A second locus for Marfan syndrome maps to chromosome 3p24.2-p25.

Marfan syndrome (MFS) is an autosomal dominant connective-tissue disorder characterized by skeletal, ocular and cardiovascular defects of highly variable expressivity. The diagnosis relies solely on clinical criteria requiring anomalies in at least two systems. By excluding the chromosome 15 disease locus, fibrillin 1 (FBN1), in a large French family with typical cardiovascular and skeletal anomalies, we raised the issue of genetic heterogeneity in MFS and the implication of a second locus (MFS2). Linkage analyses, performed in this family, have localized MFS2 to a region of 9 centiMorgans between D3S1293 and D3S1283, at 3p24.2-p25. In this region, the highest lod score was found with D3S2336, of 4.89 (theta = 0.05). By LINKMAP analyses, the most probable position for the second locus in MFS was at D3S2335.

Moderate intergenerational and somatic instability of a 55-CTG repeat in transgenic mice.

Myotonic dystrophy (DM) is associated with the expansion of a (CTG)n trinucleotide repeat in the 3' untranslated region (UTR) of the DM protein kinase gene (DMPK). The (CTG)n repeat is polymorphic and varies in size between 5 and 37 repeats in unaffected individuals whereas in affected patients there are between 50 and 4,000 CTGs. The size of the (CTG)n repeat, which increases through generations, generally correlates with clinical severity and age of onset. The instability of the CTG repeat appears to depend on its size as well as on the sex of the transmitting parent. Moreover, mitotic instability analysis of different human DM tissues shows length mosaicism between different cell lineages. The molecular mechanisms of triplet instability remain elusive. To investigate the role of genomic sequences in instability, we produced transgenic mice containing a 45-kb genomic segment with a 55-CTG repeat cloned from a mildly affected patient. In contrast to other mouse models containing CAG repeats within cDNAs, these mice showed both intergenerational and somatic repeat instability.

[Epigenetics, interface between environment and genes: role in complex diseases]. / Epigénétique, interface entre environnement et gènes: rô1e dans les maladies complexes.

Epigenetics is the study of heritable changes in gene expression or cellular phenotype caused by mechanisms other than changes in the underlying DNA sequence. Epigenetics is one of the major mechanisms explaining the "Developmental Origin of Health and Diseases" (DOHaD). Besides genetic background inherited from parents, which confers susceptibility to certain pathologies, epigenetic changes constitute the memory of previous events, either positive or negative, along the life cycle, including at the in utero stage. The later exposition to hostile environment may reveal such susceptibility, with the development of various pathologies, among them numerous chronic complex diseases. The demonstration of such a sequence of events has been shown for metabolic diseases as obesity, metabolic syndrome and type 2 diabetes, cardiovascular disease and cancer. In contrast to genetic predisposition, which is irreversible, epigenetic changes are potentially reversible, thus giving targets not only for prevention, but possibly also for the treatment of certain complex diseases.

Two anonymous DNA segments distinguish the Wilms' tumor and aniridia loci.

The association of Wilms' tumor with aniridia (the WAGR complex) in children with 11p13 chromosomal abnormalities has been established, but the paucity of molecular probes in 11p13 has hampered identification of the responsible genes. Two new anonymous DNA segments have been identified that map to the WAGR region of 11p13. Both DNA probes identify a cytologically undetectable deletion associated with a balanced chromosome translocation inherited by a patient with familial aniridia, but not Wilms' tumor. The same two DNA segments are also included in the distal p13-p14.1 deletion of another patient, who has aniridia, Wilms' tumor, and hypogonadism, but they are not included in the p12-p13 deletion of a third patient, who does not have aniridia but has had a Wilms' tumor. The discovery of this aniridia deletion and these two DNA segments that physically separate the Wilms' tumor and aniridia loci should facilitate identification of the genes in the WAGR locus, beginning with the aniridia gene.

