic and spectral properties and a mass spectral fragmentation pattern similar to that of a compound obtained by catalytic reduction of ANFT with palladium and activated carbon. Spectroscopic analyses established the structural identity of the chemical reduction product as 1-[4-(2-aminothiazolyl)]-3-cyano-1-propanone ( ATCP ). Since the chromatographic properties of the enzymatically derived product and the urinary metabolite were identical to those of a compound obtained by chemical reduction, they must be structurally the same and thus correspond to ATCP . About 5% of the urinary metabolites of FANFT is ATCP , and thus ATCP is quantitatively a minor excretory product. ATCP was far less active than was ANFT of FANFT in the Ames mutagenicity assay with Salmonella typhimurium TA.

Early induction of mouse urinary bladder ornithine decarboxylase activity by rodent vesical carcinogens.

The responses of mouse urinary bladder ornithine decarboxylase (EC 4.1.1.17) and S-adenosyl-L-methionine decarboxylase (EC 4.1.1.50) activities were studied following topical intravesical administration of N-[4-(5-nitro-2-furyl)-2-thiazolyl]-formamide (FANFT) or 2-amino-4-(5-nitro-2-furyl)thiazole (ANFT), potent rodent bladder carcinogens. A single bladder topical application of ANFT or FANFT resulted in a significant increase over controls of ornithine decarboxylase activity within 5 hr, with a return to control levels by 10 hr. S-Adenosyl-L-methionine decarboxylase activity demonstrated a lesser response to topical ANFT or FANFT, achieving a level 2 or 3 times that of controls at 5 to 8 hr, followed by a gradual decline to control levels. Stimulation of activities of both enzymes was dose dependent over a range of 4.6 to 460 nmol of ANFT. ANFT-induced ornithine decarboxylase activity was principally localized in the bladder epithelium and was inhibited in a linear dose-response relationship by the synthetic retinoid, 13-cis-retinoic acid. Mice given FANFT p.o. demonstrated a significant increase over controls in ornithine decarboxylase activity within 12 hr, followed by a gradual decline to control levels by 72 hr.

Specific binding, stimulation of rodent urinary bladder epithelial ornithine decarboxylase, and induction of transitional cell hyperplasia by the skin tumor promoter 12-O-tetradecanoylphorbol-13-acetate.

The effects of intraurethral or i.p. administration of a mouse skin tumor promoter phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA), on rodent urinary bladder transitional epithelium were studied. TPA, when instilled into the urinary bladder of inbred rats (female Fischer, F344) or mice (C3H, ICR, C57BL X DBA/2 F1) at a dose as low as 0.16 nmol, led to a significant (about 10-fold) increase in bladder ornithine decarboxylase (EC 4.1.1.17) (ODC) activity. Peak ODC activity was observed at about 6 hr, and enzyme activity returned to base levels about 14 hr after intravesical TPA. Administration of TPA i.p. in dimethyl sulfoxide also induced vesical ODC at 4 hr after treatment. The magnitude of vesical ODC induction correlated well with the ability of a series of phorbol esters to promote mouse skin tumor formation (TPA greater than phorbol didecanoate greater than phorbol dibenzoate, and phorbol diacetate or phorbol did not induce bladder ODC activity). Mezerein, a second stage mouse skin tumor promoter, induced urinary bladder ODC as much as TPA did. Increased ODC activity by TPA was the result of an increased amount of ODC protein localized mostly (greater than 60%) in urinary bladder mucosa. Intraurethrally administered TPA induced transitional cell hyperplasia starting at Day 2, and it persisted for about 7 days. The urothelium regained normal histology 13 days after TPA treatment. TPA bound specifically and with high affinity to murine bladder mucosa and muscularis particulate preparations. Scatchard analysis of mucosal binding revealed a Kd of 0.82 nM; at saturation, 2.43 pmol were bound per mg protein. Since TPA binds specifically to urinary bladder epithelium, and the induction of ODC activity is one of the properties of tumor promoters, one may conclude that TPA may promote urinary bladder carcinogenesis. Intravesical saccharin also induced urinary bladder ODC activity, but TPA at equimolar quantity was far more potent than saccharin. Thus TPA, being a structurally well-defined molecule, may be a useful compound to study the phenomenon of the tumor promotion stage in urinary bladder carcinogenesis.

