DNA damage in peripheral blood lymphocytes of patients with cirrhosis related to alcohol abuse or to hepatitis B and C viruses.

BACKGROUND: Both alcohol abuse and hepatitis B or C virus infections are implicated in the development of hepatocellular carcinoma, but it is still controversial whether the pathogenetic mechanism is epigenetic or genotoxic. AIM: Considering that alcohol promotes the generation of reactive oxygen species and both viruses infect peripheral lymphocytes, in this study we investigated the occurrence of DNA fragmentation in peripheral blood lymphocytes from patients with alcoholic cirrhosis and from patients with cirrhosis related to B and C viruses, and analyzed the correlation between the degree of DNA fragmentation and the Child-Pugh score used to assess the degree of hepatic insufficiency. METHODS: The study population consisted of two groups: group I involved 12 patients with alcoholic cirrhosis; group II involved 25 patients with hepatic B virus or hepatic C virus cirrhosis. The control group involved 20 healthy individuals. The degree of DNA fragmentation in peripheral blood lymphocytes was determined with the alkaline Comet assay that provides two indexes of the frequency of DNA single-strand breaks and alkali-labile sites, the tail length and the tail moment. RESULTS: Mean values of both tail length and tail moment were significantly increased (P<0.001) in lymphocytes from 12 patients with alcoholic cirrhosis and in lymphocytes from 25 patients with HBV or HCV cirrhosis, as compared with average tail length and tail moment values of lymphocytes from 20 healthy individuals. A significant positive correlation was found to exist between the degree of DNA fragmentation present in lymphocytes of each of the 37 patients with alcoholic or viral cirrhosis and the corresponding value of the Child-Pugh score. CONCLUSION: The occurrence of DNA fragmentation in peripheral blood lymphocytes reflects a direct genotoxic effect of either alcohol or HBV and HCV and suggests that the same genotoxic effect may operate in the liver and contribute to hepatocarcinogenesis.

1,8-Naphthyridines VI. Synthesis and anti-inflammatory activity of 5-(alkylamino)-N,N-diethyl[1,2,4]triazolo[4,3-a][1,8]naphthyridine-6-carboxamides with a new substitution pattern on the triazole ring.

On the basis of the good anti-inflammatory properties shown by the 9-alkyl-N,N-dialkyl-5-(alkylamino)[1,2,4]triazolo[4,3-a][1,8]naphthyridine-6-carboxamides 1, a series of analogues of such compounds, in which the 9-alkyl substituent was replaced by an ester or amide group (compounds 3a-i), was prepared and tested (inhibition of carrageenan-induced paw edema in the rat). Also two 5-(N-alkyl,N-acylamino) derivatives (compounds 4a,b) were synthesized and evaluated for the same purpose. Even though the general trend for these new [1,2,4]triazolo[4,3-a][1,8]naphthyridine derivatives was a decrease in activity compared with compounds 1, some of the new synthesized compounds exhibited still good anti-inflammatory properties.

DNA fragmentation and DNA repair synthesis induced in rat and human thyroid cells by chemicals carcinogenic to the rat thyroid.

Five chemicals that are known to induce in rats thyroid follicular-cell adenomas and carcinomas were assayed for their ability to induce DNA damage and DNA repair synthesis in primary cultures of human thyroid cells. Significant dose-dependent increases in the frequency of DNA single-strand breaks and alkali-labile sites, as measured by the same Comet assay, were obtained after a 20-h exposure to the following subtoxic concentrations of the five test compounds: methimazole from 2.5 to 10mM; nitrobenzene, potassium bromate, N,N'-diethylthiourea and ethylenethiourea from 1.25 to 5mM. Under the same experimental conditions, DNA repair synthesis, as evaluated by quantitative autoradiography, was present in potassium bromate-exposed thyroid cells from all the three donors and in those from two of three donors with either nitrobenzene or ethylenethiourea, but did not match the criteria for a positive response in thyroid cells from any of the donors with methimazole and N,N'-diethylthiourea. Consistently with their ability to induce thyroid tumors, all the five test compounds, administered p.o. in rats in a single dose corresponding to 1/2 LD50, induced a statistically significant degree of DNA fragmentation in the thyroid. These findings suggest that the five test compounds might be carcinogenic to thyroid in humans.

Intravesical gemcitabine in superficial bladder cancer: a phase II safety, efficacy and pharmacokinetic study.