Beckwith-Wiedemann syndrome, tumourigenesis and imprinting.

The concurrent development of cytogenetic, clinical, genetic and molecular studies has led to the recognition that the different hereditary and non-hereditary forms of the Beckwith-Wiedemann syndrome and associated tumours result from an imbalance between maternal and paternal alleles. The most exciting development in the past year was the discovery of uniparental paternal disomy and the increased understanding, arising from studies in the mouse and in hereditary cases, of the role possibly played by imprinting and somatic mosaicism in partial and complete expression of this complex syndrome.

Report on the IASO Stock Conference 2006: early and lifelong environmental epigenomic programming of metabolic syndrome, obesity and type II diabetes.

Now that analysis of the organization of the human genome sequence is reaching completion, studies of the finely tuned chromatin epigenetic networks, DNA methylation and histone modifications, are required to determine how the same DNA sequence generates different cells, lineages and organs, i.e. the phenotype. Maternal nutrition, behaviour and metabolic disturbances as well as other environmental factors have been shown to have major effects on these epigenetic processes, potentially affecting the predisposition of offspring to obesity and related adult disorders. The March 2006 Stock Conference considered the latest evidence from studies in the field of obesity and other related areas that elucidate mechanisms by which the environment can modify gene expression and the resulting individual phenotype. Presentations included evaluation of the molecular basis of epigenetic memory and the nature of relevant sequence targets, windows of susceptibility, and maternal dietary and behavioural factors that determine epigenetic changes. Imprinted genes, age and tissue-related exposures, transgenerational and potential interventions were also discussed. In summary, it is clear that epigenetic alterations can no longer be ignored in evaluations of the causes of obesity and its associated disorders. There is a need for systematic large-scale epigenetic studies of obesity, employing appropriate strategies and techniques and appropriately chosen environmental factors in critical spatio-temporal windows.

Anderson's disease: genetic exclusion of the apolipoprotein-B gene in two families.

Anderson's disease is a recessive disorder characterized by intestinal fat malabsorption, absence of postprandial chylomicrons, and reduced levels of cholesterol, triglycerides, and apoproteins B, AI, and C. We have studied two families with, respectively, three and two children with Anderson's disease. Intestinal apo-B and apo-AIV mRNAs from two Anderson's patients were normal in size but their concentration was decreased fivefold compared with controls. After DNA digestion with seven restriction enzymes, restriction fragment length polymorphisms of apo-B gene did not show conclusive information except for Xba1, which revealed a lack of cosegregation between the restriction fragment length polymorphism and the Anderson's phenotype. Linkage analysis was performed using the polymorphism of the apo-B gene 3'minisatellite. Genomic DNA from parents and children was amplified by polymerase chain reaction using oligonucleotide primers flanking the apo-B gene 3'hypervariable locus. In both families each child inherited different apo-B alleles from at least one parent. According to the recessive mode of transmission of the disease, our results are incompatible with the involvement of the apo-B gene. More likely a posttranslational defect or a mutation in another gene encoding a protein essential for lipoprotein assembly or secretion may be involved.

Paternal mutation of the sulfonylurea receptor (SUR1) gene and maternal loss of 11p15 imprinted genes lead to persistent hyperinsulinism in focal adenomatous hyperplasia.