Mutagenicity of nitrofurans, nitrothiophenes, nitropyrroles, nitroimidazole, aminothiophenes, and aminothiazoles in Salmonella typhimurium.

Thirty-two heterocyclic compounds, including 24 nitroheterocycles, 7 aminoheterocycles and derivatives, and 1 thiophene lacking a nitro group, were tested for mutagenic activity in Salmonella typhimurium TA 98 and TA 100. All the nitroheterocycles (11 new), including nitrofurans, nitrothiophenes, nitropyrroles, and 1 nitroimidazole, were mutagenic in TA 100; 13 were also mutagenic in TA 98. 5-Nitro-2-furoic acid, a noncarcinogen, was mutagenic in TA 100. Seven carcinogenic nitroheterocycles were mutagenic in both strains. Seven aminoheterocycles (4 new), aminothiophenes and aminothiazole derivatives, and 1 thiophene without a nitro group were not mutagenic. Both TA 98 and TA 100 were uvrB and lacked the ability of excision repair of DNA. Among the 24 mutagenic nitroheterocycles, only 13 compounds exhibited bacterial killing effects, suggesting that more than 1 mechanism may be involved in the interaction of nitroheterocycles with bacterial DNA.

Early induction of rat colonic epithelial ornithine and S-adenosyl-L-methionine decarboxylase activities by N-methyl-N'-nitro-N-nitrosoguanidine or bile salts.

The responses of male noninbred rat colonic epithelial ornithine decarboxylase (EC 4.1.1.17) (ODC) and S-adenosyl-L-methionine decarboxylase (EC 4.1.1.50) (SAMD) activities following topical administration of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) or bile salts were studied. A single intrarectal installation of 13 mumol of MNNG resulted in a significant (p < 0.001) 20-fold peak ODC activity after 4 hr, with a prompt return to control levels by 12 hr. Stimulation of SAMD activity was less pronounced but significant (p < 0.01), with a broad 2-fold peak over controls. No significant responses of colonic epithelial enzyme activities were detected following a single intrarectal instillation of N-methyl-N'-nitroguanidine, a noncarcinogenic and nonmutagenic metabolite of MNNG, at a dose equimolar to that of MNNG. Bile salts significantly (p < 0.001) induced ODC with almost the same kinetic pattern as that observed after MNNG administration in the following order: sodium deoxycholate > sodium chenodeoxycholate > sodium cholate. Activations of SAMD were similar for these 3 bile salts. Glycine- or taurine-conjugated deoxycholate showed ODC and SAMD enzyme activations similar to that of nonconjugated deoxycholate. No significant enzyme response was seen after sodium dehydrocholate treatment. Stimulation of activities of both enzymes was directly dependent on bile salt dose. Induced ODC and SAMD activities were principally localized in colonic epithelium. Deoxycholate-stimulated enzyme activities were significantly inhibited by cycloheximide. Enzyme stimulations by active compounds were accompanied by morphological changes such as mucosal cell degeneration, mucus depletion, submucosal congestion, and punctate hemorrhage, followed by submucosal leukocytic cellular infiltration. These data support the concept that initiating and promoting events may be involved in colon carcinogenesis.

Mutagenicity, carcinogenicity, distribution, and nitroreduction of 4-(5-nitro-2-furyl)thiazole in the rat.

