Joseph L. Wiemels

Aging and environmental exposures alter tissue-specific DNA methylation dependent upon CPG island context

Epigenetic control of gene transcription is critical for normal human development and cellular differentiation. While alterations of epigenetic marks such as DNA methylation have been linked to cancers and many other human diseases, interindividual epigenetic variations in normal tissues due to aging, environmental factors, or innate susceptibility are poorly characterized. The plasticity, tissue-specific nature, and variability of gene expression are related to epigenomic states that vary across individuals. Thus, population-based investigations are needed to further our understanding of the fundamental dynamics of normal individual epigenomes. We analyzed 217 non-pathologic human tissues from 10 anatomic sites at 1,413 autosomal CpG loci associated with 773 genes to investigate tissue-specific differences in DNA methylation and to discern how aging and exposures contribute to normal variation in methylation. Methylation profile classes derived from unsupervised modeling were significantly associated with age (P<0.0001) and were significant predictors of tissue origin (P<0.0001). In solid tissues (n = 119) we found striking, highly significant CpG island–dependent correlations between age and methylation; loci in CpG islands gained methylation with age, loci not in CpG islands lost methylation with age (P<0.001), and this pattern was consistent across tissues and in an analysis of blood-derived DNA. Our data clearly demonstrate age- and exposure-related differences in tissue-specific methylation and significant age-associated methylation patterns which are CpG island context-dependent. This work provides novel insight into the role of aging and the environment in susceptibility to diseases such as cancer and critically informs the field of epigenomics by providing evidence of epigenetic dysregulation by age-related methylation alterations. Collectively we reveal key issues to consider both in the construction of reference and disease-related epigenomes and in the interpretation of potentially pathologically important alterations.

Origins of chromosome translocations in childhood leukaemia

Chromosome translocations are often early or initiating events in leukaemogenesis, occurring prenatally in most cases of childhood leukaemia. Although these genetic changes are necessary, they are usually not sufficient to cause leukaemia. How, when and where do translocations arise? And can these insights aid our understanding of the natural history, pathogenesis and causes of leukaemia?

Prenatal origin of acute lymphoblastic leukaemia in children.

BACKGROUND There is little current insight into the natural history of childhood leukaemia or the timing of relevant mutational events. TEL-AML1 gene fusion due to chromosomal translocation is frequently seen in the common form of childhood acute lymphoblastic leukaemia. We investigated whether this abnormality arises prenatally. METHODS We identified, by reverse-transcriptase PCR screening of blood or bone marrow, TEL-AML1 fusion in 12 children, plus a pair of identical twins, aged 2-5 years from Italy and the UK, who had newly diagnosed acute lymphoblastic leukaemia. We amplified and sequenced the genomic TEL-AML1 fusion gene with a long-distance inverse PCR method. Primers were designed that could be used in short-range PCR to screen for patient-specific, leukaemia clone-specific TEL-AML1 genomic fusion sequences in neonatal blood spots from each child. FINDINGS We initially identified TEL-AML1 fusion sequences in blood spots from the identical twins, diagnosed with concordant acute lymphoblastic leukaemia at age 4 years, who shared a single or clonotypic TEL-AML1 sequence that suggested prenatal origin in one twin. Three children were excluded because control genes could not be amplified. Of the other nine patients, six had positive blood spots. Blood spots that were classified as negative were uninformative. INTERPRETATION Our findings showed that childhood acute lymphoblastic leukaemia is frequently initiated by a chromosome translocation event in utero. Studies in identical twins show however that such an event is insufficient for clinical leukaemia and that a postnatal promotional event is also required.

Glioma Groups Based on 1p/19q, IDH, and TERT Promoter Mutations in Tumors

BACKGROUND The prediction of clinical behavior, response to therapy, and outcome of infiltrative glioma is challenging. On the basis of previous studies of tumor biology, we defined five glioma molecular groups with the use of three alterations: mutations in the TERT promoter, mutations in IDH, and codeletion of chromosome arms 1p and 19q (1p/19q codeletion). We tested the hypothesis that within groups based on these features, tumors would have similar clinical variables, acquired somatic alterations, and germline variants. METHODS We scored tumors as negative or positive for each of these markers in 1087 gliomas and compared acquired alterations and patient characteristics among the five primary molecular groups. Using 11,590 controls, we assessed associations between these groups and known glioma germline variants. RESULTS Among 615 grade II or III gliomas, 29% had all three alterations (i.e., were triple-positive), 5% had TERT and IDH mutations, 45% had only IDH mutations, 7% were triple-negative, and 10% had only TERT mutations; 5% had other combinations. Among 472 grade IV gliomas, less than 1% were triple-positive, 2% had TERT and IDH mutations, 7% had only IDH mutations, 17% were triple-negative, and 74% had only TERT mutations. The mean age at diagnosis was lowest (37 years) among patients who had gliomas with only IDH mutations and was highest (59 years) among patients who had gliomas with only TERT mutations. The molecular groups were independently associated with overall survival among patients with grade II or III gliomas but not among patients with grade IV gliomas. The molecular groups were associated with specific germline variants. CONCLUSIONS Gliomas were classified into five principal groups on the basis of three tumor markers. The groups had different ages at onset, overall survival, and associations with germline variants, which implies that they are characterized by distinct mechanisms of pathogenesis. (Funded by the National Institutes of Health and others.).

