Epigenetic alterations are a common event in lung cancer and their identification can serve to inform on the carcinogenic process and provide clinically relevant biomarkers. Using paired tumor and non-tumor lung tissues from 146 individuals from three independent populations we sought to identify common changes in DNA methylation associated with the development of non-small cell lung cancer. Pathologically normal lung tissue taken at the time of cancer resection was matched to tumorous lung tissue and together were probed for methylation using Illumina GoldenGate arrays in the discovery set (n = 47 pairs) followed by bisulfite pyrosequencing for validation sets (n = 99 pairs). For each matched pair the change in methylation at each CpG was calculated (the odds ratio), and these ratios were averaged across individuals and ranked by magnitude to identify the CpGs with the greatest change in methylation associated with tumor development. We identified the top gene-loci representing an increase in methylation (HOXA9, 10.3-fold and SOX1, 5.9-fold) and decrease in methylation (DDR1, 8.1-fold). In replication testing sets, methylation was higher in tumors for HOXA9 (p < 2.2 × 10−16) and SOX1 (p < 2.2 × 10−16) and lower for DDR1 (p < 2.2 × 10−16). The magnitude and strength of these changes were consistent across squamous cell and adenocarcinoma tumors. Our data indicate that the identified genes consistently have altered methylation in lung tumors. Our identified genes should be included in translational studies that aim to develop screening for early disease detection.
DNA Methylation; goldengate; lung cancer; molecular epidemiology; pyrosequencing
Studies in European and East Asian populations have identified lung cancer susceptibility loci in nicotinic acetylcholine receptor (nAChR) genes on chromosome 15q25.1 which also appear to influence smoking behaviors. We sought to determine if genetic variation in nAChR genes influences lung cancer susceptibly in African-Americans, and evaluated the association of these cancer susceptibility loci with smoking behavior. A total of 1308 African-Americans with lung cancer and 1241 African-American controls from three centers were genotyped for 378 single nucleotide polymorphisms (SNPs) spanning the sixteen human nAChR genes. Associations between SNPs and the risk of lung cancer were estimated using logistic regression, adjusted for relevant covariates. Seven SNPs in three nAChR genes were significantly associated with lung cancer at a strict Bonferroni-corrected level, including a novel association on chromosome 2 near the promoter of CHRNA1 (rs3755486: OR = 1.40, 95% CI = 1.18-1.67, P = 1.0 × 10−4). Association analysis of an additional 305 imputed SNPs on 2q31.1 supported this association. Publicly available expression data demonstrated that the rs3755486 risk allele correlates with increased CHRNA1 gene expression. Additional SNP associations were observed on 15q25.1 in genes previously associated with lung cancer, including a missense variant in CHRNA5 (rs16969968: OR = 1.60, 95% CI = 1.27-2.01, P = 5.9 × 10−5). Risk alleles on 15q25.1 also correlated with an increased number of cigarettes smoked per day among the controls. These findings identify a novel lung cancer risk locus on 2q31.1 which correlates with CHRNA1 expression and replicate previous associations on 15q25.1 in African-Americans.
Lung cancer; nicotine dependence; African-Americans; genetic association; smoking
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.
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.
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).
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.
Acute lymphoblastic leukemia (ALL) likely has a multistep etiology, with initial genetic aberrations occurring early in life. An abnormal immune response to common infections has emerged as a plausible candidate for triggering the proliferation of pre-leukemic clones and the fixation of secondary genetic mutations and epigenetic alterations. We investigated whether evidence of infection with a specific common myelotropic childhood virus, parvovirus B19 (PVB19), relates to patterns of gene promoter DNA methylation in ALL patients. We serologically tested bone marrow samples at diagnosis of B-cell ALL for PVB19 infection and DNA methylation using a high-throughput bead array and found that 4.2% and 36.7% of samples were seroreactive to PVB19 IgM and IgG, respectively. Leukemia samples were grouped by DNA methylation pattern. Controlling for age and immunophenotype, unsupervised modeling confirmed that the DNA methylation pattern was associated with history of PVB19 (assessed by IgG, p = 0.02), but not recent infection (assessed by IgM). Replication assays on single genes were consistent with the association. The data indicate that a common viral illness may drive specific DNA methylation patterns in susceptible B-precursor cells, contributing to the leukemogenic potential of such cells. Infections may impact childhood leukemia by altering DNA methylation patterns and specific key genes in susceptible cells; these changes may be retained even after the clearance of infection.
