The ICF syndrome (i̲mmunodeficiency, c̲entromeric region instability, f̲acial anomalies) is a unique DNA methylation deficiency disease diagnosed by an extraordinary collection of chromosomal anomalies specifically in the vicinity of the centromeres of chromosomes 1 and 16 (Chr1 and Chr16) in mitogen-stimulated lymphocytes. These aberrations include decondensation of centromere-adjacent (qh) heterochromatin, multiradial chromosomes with up to 12 arms, and whole-arm deletions. We demonstrate that lymphoblastoid cell lines from two ICF patients exhibit these Chr1 and Chr16 anomalies in 61% of the cells and continuously generate 1qh or 16qh breaks. No other consistent chromosomal abnormality was seen except for various telomeric associations, which had not been previously noted in ICF cells. Surprisingly, multiradials composed of arms of both Chr1 and Chr16 were favored over homologous associations and cells containing multiradials with 3 or >4 arms almost always displayed losses or gains of Chr1 or Chr16 arms from the metaphase. Our results suggest that decondensation of 1qh and 16qh often leads to unresolved Holliday junctions, chromosome breakage, arm missegregation, and the formation of multiradials that may yield more stable chromosomal abnormalities, such as translocations. These cell lines maintained the abnormal hypomethylation in 1qh and 16qh seen in ICF tissues. The ICF-specific hypomethylation occurs in only a small percentage of the genome, e.g., ICF brain DNA had 7% less 5-methylcytosine than normal brain DNA. The ICF lymphoblastoid cell lines, therefore, retain not only the ICF-specific pattern of chromosome rearrangements, but also of targeted DNA hypomethylation. This hypomethylation of heterochromatic DNA sequences is seen in many cancers and may predispose to chromosome rearrangements in cancer as well as in ICF.

We quantitatively analysed hypermethylation at CpG islands in the 5' ends of 12 genes and one non-CpG island 5' region (MTHFR) in 31 Wilms tumors. We also determined their global genomic 5-methylcytosine content. Compared with various normal postnatal tissues, *40 -90% of these pediatric kidney cancers were hypermethylated in four of the genes, MCJ, RASSF1A, TNFRSF12 and CALCA as determined by a quantitative bisulfite-based assay (MethyLight). Interestingly, the non-CpG island 5' region of MTHFR was less methylated in most tumors relative to the normal tissues. By chromatographic analysis of DNA digested to deoxynucleosides, about 60% of the Wilms tumors were found to be deficient in their overall levels of DNA methylation. We also analysed expression of the three known functional DNA methyltransferase genes. No relationship was observed between global genomic 5-methylcytosine levels and relative amounts of RNA for DNA methyltransferases DNMT1, DNMT3A, and DNMT3B. Importantly, no association was seen between CpG island hypermethylation and global DNA hypomethylation in these cancers. Therefore, the overall genomic hypomethylation frequently observed in cancers is probably not just a response or a prelude to hypermethylation elsewhere in the genome. This suggests that the DNA hypomethylation contributes independently to oncogenesis or tumor progression.

1. The induction of phase I and II enzymes in the liver of the male F344 rat drinking 2% (w/v) solutions of green or black tea for 6 weeks was investigated. Also studied were glutathione (GSH) and cyst(e)ine in blood, liver and kidney, as well as serum cholesterol, HDL cholesterol, triglycerides, and total and free testosterone. 2. The total carbon monoxide-discernible liver P450, b5 and NADPH-cytochrome c(P450) reductase activities were similar in all groups. 3. There were significant increases in liver of rat drinking green or black tea of P4501A1, 1A2 and 2B1 activities, but no change in P4502E1 and 3A4 activities. Of the phase II enzymes, UDP-glucuronyltransferase was increased, but glutathione S-transferase was not. 4. Serum GSH was higher in the group administered black tea, but GSH and cyst(e)ine in other groups was at control levels. Serum cholesterol was lower in rat given black compared with green tea. Triglycerides had a declining trend after green and black tea exposure compared with water controls. Free and total testosterone were not affected. 5. Thus, beverages widely used by man altered host biochemistry as regards specific phase I and II enzymes in liver of rat and specific serum parameters.

The metabolism of azoxymethane (AOM), methylazoxymethanol (MAM) and N-nitrosodimethylamine (NDMA) by liver microsomes from acetone-induced rats as well as by a reconstituted system containing purified cytochrome P450IIE1 was examined. The products consisted of MAM from AOM; methanol and formic acid from MAM; and methylamine, formaldehyde, methanol, methylphosphate and formic acid from NDMA. Compared to liver microsomes from untreated rats, the metabolic activity of acetone-induced microsomes was approximately 4 times higher for all three carcinogens. Using the reconstituted system, the enzyme activities (nmol substrate metabolized/nmol P450/min) for AOM, MAM and NDMA were 2.88 +/- 1.14, 2.87 +/- 0.59 and 9.47 +/- 2.24 respectively. Incubations carried out in the presence of a monoclonal antibody to cytochrome P450IIE1 resulted in a 85-90% inhibition of all three reactions in this system. These results provide conclusive evidence that AOM, MAM and NDMA are metabolized by the same form of rat liver cytochrome P450. In addition, the stoichiometry of NDMA products formed in these reactions indicates that denitrosation, a presumed detoxication process, and alpha-hydroxylation, an activation reaction, are also catalyzed by the same cytochrome P450 isozyme.

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