Congenital hyperinsulinism, or persistent hyperinsulinemic hypoglycemia of infancy (PHHI), is a glucose metabolism disorder characterized by unregulated secretion of insulin and profound hypoglycemia. From a morphological standpoint, there are two types of histopathological lesions, a focal adenomatous hyperplasia of islet cells of the pancreas in approximately 30% of operated sporadic cases, and a diffuse form. In sporadic focal forms, specific losses of maternal alleles (LOH) of the imprinted chromosomal region 11p15, restricted to the hyperplastic area of the pancreas, were observed. Similar mechanisms are observed in embryonal tumors and in the Beckwith-Wiedemann syndrome (BWS), also associated with neonatal but transient hyperinsulinism. However, this region also contains the sulfonylurea receptor (SUR1) gene and the inward rectifying potassium channel subunit (KIR6.2) gene, involved in recessive familial forms of PHHI, but not known to be imprinted. Although the parental bias in loss of maternal alleles did not argue in favor of their direct involvement, the LOH may also unmask a recessive mutation leading to persistent hyperinsulinism. We now report somatic reduction to hemizygosity or homozygosity of a paternal SUR1 constitutional heterozygous mutation in four patients with a focal form of PHHI. Thus, this somatic event which leads both to beta cell proliferation and to hyperinsulinism can be considered as the somatic equivalent, restricted to a microscopic focal lesion, of constitutional uniparental disomy associated with unmasking of a heterozygous parental mutation leading to a somatic recessive disorder.

Somatic deletion of the imprinted 11p15 region in sporadic persistent hyperinsulinemic hypoglycemia of infancy is specific of focal adenomatous hyperplasia and endorses partial pancreatectomy.

Sporadic persistent hyperinsulinemic hypoglycemia of infancy (PHHI) or nesidioblastosis is a heterogeneous disorder characterized by profound hypoglycemia due to inappropriate hypersecretion of insulin. An important diagnostic goal is to distinguish patients with a focal hyperplasia of islet cells of the pancreas (FoPHHI) from those with a diffuse abnormality of islets (DiPHHI) because management strategies differ significantly. 16 infants with sporadic PHHI resistant to diazoxide and who underwent pancreatectomy were investigated. Selective pancreatic venous sampling coupled with peroperative surgical examination and analysis of extemporaneous frozen sections allowed us to identify 10 cases with FoPHHI and 6 cases with DiPHHI. We show here that in cases of FoPHHI, but not those of DiPHHI, there was specific loss of maternal alleles of the imprinted chromosome region 11p15 in cells of the hyperplastic area of the pancreas but not in normal pancreatic cells. This somatic event is consistent with a proliferative monoclonal lesion. It involves disruption of the balance between monoallelic expression of several maternally and paternally expressed genes. Thus, we provide the first molecular explanation of the heterogeneity of sporadic forms of PHHI such that it is possible to perform only partial pancreatectomy, limited to the focal somatic lesion, so as to avoid iatrogenic diabetes in patients with focal adenomatous hyperplasia.

Congenital hyperinsulinism and mosaic abnormalities of the ploidy.

BACKGROUND: Congenital hyperinsulinism and Beckwith-Wiedemann syndrome both lead to beta islet hyperplasia and neonatal hypoglycaemia. They may be related to complex genetic/epigenetic abnormalities of the imprinted 11p15 region. The possibility of common pathophysiological determinants has not been thoroughly investigated. OBJECTIVE: To report abnormalities of the ploidy in two unrelated patients with congenital hyperinsulinism. METHODS: Two patients with severe congenital hyperinsulinism, one overlapping with Beckwith-Wiedemann syndrome, had pancreatic histology, ex vivo potassium channel electrophysiological studies, and mutation detection of the encoding genes. The parental genetic contribution was explored using genome-wide polymorphism, fluorescent in situ hybridisation (FISH), and blood group typing studies. RESULTS: Histological findings diverged from those described in focal congenital hyperinsulinism or Beckwith-Wiedemann syndrome. No potassium channel dysfunction and no mutation of its encoding genes (SUR1, KIR6.2) were detected. In patient 1 with congenital hyperinsulinism and Beckwith-Wiedemann syndrome, paternal isodisomy for the whole haploid set was homogeneous in the pancreatic lesion, and mosaic in the leucocytes and skin fibroblasts (hemihypertrophic segment). Blood group typing confirmed the presence of two erythroid populations (bi-parental v paternal only contribution). Patient 2 had two pancreatic lesions, both revealing triploidy with paternal heterodisomy. Karyotype and FISH analyses done on the fibroblasts and leucocytes of both patients were unremarkable (diploidy). CONCLUSIONS: Diploid (biparental/paternal-only) mosaicism and diploid/triploid mosaicism were present in two distinct patients with congenital hyperinsulinism. These chromosomal abnormalities led to paternal disomy for the whole haploid set in pancreatic lesions (with isodisomy or heterodisomy), thereby extending the range and complexity of the mechanisms underlying congenital hyperinsulinism, associated or not with Beckwith-Wiedemann syndrome.