childhood leukemia; DNA methylation; parvovirus B19; serology
Approximately 500,000 individuals diagnosed with bladder cancer in the U.S. require routine cystoscopic follow-up to monitor for disease recurrences or progression, resulting in over $2 billion in annual expenditures. Identification of new diagnostic and monitoring strategies are clearly needed, and markers related to DNA methylation alterations hold great promise due to their stability, objective measurement, and known associations with the disease and with its clinical features. To identify novel epigenetic markers of aggressive bladder cancer, we utilized a high-throughput DNA methylation bead-array in two distinct population-based series of incident bladder cancer (n = 73 and n = 264, respectively). We then validated the association between methylation of these candidate loci with tumor grade in a third population (n = 245) through bisulfite pyrosequencing of candidate loci. Array based analyses identified 5 loci for further confirmation with bisulfite pyrosequencing. We identified and confirmed that increased promoter methylation of HOXB2 is significantly and independently associated with invasive bladder cancer and methylation of HOXB2, KRT13 and FRZB together significantly predict high-grade non-invasive disease. Methylation of these genes may be useful as clinical markers of the disease and may point to genes and pathways worthy of additional examination as novel targets for therapeutic treatment.
Motivation: Integration of various genome-scale measures of molecular alterations is of great interest to researchers aiming to better define disease processes or identify novel targets with clinical utility. Particularly important in cancer are measures of gene copy number DNA methylation. However, copy number variation may bias the measurement of DNA methylation. To investigate possible bias, we analyzed integrated data obtained from 19 head and neck squamous cell carcinoma (HNSCC) tumors and 23 mesothelioma tumors.
Results: Statistical analysis of observational data produced results consistent with those anticipated from theoretical mathematical properties. Average beta value reported by Illumina GoldenGate (a bead-array platform) was significantly smaller than a similar measure constructed from the ratio of average dye intensities. Among CpGs that had only small variations in measured methylation across tumors (filtering out clearly biological methylation signatures), there were no systematic copy number effects on methylation for three and more than four copies; however, one copy led to small systematic negative effects, and no copies led to substantial significant negative effects.
Conclusions: Since mathematical considerations suggest little bias in methylation assayed using bead-arrays, the consistency of observational data with anticipated properties suggests little bias. However, further analysis of systematic copy number effects across CpGs suggest that though there may be little bias when there are copy number gains, small biases may result when one allele is lost, and substantial biases when both alleles are lost. These results suggest that further integration of these measures can be useful for characterizing the biological relationships between these somatic events.
Supplementary information: Supplementary data are available at Bioinformatics online.
Pathologic differentiation of tissue of origin in tumors found in the lung can be challenging, with differentiation of mesothelioma and lung adenocarcinoma emblematic of this problem. Indeed, proper classification is essential for determination of treatment regimen for these diseases, making accurate and early diagnosis critical. Here we investigate the potential of epigenetic profiles of lung adenocarcinoma, mesothelioma, and non-malignant pulmonary tissues (n=285) as differentiation markers in an analysis of DNA methylation at 1413 autosomal CpG loci associated with 773 cancer-related genes. Using an unsupervised recursively-partitioned mixture modeling technique for all samples, the derived methylation profile classes were significantly associated with sample type (P < 0.0001). In a similar analysis restricted to tumors, methylation profile classes significantly predicted tumor type (P < 0.0001). Random forests classification of CpG methylation of tumors - which splits the data into training and test sets - accurately differentiated MPM from lung adenocarcinoma over 99% of the time (P < 0.0001). In a locus-by-locus comparison of CpG methylation between tumor types, 1266 CpG loci had significantly different methylation between tumors following correction for multiple comparisons (Q < 0.05); 61% had higher methylation in adenocarcinoma. Using the CpG loci with significant differential methylation in a pathways analysis revealed significant enrichment of methylated gene-loci in Cell Cycle Regulation, DNA Damage Response, PTEN Signaling, and Apoptosis Signaling pathways in lung adenocarcinoma when compared to mesothelioma. Methylation-profile-based differentiation of lung adenocarcinoma and mesothelioma is highly accurate, informs on the distinct etiologies of these diseases, and holds promise for clinical application.