Brain tumor epidemiology: consensus from the Brain Tumor Epidemiology Consortium.

Epidemiologists in the Brain Tumor Epidemiology Consortium (BTEC) have prioritized areas for further research. Although many risk factors have been examined over the past several decades, there are few consistent findings, possibly because of small sample sizes in individual studies and differences between studies in patients, tumor types, and methods of classification. Individual studies generally have lacked samples of sufficient size to examine interactions. A major priority based on available evidence and technologies includes expanding research in genetics and molecular epidemiology of brain tumors. BTEC has taken an active role in promoting understudied groups, such as pediatric brain tumors; the etiology of rare glioma subtypes, such as oligodendroglioma; and meningioma, which, although it is not uncommon, has only recently been registered systematically in the United States. There also is a pressing need for more researchers, especially junior investigators, to study brain tumor epidemiology. However, relatively poor funding for brain tumor research has made it difficult to encourage careers in this area. In this report, BTEC epidemiologists reviewed the groups consensus on the current state of scientific findings, and they present a consensus on research priorities to identify which important areas the science should move to address. Cancer 2008;113(7 suppl):1953–68.

EPIDEMIOLOGY OF INTRACRANIAL MENINGIOMA

Meningiomas are the most frequently reported primary intracranial neoplasms, accounting for approximately 25% of all such lesions diagnosed in the United States. Few studies have examined the risk factors associated with a diagnosis of meningioma with two categories of exposure, hormones (both endogenous and exogenous) and radiation, most strongly associated with meningioma risk. Limited data are also available on long-term outcomes for meningioma patients, although it is clear that the disease is associated with significant morbidity and mortality. Recent legislation passed in the United States (The Benign Brain Tumor Cancer Registries Amendment Act [H.R. 5204]) mandates registration of benign brain tumors such as meningioma. This will increase the focus on this disease over the coming years as well as likely increase the reported prevalence of the disease. The increased emphasis on research dedicated to the study of brain tumors coupled with the advent of new tools in genetic and molecular epidemiology make the current era an ideal time to advance knowledge for intracranial meningioma. This review highlights current knowledge of meningioma epidemiology and new directions for research efforts in this field.

Variants in the CDKN2B and RTEL1 regions are associated with high-grade glioma susceptibility.

The causes of glioblastoma and other gliomas remain obscure. To discover new candidate genes influencing glioma susceptibility, we conducted a principal component–adjusted genome-wide association study (GWAS) of 275,895 autosomal variants among 692 adult high-grade glioma cases (622 from the San Francisco Adult Glioma Study (AGS) and 70 from the Cancer Genome Atlas (TCGA)) and 3,992 controls (602 from AGS and 3,390 from Illumina iControlDB (iControls)). For replication, we analyzed the 13 SNPs with P < 10−6 using independent data from 176 high-grade glioma cases and 174 controls from the Mayo Clinic. On 9p21, rs1412829 near CDKN2B had discovery P = 3.4 × 10−8, replication P = 0.0038 and combined P = 1.85 × 10−10. On 20q13.3, rs6010620 intronic to RTEL1 had discovery P = 1.5 × 10−7, replication P = 0.00035 and combined P = 3.40 × 10−9. For both SNPs, the direction of association was the same in discovery and replication phases.

Epidemiology and etiology of meningioma

Although most meningiomas are encapsulated and benign tumors with limited numbers of genetic aberrations, their intracranial location often leads to serious and potentially lethal consequences. They are the most frequently diagnosed primary brain tumor accounting for 33.8% of all primary brain and central nervous system tumors reported in the United States between 2002 and 2006. Inherited susceptibility to meningioma is suggested both by family history and candidate gene studies in DNA repair genes. People with certain mutations in the neurofibromatosis gene (NF2) have a very substantial increased risk for meningioma. High dose ionizing radiation exposure is an established risk factor for meningioma, and lower doses may also increase risk, but which types and doses are controversial or understudied. Because women are twice as likely as men to develop meningiomas and these tumors harbor hormone receptors, an etiologic role for hormones (both endogenous and exogenous) has been hypothesized. The extent to which immunologic factors influence meningioma etiology has been largely unexplored. Growing emphasis on brain tumor research coupled with the advent of new genetic and molecular epidemiologic tools in genetic and molecular epidemiology promise hope for advancing knowledge about the causes of intra-cranial meningioma. In this review, we highlight current knowledge about meningioma epidemiology and etiology and suggest future research directions.