BACKGROUND: We investigated the safety, efficacy and pharmacokinetics of the intravesical administration of 2000 mg gemcitabine once a week in the four weeks before transurethral resection of superficial bladder cancer (TUR), and in the four successive weeks. MATERIALS AND METHODS: Nine patients with superficial transitional cell bladder carcinoma were studied. Two thousand mg of gemcitabine dissolved in 50 ml of distilled water were administered intravesically. The dwell time was 60 min. The pharmacokinetics of gemcitabine and its metabolite, 2',2'-difluorodeoxyuridine (dFdU), were studied in plasma and urine before and after TUR. Cystoscopy was repeated 30 days after completion of the TUR treatment and subsequently at time intervals of one or two months. RESULTS: No systemic toxicity was noted, and only three patients displayed modest signs of local toxicity. One patient had recurrence 1 month after TUR, three between 3 and 6 months, and another three after 8, 11 and 18 months, respectively; two were recurrence-free after 21 and 22 months, respectively. The peak plasma concentrations of gemcitabine never exceeded 1000 ng/ml before TUR and 350 ng/ml after TUR, and declined rapidly. The plasma levels of dFdU were higher than those of gemcitabine, increased until 60 min and then declined little. Between 52% and 100% of the gemcitabine dose was present in voided urine. CONCLUSION: Intravesical gemcitabine, at the dose of 2000 mg, is well tolerated, is associated with minimal systemic absorption and has a moderate efficacy in the treatment of superficial bladder cancer.

DNA fragmentation and DNA repair synthesis induced in rat and human thyroid cells by four rat thyroid carcinogens.

Four chemicals that are known to induce in rats thyroid follicular-cell adenomas and carcinomas were assayed for their ability to induce DNA damage and DNA repair synthesis in primary cultures of human thyroid cells. Significant dose-dependent increases in the frequency of DNA single-strand breaks and alkali-labile sites, as measures by the Comet assay, were obtained after a 20-h exposure to the following subtoxic concentrations of the four test compounds: 2,4-diaminoanisole (DAA) from 0.10 to 1.0 mM, 4,4'-methylene-bis(N,N-dimethyl)benzenamine (MDB) from 0.32 to 1.8 mM, propylthiouracil (PTU) from 1.8 to 5.6 mM, and 4,4'-thiodianiline (THA) from 0.032 to 0.18 mM. Under the same experimental conditions, DNA repair synthesis, as evaluated by quantitative autoradiography, was present in thyreocytes exposed to DAA but absent after treatment with MDB, PTU, and THA. Consistent with their thyroid-specific carcinogenic activity, all the four chemicals, administered p.o. in rats in a single dose corresponding to 1/2 LD50, induced a statistically significant degree of DNA fragmentation in the thyroid, whereas any substantial evidence of DNA lesions was absent in liver, kidney, and lung, which, with the exception of liver tumors caused by THA, are not targets of the carcinogenic activity of the four test compounds. These findings indicate that the DNA damage observed in thyroid cells was consistent with the carcinogenicity of the four test compounds, and suggest that DAA, MDB, PTU, and THA might be carcinogenic to thyroid in humans.

DNA fragmentation, DNA repair and apoptosis induced in primary rat hepatocytes by dienogest, dydrogesterone and 1,4,6-androstatriene-17beta-ol-3-one acetate.

Four steroids that share the 17-hydroxy-3-oxopregna-4,6-diene structure - cyproterone acetate, chlormadinone acetate, megestrol acetate, and potassium canrenoate - have been shown previously to behave with different potency as liver-specific genotoxic agents, the response being markedly higher in female than in male rats, but similar in humans of both genders. In this study, performed to better define the relationship between chemical structure and genotoxicity, dydrogesterone (DGT) with double bonds C4=C5 and C6=C7, dienogest (DNG) with double bonds C4=C5 and C9=C10, and 1,4,6-androstatriene-17beta-ol-3-one acetate (ADT) with double bonds C1=C2, C4=C5 and C6=C7, were compared with cyproterone acetate (CPA) for their ability to induce DNA fragmentation and DNA repair synthesis in primary cultures of hepatocytes from three rats of each sex. At subtoxic concentrations, ranging from 10 to 90 microM, all four steroids consistently induced a dose-dependent increase of DNA fragmentation, which in all cases was higher in females than in males; their DNA damaging potency decreased in the order CPA > DNG > ADT > DGT. Under the same experimental conditions, the responses provided by the DNA repair-synthesis assay were positive or inconclusive in hepatocytes from female rats and consistently negative in hepatocytes from male rats. In the induction of apoptotic cells, examined in primary hepatocytes from female rats, CPA was more active than ADT and DGT, and DNG was inactive. Considered as a whole these findings suggest that a liver-specific genotoxic effect more marked in female than in male rats might be a common property of steroids with two or three double bonds.