Head and neck squamous cell carcinomas (HNSCCs) represent clinically and etiologically heterogeneous tumors affecting >40 000 patients per year in the USA. Previous research has identified individual epigenetic alterations and, in some cases, the relationship of these alterations with carcinogen exposure or patient outcomes, suggesting that specific exposures give rise to specific types of molecular alterations in HNSCCs. Here, we describe how different etiologic factors are reflected in the molecular character and clinical outcome of these tumors. In a case series of primary, incident HNSCC (n = 68), we examined the DNA methylation profile of 1413 autosomal CpG loci in 773 genes, in relation to exposures and etiologic factors. The overall pattern of epigenetic alteration could significantly distinguish tumor from normal head and neck epithelial tissues (P < 0.0001) more effectively than specific gene methylation events. Among tumors, there were significant associations between specific DNA methylation profile classes and tobacco smoking and alcohol exposures. Although there was a significant association between methylation profile and tumor stage (P < 0.01), we did not observe an association between these profiles and overall patient survival after adjustment for stage; although methylation of a number of specific loci falling in different cellular pathways was associated with overall patient survival. We found that the etiologic heterogeneity of HNSCC is reflected in specific patterns of molecular epigenetic alterations within the tumors and that the DNA methylation profiles may hold clinical promise worthy of further study.
Mechanisms of action of non-mutagenic carcinogens such as asbestos remain poorly characterized. As pleural mesothelioma is known to have limited numbers of genetic mutations, we aimed to characterize the relationships among gene-locus specific methylation alterations, disease status, asbestos burden, and survival in this rapidly-fatal asbestos-associated tumor. Methylation of 1505 CpG loci associated with 803 cancer-related genes were studied in 158 pleural mesotheliomas and 18 normal pleura. After false-discovery rate correction, 969 CpG loci were independently associated with disease status (Q < 0.05). Classifying samples based upon CpG methylation profile with a mixture model approach, methylation classes discriminated tumor from normal pleura (permutation P < 0.0001). In a random forests classification the overall misclassification error rate was 3.4%, with <1% (n=1) of tumors misclassified as normal (P < 0.0001). Among tumors, methylation class membership was significantly associated with lung tissue asbestos body burden (P < 0.03), and significantly predicted survival (likelihood ratio P < 0.01). Consistent with prior work, asbestos burden was associated with an increased risk of death (HR = 1.4, 95% CI, 1.1 – 1.8). Our results have shown that methylation profiles powerfully differentiate diseased pleura from non-tumor pleura and that asbestos burden and methylation profiles are independent predictors of mesothelioma patient survival. We have added to the growing body of evidence that cellular epigenetic dysregulation is a critical mode of action for asbestos in the induction of pleural mesothelioma. Importantly, these findings hold great promise for using epigenetic profiling in the diagnosis and prognosis of human cancers.
Methylation; asbestos; mesothelioma
Although tumor size and lymph node involvement are the current cornerstones of breast cancer prognosis, they have not been extensively explored in relation to tumor methylation attributes in conjunction with other tumor and patient dietary and hormonal characteristics. Using primary breast tumors from 162 (AJCC stage I–IV) women from the Kaiser Division of Research Pathways Study and the Illumina GoldenGate methylation bead-array platform, we measured 1,413 autosomal CpG loci associated with 773 cancer-related genes and validated select CpG loci with Sequenom EpiTYPER. Tumor grade, size, estrogen and progesterone receptor status, and triple negative status were significantly (Q-values <0.05) associated with altered methylation of 209, 74, 183, 69, and 130 loci, respectively. Unsupervised clustering, using a recursively partitioned mixture model (RPMM), of all autosomal CpG loci revealed eight distinct methylation classes. Methylation class membership was significantly associated with patient race (P<0.02) and tumor size (P<0.001) in univariate tests. Using multinomial logistic regression to adjust for potential confounders, patient age and tumor size, as well as known disease risk factors of alcohol intake and total dietary folate, were all significantly (P<0.0001) associated with methylation class membership. Breast cancer prognostic characteristics and risk-related exposures appear to be associated with gene-specific tumor methylation, as well as overall methylation patterns.