Methylenetetrahydrofolate reductase (MTHFR) polymorphisms and risk of molecularly defined subtypes of childhood acute leukemia

Low folate intake as well as alterations in folate metabolism as a result of polymorphisms in the enzyme methylenetetrahydrofolate reductase (MTHFR) have been associated with an increased incidence of neural tube defects, vascular disease, and some cancers. Polymorphic variants of MTHFR lead to enhanced thymidine pools and better quality DNA synthesis that could afford some protection from the development of leukemias, particularly those with translocations. We now report associations of MTHFR polymorphisms in three subgroups of pediatric leukemias: infant lymphoblastic or myeloblastic leukemias with MLL rearrangements and childhood lymphoblastic leukemias with either TEL-AML1 fusions or hyperdiploid karyotypes. Pediatric leukemia patients (n = 253 total) and healthy newborn controls (n = 200) were genotyped for MTHFR polymorphisms at nucleotides 677 (C→T) and 1,298 (A→C). A significant association for carriers of C677T was demonstrated for leukemias with MLL translocations (MLL+, n = 37) when compared with controls [adjusted odd ratios (OR) = 0.36 with a 95% confidence interval (CI) of 0.15–0.85; P = 0.017]. This protective effect was not evident for A1298C alleles (OR = 1.14). In contrast, associations for A1298C homozygotes (CC; OR = 0.26 with a 95% CI of 0.07–0.81) and C677T homozygotes (TT; OR = 0.49 with a 95% CI of 0.20–1.17) were observed for hyperdiploid leukemias (n = 138). No significant associations were evident for either polymorphism with TEL-AML1+ leukemias (n = 78). These differences in allelic associations may point to discrete attributes of the two alleles in their ability to alter folate and one-carbon metabolite pools and impact after DNA synthesis and methylation pathways, but should be viewed cautiously pending larger follow-up studies. The data provide evidence that molecularly defined subgroups of pediatric leukemias have different etiologies and also suggest a role of folate in the development of childhood leukemia.

DNA Methylation, Isocitrate Dehydrogenase Mutation, and Survival in Glioma

BACKGROUND Although much is known about molecular and chromosomal characteristics that distinguish glioma histological subtypes, DNA methylation patterns of gliomas and their association with other tumor features such as mutation of isocitrate dehydrogenase (IDH) genes have only recently begun to be investigated. METHODS DNA methylation of glioblastomas, astrocytomas, oligodendrogliomas, oligoastrocytomas, ependymomas, and pilocytic astrocytomas (n = 131) from the Brain Tumor Research Center at the University of California San Francisco, as well as nontumor brain tissues (n = 7), was assessed with the Illumina GoldenGate methylation array. Methylation data were subjected to recursively partitioned mixture modeling (RPMM) to derive methylation classes. Differential DNA methylation between tumor and nontumor was also assessed. The association between methylation class and IDH mutation (IDH1 and IDH2) was tested using univariate and multivariable analysis for tumors (n = 95) with available substrate for sequencing. Survival of glioma patients carrying mutant IDH (n = 57) was compared with patients carrying wild-type IDH (n = 38) using a multivariable Cox proportional hazards model and Kaplan-Meier analysis. All statistical tests were two-sided. RESULTS We observed a statistically significant association between RPMM methylation class and glioma histological subtype (P < 2.2 × 10(-16)). Compared with nontumor brain tissues, across glioma tumor histological subtypes, the differential methylation ratios of CpG loci were statistically significantly different (permutation P < .0001). Methylation class was strongly associated with IDH mutation in gliomas (P = 3.0 × 10(-16)). Compared with glioma patients whose tumors harbored wild-type IDH, patients whose tumors harbored mutant IDH showed statistically significantly improved survival (hazard ratio of death = 0.27, 95% confidence interval = 0.10 to 0.72). CONCLUSION The homogeneity of methylation classes for gliomas with IDH mutation, despite their histological diversity, suggests that IDH mutation is associated with a distinct DNA methylation phenotype and an altered metabolic profile in glioma.