Albino noninbred weanling female Sprague-Dawley rats were fed a powdered basic grain diet (Group 1) or a basic diet supplemented with 1540 ppm of 4-(5-nitro-2-furyl)thiazole (NFT) (Group 2). Group 2 rats consumed an estimated mean NFT cumulative dose of 42 mmol/rat, exhibited significant growth retardation and hepatomegaly, and displayed 46 neoplasms (24 multiple mammary fibroadenomas, 19 forestomach squamous cell carcinomas, and 3 other malignant tumors) in 31 of 35 rats histologically evaluated. Six of 36 control rats had solitary, benign mammary fibroadenomas. After p.o. administration of NFT, extraction of urine with chloroform:diethyl ether followed by gas chromatography provided a major peak with a retention time of about 4 min. Catalytic hydrogenation of NFT with palladium on activated carbon afforded a product with the same retention time. The isolated urinary metabolite of NFT exhibited mass spectral fragmentation patterns and gas and high-pressure liquid chromatographic retention times similar to those of the chemical reduction product. These data demonstrate the identical chemical characteristics of the in vivo urinary metabolite of NFT and the compound obtained by chemical reduction of NFT. Spectroscopic analyses established the structural identity of this reduced product as 1-(4-thiazolyl)-3-cyano-1-propanone. Forty-eight hr after the intragastric administration of [14C]NFT, 32% of radioactivity was recovered in urine, 57% was recovered in gastrointestinal contents and feces, and 5.5% was recovered in expired 14CO2. About 2% of the urinary radioactivity was extracted in chloroform:diethyl ether, suggesting that 1-(4-thiazolyl)-3-cyano-1-propanone is quantitatively a minor urinary metabolite of NFT. 1-(4-Thiazolyl)-3-cyano-1-propanone was 1.1 x 10(4)-fold less active than was NFT in the Ames mutagenicity assay with Salmonella typhimurium TA 100.

Effect of avitaminosis A and hypervitaminosis A on urinary bladder carcinogenicity of N-(4-(5-Nitro-2-furyl)-2-thiazolyl)formamide.

The effect of vitamin A deficiency and hypervitaminosis A on the urothelial carcinogenicity of N-[4-(5-nitro-2-furyl)-2-thiazolyl]formanmide (FANFT) was determined in female weanling Sprague-Dawley rats. Vitamin A deficiency resulted in squamous metaplasia of the urinary bladder and high incidences of cystitis, ureteritis, and pyelonephritis. Administration of FANFT to vitamin A-deficient rats appeared to accelerate the carcinogenic process, with earlier appearance of urinary bladder tumors and the development of ureteral and renal pelvic carcinomas. Most of these tumors were squamous cell, occasionally with transitional cell foci. Hypervitaminosis A prevented the appearance of squamous metaplasia and squamous cell neoplasia in rats fed FANFT, but it did not inhibit the formation of transitional cell hyperplasia or neoplasia in comparison to rats receiving normal levels of vitamin A and FANFT.

Suppression of antibody-mediated and cell-mediated murine immunity by the carcinogen N-[4-(5-nitro-2-furyl)-2-thiazolyl]acetamide.

N-[4-(5-Nitro-2-furyl)-2-thiazolyl]acetamide (NFTA) administered at 1000 ppm in diet to mice for 12 weeks induced a high incidence of lymphocytic leukemia. Effects of NFTA on antibody-mediated immunity and cell-mediated immunity of BALB/c mice were studied using the spleen plaque assay for detection of immunoglobulin M-producing cells and the graft-versus-host (GVH) reaction, respectively. NFTA suppressed both responses. With the spleen plaque assay, the number of antibody-forming cells (AFC) to sheep red blood cells was significantly less than in unmedicated, control mice after treated mice received NFTA at 1000 ppm for 6 days. The GVH reaction was not suppressed at 21 days, but was severely suppressed at 70 days, prior to the histological appearance of leukemia. Effect of dose was studied by administering NFTA at 100, 250, 500, and 1000 ppm of diet for 13 to 14 weeks and then determining the response in the spleen plaque assay and GVH reactions. The ratio of AFC/spleen of NFTA-treated groups to AFC/spleen of an unmedicated control group, at the above specified doses, was 0.86, 0.22, 0.33, and 0.54 in ascending dosage order beginning with 100 ppm. For the GVH reaction, the suppression of the cell-mediated immunity was directly proportional to the dose of NFTA. Suppression of the antibody-mediated immunity in relation to the induction of leukemia at 28 weeks was studied by feeding NFTA at 500 ppm for 14 weeks, followed by unmedicated diet for 14 weeks. During the 11th week, mice were immunized with SRBC; 5 days later the spleens were removed and the spleen plaque assay was performed. Eight of 18 mice fed NFTA developed leukemia. The number of AFC/spleen was 78 X 10(3) +/- 34 for those with leukemia and 68 X 10(3) +/- 24 (p greater than 0.5) for those without leukemia, compared with 170 X 10(3) +/- 74 for the control mice (p less than 0.01 for both groups, compared with controls). A closely related carcinogenic nitrofuran, N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide, did not suppress the antibody-mediated immunity response measured during the 11th week of administration.