An oligonucleotide microarray for mouse imprinted genes profiling.

Genomic imprinting is an epigenetic phenomenon unique to mammals that causes some genes to be expressed according to their parental origin. It results in developmental asymmetry in the function of the parental genomes. We describe here a method for the profiling of imprinted genes based on the development of a mouse imprinting microchip containing oligonucleotides corresponding to 493 genes, including most of the known imprinted genes (IG = 63), genes involved in epigenetic processes (EPI = 15), in metabolism (= 147), in obesity (= 10) and in neurotransmission (= 256) and housekeeping reference genes (= 2). This custom oligonucleotide microarray has been constructed to make data analysis and handling more manageable than pangenomic microarrays. As a proof of concept we present the differential expression of these 493 genes in different tissues (liver, placenta, embryo) of C57BL6/J mice fed different diets. Appropriate experimental strategies and statistical tools were defined at each step of the data analysis process with regard to the different sources of constraints. Data were confirmed by expression analyses based on quantitative real-time PCR. These oligochips should make it possible to increase our understanding of the involvement of imprinted genes in the timing of expression programs, tissue by tissue, stage by stage, in response to nutrients, lifestyles and other as yet unknown critical environmental factors in a variety of physiopathological situations, and in animals of different strains, ages and sexes. The use of oligonucleotides makes it possible to expand this microchip to include the increasing number of imprinted genes discovered.

Seven megabase yeast artificial chromosome contig at region 11p15: identification of a yeast artificial chromosome spanning the breakpoint of a chromosomal translocation found in a case of Beckwith-Wiedemann syndrome.

Genetic alterations of chromosome region 11p15 have been detected in neoplastic diseases as well as in cancer-predisposing syndromes. The cloning of the entire chromosomal region will be important for the identification and characterization of critical tumor suppressor genes. We have developed a yeast artificial chromosome contig that covers up to 7 Mb of this chromosome band. The most centromeric marker included in the contig is D11S932 and the most telomeric is D11S470. We have developed 18 new STS markers, which have been located in the contig in relation to 16 known markers. One of the yeast artificial chromosome clones was found to span the chromosome 11 breakpoint of the translocation t(11;18), associated with a case of Beckwith-Wiedemann syndrome. Cloning the regions in proximity to this translocation might reveal the presence of a gene altered in association with the development of Beckwith-Wiedemann syndrome.

Isolation of kidney complementary DNAs down-expressed in Wilms' tumor by a subtractive hybridization approach.

We applied a subtractive hybridization approach to isolate genes differentially expressed between mature kidney and Wilms' tumor. We constructed a complementary DNA library from a total mature kidney complementary DNA subtracted by an excess of mRNA from a Wilms' tumor, WAGR4, with a germline deletion of 11p13 and a somatic loss of alleles at 11p15. Six clones presenting a differential pattern of expression, positive with mRNA from the mature kidney and negative with mRNA from the Wilms' tumor WAGR4, were characterized. Among these clones were two as yet unknown expressed sequences (D11S877E and D15S109E) and four sequences from known genes: renal dipeptidase (DPEP1), alpha B-crystallin (CRYA2), uromodulin (UMOD), and plasma glutathione peroxidase (GPX2). The different patterns of expression of these genes in 11 Wilms' tumors, whether or not they are hereditary, reflect the well-documented pathogenetic heterogeneity for Wilms' tumors. We propose that these clones could be helpful for an improved histological characterization of Wilms' tumors.