The current standard prognostic indicator for breast cancer is tumor-node-metastasis staging; though, as population-based studies and clinical trials are conducted, molecular characterization of disease is beginning to allow improved markers of prognosis and assist clinicians in choosing the most appropriate therapies. We investigated DNA methylation profiles in over 160 well annotated breast tumor samples and found significant relationships with standard and other known predictors of prognosis, as well as established risk factors for disease: alcohol intake and dietary folate. Recently the United States National Cancer Institute Cancer Biomarkers Research Group articulated a need for a “Strategic Approach to Validating Methylated Genes as Biomarkers for Breast Cancer,” and our work is extremely responsive to this call for a national strategy. Recognizing the increasing use of pre-operative chemotherapy for patients with operable, early-stage disease, there is added complexity in breast cancer staging. Since chemotherapy can considerably decrease tumor size, it is still unclear whether pre-operative or post-operative stage best informs prognosis and treatment decisions for patients electing pre-operative chemotherapy. However, our data clearly illustrate the promise of tumor DNA methylation for augmenting tumor staging and can be attained with minimal tissue in a pre-operative 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.
The causes and extent of tissue-specific interindividual variation in human epigenomes are underappreciated and, hence, poorly characterized. We surveyed over 200 carefully annotated human tissue samples from ten anatosites at 1,413 CpGs for methylation alterations to appraise the nature of phenotypically, and hence potentially clinically important epigenomic alterations. Within tissue types, across individuals, we found variation in methylation that was significantly related to aging and environmental exposures such as tobacco smoking. Individual variation in age- and exposure-related methylation may significantly contribute to increased susceptibility to several diseases. As the NIH–funded HapMap project is critically contributing to annotating the human reference genome defining normal genetic variability, our work raises key issues to consider in the construction of reference epigenomes. It is well recognized that understanding genetic variation is essential to understanding disease. Our work, and the known interplay of epigenetics and genetics, makes it equally clear that a more complete characterization of epigenetic variation and its sources must be accomplished to reach the goal of a complete understanding of disease. Additional research is absolutely necessary to define the mechanisms controlling epigenomic variation. We have begun to lay the foundations for essential normal tissue controls for comparison to diseased tissue, which will allow the identification of the most crucial disease-related alterations and provide more robust targets for novel treatments.
Considerable controversy surrounds the carcinogenic potential of asphalt and tar. Since minority individuals may have had relatively high historical exposures, we investigated asphalt and tar exposure and lung cancer risk among African Americans and Latino Americans.
We conducted a case-control study of lung cancer among African Americans and Latino Americans in the San Francisco Bay area (422 cases, 894 controls). A questionnaire was used to obtain detailed work histories and exposure information. Self-reported exposure to asphalt and tar as well as other factors (eg. smoking, automobile exhaust, and asbestos) were evaluated as predictors of lung cancer risk. Potential effect modification by cytochrome P450 (CYP) 1A1 was also explored.
Self-reported duration of exposure to asphalt and tar was associated with a statistically significant excess risk of lung cancer in the overall population (OR: 1.11, 95%CI: 1.01–1.22), evaluating risk per year of exposure. Years of exposure to automobile exhaust (OR: 1.02, 95%CI: 1.00–1.05) and asbestos (OR: 1.04, 95%CI: 1.02–1.06) were also associated with statistically significant elevations in risk. In Latino Americans, the lung cancer risks associated with polycyclic aromatic hydrocarbon-related exposures were consistently higher in the CYP1A1 wildtype subjects as compared to the variant genotype subjects, and the interaction was statistically significant for smoking and the CYP1A1 M2 polymorphism (p-valueinteraction=0.02).
These data are consistent with the literature suggesting that exposure to asphalt and tar may increase risk of lung cancer. However, it was not possible to separate the effects and asphalt and tar in this study.
asphalt; tar; lung cancer; minorities
Allergy history has been consistently inversely associated with glioma risk. Two serologic markers, soluble CD23 (sCD23) and soluble CD14 (sCD14), are part of the innate and adaptive humoral immune systems and modulate allergic responses in opposite directions, with sCD23 enhancing and sCD14 blunting inflammatory responses. We measured sCD23 and sCD14 in serum from blood that was drawn at a single time point from 1079 glioma patients post diagnosis and 736 healthy controls. Glioma was strongly associated with high sCD14 (highest vs. lowest quartile OR = 3.94 (95% CI: 2.98-5.21) and low sCD23 (lowest vs. highest quartile OR=2.5 (95% CI: 1.89-3.23)). Results were consistent across glioma histologic types and grades, but were strongest for glioblastoma. While temozolomide treatment was not associated with either sCD14 or sCD23 levels among cases, those taking dexamethasone had somewhat lower sCD23 levels than those not taking dexamethasone. However, sCD23 was associated with case status regardless of dexamethasone treatment. These results augment the long observed association between allergies and glioma and support a role for the innate and adaptive humoral functions of the immune system, and in particular immunoregulatory proteins, in gliomagenesis.