Candidate genetic modifiers of individual susceptibility to renal cell carcinoma: a study of polymorphic human xenobiotic-metabolizing enzymes.

The steady increase in sporadic renal cell carcinoma (RCC) observed in industrialized countries supports the notion that certain carcinogens present in the environment (tobacco smoke, drugs, pollutants, and dietary constituents) may affect the occurrence of RCC. Many of the enzymes dealing with such environmental factors are polymorphic and may, therefore, confer variable susceptibility to RCC. This case-control study was designed to test for an association between genetic polymorphism of enzymes involved in xenobiotic metabolism and the risk of sporadic RCC. Genomic DNA was obtained from 173 patients with RCC and 211 controls of Caucasian origin. We used PCR-RFLP to investigate polymorphism for the most common alleles at two cytochrome-P450 mono-oxygenases (CYP1A1 and CYP2D6), one NAD[P]H:quinone oxidoreductase (NQO1), three glutathione S-transferases (GSTM1, GSTT1, and GSTP1), and one N-acetyltransferase (NAT2) loci. The CYP1A1 (m) "variant" genotype, which contains at least one copy of the CYP1A1 variant alleles, was found to be associated with a 2.1-fold [95% confidence interval (CI), 1.1-3.9] increase in the risk of RCC. There was also a higher risk of RCC for subjects with the CYP1A1 (m) variant genotype combined with any of the following genotypes: GSTT1 (+) "active" [odds ratio (OR), 2.3; 95% CI, 1.2-4.5], GSTP1 (m) variant (OR, 2.4; 95% CI, 1.0-5.4), or NAT2 (-) "slow acetylator" (OR, 2.5; 95% CI, 1.1-5.5). A significant association was also found for the GSTM1 (-) "null" and GSTP1 (m) genotypes combined with either NAT2 (-) (OR, 2.6; 95% CI, 1.2-5.8) or CYP1A1 (m) (OR, 3.5; 95% CI, 1.1-11.2). The CYP2D6 (-) "poor metabolizer " and the NQO1 (-) "defective" genotypes were not clearly associated with a higher risk of RCC. Our data demonstrate for the first time a significant association between a group of pharmacogenetic polymorphisms and RCC risk. These positive findings suggest that interindividual variation in the metabolic pathways involved in the functionalization and detoxification of specific xenobiotics is an important susceptibility factor for RCC in Caucasians.

Relationships among electrophysiological findings and clinical status, heart function, and extent of DNA mutation in myotonic dystrophy.

BACKGROUND: Impulse-conduction abnormalities and arrhythmias are common in myotonic dystrophy (MD). This study was performed to determine whether a correlation exists between electrophysiological (EP) testing data and clinical status, heart function, or size of the DNA abnormality (cytosine-thymine-guanine sequence repeat). METHODS AND RESULTS: Eighty-three MD patients underwent invasive EP studies prompted primarily by the presence of asymptomatic conduction abnormalities. AV conduction disturbances were common and mainly distal (HV interval, 66.2+/-14 ms). AV conduction observed from the surface ECG was generally concordant with endocardial measurements. However, 11 of 20 patients with normal surface ECGs had abnormal subhisian conduction. Atrial arrhythmias were inducible in 41% of cases and correlated with prolongation of the AH interval (P=0.02) and a shorter atrial refractory period (P=0.04). Induction of ventricular arrhythmias (18%) correlated strongly with age (P=0. 0003). After adjustment for age, the extent of DNA mutation correlated with the Walton score (P=0.0018) but not with conduction abnormalities or induction of arrhythmias. CONCLUSIONS: Prolongation of the HV interval is the most common conduction abnormality in MD and can be reliably recognized only by invasive EP testing. It raises the issue of prophylactic pacing to limit the incidence of sudden death in MD. Atrial and ventricular arrhythmias are often inducible, although their predictive value remains to be determined. Young age emerged as the most powerful predictor of inducible ventricular tachyarrhythmias. Conversely, we found no relationship between ECG or EP abnormalities recorded during invasive testing and the DNA mutation size or severity of peripheral muscle involvement.

R3531C mutation in the apolipoprotein B gene is not sufficient to cause hypercholesterolemia.

Familial hypercholesterolemia and familial ligand-defective apolipoprotein B-100 (FDB) are dominantly inherited disorders leading to impaired low-density lipoprotein receptor (LDLR) and apolipoprotein B-100 (APOB) interaction, plasma LDL elevation, and hypercholesterolemia. We previously identified the first French FDB-R3531C proband, a woman with very high total cholesterol, in a group of type IIa hypercholesterolemic families. We report here the investigation of her family at large that revealed the total absence of cosegregation with hypercholesterolemia. Six of the 10 subjects heterozygous for the R3531C mutation had plasma cholesterol lower than the 97.5th percentile for their age and gender, and mean cholesterol levels were not significantly different between affected and unaffected persons. Furthermore, 2 family members with similar high LDL-cholesterol levels were not carriers of the R3531C substitution, suggesting the implication of another mutation. Segregation analysis of the LDLR gene revealed statistically significant genetic linkage with hypercholesterolemia, and analysis of the proband LDLR gene led to the identification of the 664 proline to leucine defective mutation and its detection in all 6 hypercholesterolemic-related members of this family. Therefore, our results show that the family presents with familial hypercholesterolemia and give evidence that the R3531C substitution in the APOB gene is not an allelic variant leading to FDB. Furthermore, thorough analysis of our data suggests that the APOB-R3531C mutation enhances the hypercholesterolemic effect of the LDLR-P664L defect, suggesting that it is a susceptibility mutation.

WT1 splicing alterations in Wilms' tumors.

Hereditary and sporadic forms of tumors are generally related to germ-line and somatic mutations of the same tumor suppressor gene. Unexpectedly, in Wilms' tumor, somatic mutations of the WT1 gene were found only occasionally in sporadic cases, although constitutional mutations of this gene are clearly associated with predisposition. It has been suggested that abnormal splicing may be another mode of somatic WT1 alteration. However, this idea was based on the analysis of a small series of tumors, precluding accurate evaluation of the frequency of such changes. To investigate WT1 changes at the somatic level in more detail, we analyzed the levels of the four isoform transcripts produced by alternative splicing events in a large series of 50 tumors, normal mature kidneys, and fetal kidneys. We characterized splicing alterations in 63% of sporadic Wilms' tumors. Moreover, taking into account the decreased and increased overall levels of WT1 mRNA, the percentage of sporadic tumors with changes in WT1 expression reached 90%. Whether and how these alterations of expression play a role in the tumorigenic process remain to be evaluated.

Silencing of CDKN1C (p57KIP2) is associated with hypomethylation at KvDMR1 in Beckwith-Wiedemann syndrome.

CONTEXT: Beckwith-Wiedemann syndrome (BWS) arises by several genetic and epigenetic mechanisms affecting the balance of imprinted gene expression in chromosome 11p15.5. The most frequent alteration associated with BWS is the absence of methylation at the maternal allele of KvDMR1, an intronic CpG island within the KCNQ1 gene. Targeted deletion of KvDMR1 suggests that this locus is an imprinting control region (ICR) that regulates multiple genes in 11p15.5. Cell culture based enhancer blocking assays indicate that KvDMR1 may function as a methylation modulated chromatin insulator and/or silencer. OBJECTIVE: To determine the potential consequence of loss of methylation (LOM) at KvDMR1 in the development of BWS. METHODS: The steady state levels of CDKN1C gene expression in fibroblast cells from normal individuals, and from persons with BWS who have LOM at KvDMR1, was determined by both real time quantitative polymerase chain reaction (qPCR) and ribonuclease protection assay (RPA). Methylation of the CDKN1C promoter region was assessed by Southern hybridisation using a methylation sensitive restriction endonuclease. RESULTS: Both qPCR and RPA clearly demonstrated a marked decrease (86-93%) in the expression level of the CDKN1C gene in cells derived from patients with BWS, who had LOM at KvDMR1. Southern analysis indicated that downregulation of CDKN1C in these patients was not associated with hypermethylation at the presumptive CDKN1C promoter. CONCLUSIONS: An epimutation at KvDMR1, the absence of maternal methylation, causes the aberrant silencing of CDKN1C, some 180 kb away on the maternal chromosome. Similar to mutations at this locus, this silencing may give rise to BWS.