CD14; CD23; glioma; brain tumor; immune; tumor risk; epidemiology
We explored the associations of aberrant DNA methylation patterns in 12 candidate genes with adult glioma subtype, patient survival, and gene expression of enhancer of zeste human homolog 2 (EZH2) and insulin-like growth factor-binding protein 2 (IGFBP2). We analyzed 154 primary glioma tumors (37 astrocytoma II and III, 52 primary glioblastoma multiforme (GBM), 11 secondary GBM, 54 oligodendroglioma/oligoastrocytoma II and III) and 13 nonmalignant brain tissues for aberrant methylation with quantitative methylation-specific PCR (qMS-PCR) and for EZH2 and IGFBP2 expression with quantitative reverse transcription PCR (qRT-PCR). Global methylation was assessed by measuring long interspersed nuclear element-1 (LINE1) methylation. Unsupervised clustering analyses yielded 3 methylation patterns (classes). Class 1 (MGMT, PTEN, RASSF1A, TMS1, ZNF342, EMP3, SOCS1, RFX1) was highly methylated in 82% (75/91) of lower-grade astrocytic and oligodendroglial tumors, 73% (8/11) of secondary GBMs, and 12% (6/52) of primary GBMs. The primary GBMs in this class were early onset (median age 37 years). Class 2 (HOXA9 and SLIT2) was highly methylated in 37% (19/52) of primary GBMs. None of the 10 genes for class 3 that were differentially methylated in classes 1 and 2 were hypermethylated in 92% (12/13) of nonmalignant brain tissues and 52% (27/52) of primary GBMs. Class 1 tumors had elevated EZH2 expression but not elevated IGFBP2; class 2 tumors had both high IGFBP2 and high EZH2 expressions. The gene-specific hypermethylation class correlated with higher levels of global LINE1 methylation and longer patient survival times. These findings indicate a generalized hypermethylation phenotype in glioma linked to improved survival and low IGFBP2. DNA methylation markers are useful in characterizing distinct glioma subtypes and may hold promise for clinical applications.
glioma; DNA methylation; EZH2; Polycomb; PI3K/Akt
Lung cancer is the leading cause of cancer death among US Asian/Pacific Islander (API) and Latina women, despite low smoking prevalence. This study examined survival patterns following non-small cell lung cancer in a population-based sample of lung cancer cases from the San Francisco Bay Area Lung Cancer Study (SFBALCS).
Women diagnosed with lung cancer from 1998–2003 and 2005–2008 and identified through the Greater Bay Area Cancer Registry were telephone-screened for eligibility for the SFBALCS. The screener data were linked to the cancer registry data to determine follow-up. This analysis included 187 non-Hispanic White, 23 US-born Latina, 32 foreign-born Latina, 30 US-born API, and 190 foreign-born API never smokers diagnosed with lung cancer and followed through 2008.
All-cause survival was poorer among APIs (hazard ratio (HR) and 95% confidence interval (CI) = 1.7 (1.0–2.8) among US-born APIs; 1.2 (0.9–1.5) among foreign-born APIs), and Latinas (HR (95% CI) = 2.1 (1.2–3.6) among US-born Latinas; 1.4 (0.9–2.3) among foreign-born Latinas), relative to non-Hispanic Whites. These survival differences were not explained by differences in selected sociodemographic or clinical factors.
Further research should focus on factors such as cultural behaviors, access to or attitudes toward health care, and genetic variations, as possible explanations for these striking racial/ethnic differences.
Latina and API female never smokers diagnosed with lung cancer were up to two-times more likely to die than non-Hispanic Whites, highlighting the need for additional research to identify the underlying reasons for the disparities, as well as heightened clinical awareness.