[Nutritionnal epigenomics: consequences of unbalanced diets on epigenetics processes of programming during lifespan and between generations]. / Epigénomique nutritionnelle: impact de régimes alimentaires déséquilibrés sur les processus épigénétiques de programmation au cours de la vie et transgénérationnels.

Epigenetic changes associated with DNA methylation and histone modifications leading to chromatin remodeling and regulation of gene expression underlie the developmental programming of obesity, type 2 diabetes, cardiovascular diseases and metabolic syndrome. This review focuses on converging data supporting the hypothesis that, in addition to "thrifty genotype" inheritance, individuals with obesity, type 2 diabetes, and metabolic syndrome (MetS) with an increased risk of cardiovascular diseases have suffered improper "epigenetic programming" during their fetal/postnatal development due to maternal inadequate nutrition and metabolic disturbances and also during their lifetime, that could even be transmitted to the next generation(s). We highlight the susceptibility of epigenetic mechanisms controlling gene expression to environmental influences due to their inherent malleability, emphasizing the participation of transposable elements and the potential role of imprinted genes during critical time windows in epigenetic programming, from the very beginning of development, throughout life. Increasing our understanding on epigenetic patterns significance and their role in development, evolution and adaptation and on small molecules (nutrients, drugs) that reverse epigenetic (in)activation should provide us with the means to "unlock" silenced (enhanced) genes, and to "convert" the obsolete human thrifty genotype into a "squandering" phenotype.

Association of GSTT1 non-null and NAT1 slow/rapid genotypes with von Hippel-Lindau tumour suppressor gene transversions in sporadic renal cell carcinoma.

The von Hippel-Lindau (VHL) tumour suppressor gene is commonly mutated in renal cell carcinoma of clear cell type (CCRCC). We investigated the possible relationship between VHL mutations in sporadic CCRCC and polymorphism of genes encoding enzymes involved in carcinogen metabolism: two cytochrome P450 monooxygenases (CYP1A1 and CYP2D6), one NAD[P]H:quinone oxidoreductase (NQO1), three glutathione S-transferases (GSTM1, GSTT1 and GSTP1) and two arylamine N-acetyltransferases (NAT1 and NAT2). We analysed DNA from tumour and nontumoural kidney tissue from 195 CCRCC patients. Single VHL mutations were identified in 88 patients and double mutations were present in two patients. Nine of 18 transversions were GC to TA, four were AT to TA, four were GC to CG and one was AT to CG. Ten of 19 transitions were GC to AT and nine were AT to GC. We also identified 53 frameshifts and two GC to AT at CpG. An excess of transversions was observed in a subset of patients with active GSTT1 [GSTT1 (+) genotype] and probably defective NAT1 (NAT1 S/R variant genotype). All 18 transversions were in GSTT1 (+) patients, whereas only 76% of transitions (P = 0.05) and 81% of the other mutations (P = 0.06) occurred in this genotype. We found that 28% of the transversions were in the NAT1 S/R genotype versus 12% of the transitions (P = 0.40) and 4% of the other mutations (P = 0.01). This suggests that pharmacogenetic polymorphisms may be associated with the type of acquired VHL mutation, which may modulate CCRCC development.