lung cancer survival; Asian; Latina; Hispanic; never smokers; nativity
Oligodendroglial tumors are rare subtypes of brain tumors and are often combined with other glial tumors in epidemiological analyses. However, different demographic associations and clinical characteristics suggest potentially different risk factors. The purpose of this study was to investigate possible risk factors for oligodendroglial tumors (including oligodendroglioma, anaplastic oligodendroglioma, and mixed glioma). Data from 7 case–control studies (5 US and 2 Scandinavian) were pooled. Unconditional logistic regression was used to calculate odds ratios (ORs) and 95% confidence intervals (CIs), adjusted for age group, gender, and study site. Data on 617 cases and 1260 controls were available for analyses. Using data from all 7 studies, history of allergies and/or asthma was associated with a decreased risk of anaplastic oligodendroglioma (OR = 0.6; 95% CI: 0.4–0.9), and history of asthma only was associated with a decreased risk of oligodendroglioma (OR = 0.5; 95% CI: 0.3–0.9) and anaplastic oligodendroglioma (OR = 0.3; 95% CI: 0.1–0.9). A family history of brain tumors was associated with an increased risk of anaplastic oligodendroglioma (OR = 2.2; 95% CI: 1.1–4.5). Having had chicken pox was associated with a decreased risk of oligodendroglioma (OR = 0.6; 95% CI: 0.4–0.9) and anaplastic oligodendroglioma (OR = 0.5; 95% CI: 0.3–0.9) in the US studies. Although there is some overlap in risk factors between oligodendroglial tumors and gliomas as a group, it is likely that additional factors specific to oligodendroglial tumors have yet to be identified. Large, multi-institution international studies will be necessary to better characterize these etiological risk factors.
anaplastic oligodendroglioma; epidemiology; mixed glioma; oligodendroglioma; risk factors
Two recent genome-wide association studies reported that single nucleotide polymorphisms (SNPs) in (or near) TERT (5p15), CCDC26 (8q24), CDKN2A/B (9p21), PHLDB1 (11q23), and RTEL1 (20q13) are associated with infiltrating glioma. From these reports it was not clear if the SNP associations predispose to glioma in general or whether they are specific to certain glioma grades or morphologic subtypes. To identify hypothesized associations between susceptibility loci and tumor subtype, we genotyped two case/control groups composed of the spectrum of infiltrating glioma subtypes, and stratified the analyses by type. We report that specific germline polymorphisms are associated with different glioma subtypes. CCDC26 (8q24) region polymorphisms are strongly associated with oligodendroglial tumor risk (rs4295627, OR=2.05, p=8.3*10−11), but not glioblastoma risk. The opposite is true of RTEL (20q13) region polymorphisms which are significantly associated with glioblastoma (rs2297440, OR = 0.56, p= 4.6*10−10) but not oligodendroglial tumor. The SNPs in or near CCDC26 (8q24) are associated with oligodendroglial tumors regardless of combined 1p and 19q deletion status; however, the association is greatest for those with combined deletion (rs4295627, OR=2.77, p=2.6*10-9). These observations generate hypotheses concerning the possible mechanisms by which specific SNPs (or alterations in linkage disequilibrium with such SNPs) are associated with glioma development.
Glioma; Genetic Association; 1p/19q deletion
In genome-wide association studies, inherited risk of glioma has been demonstrated for rare familial syndromes and with common variants from 3–5 chromosomal regions. To assess the degree of familial aggregation of glioma, the authors performed a pooled analysis of data from 2 large glioma case-control studies in the United States (MD Anderson Cancer Center, Houston, Texas (1994–2006) and University of California, San Francisco (1991–2004)) and from the Swedish Cancer Registry (1958–2006) to measure excess cases of cancer among first-degree relatives of glioma probands. This analysis included 20,377 probands with glioma and 52,714 first-degree relatives. No overall increase was found in the expected number of cancers among family members; however, there were 77% more gliomas than expected. There were also significantly more sarcoma and melanoma cases than expected, which is supported by evidence in the literature, whereas there were significantly fewer-than-expected cases of leukemia, non-Hodgkin lymphoma, and bladder, lung, pancreatic, prostate, and uterine cancers. This large pooled analysis provided sufficient numbers of related family members to examine the genetic mechanisms involved in the aggregation of glioma with other cancers in these families. However, misclassification due to unvalidated cancers among family members could account for the differences seen by study site.
family; glioma; meta-analysis; neoplasms
A growing body of evidence supports the significant interplay between the immune system and glioma pathogenesis. Here we investigate whether the extent of local glioma-associated CD8+ T-cell infiltrate at initial presentation correlates with long-term survival in patients with glioblastoma multiforme (GBM). The study was conducted by the University of California San Francisco Brain Tumor Research Center as part of the San Francisco Bay Area Adult Glioma Study, which included over 519 patients with GBM. A central neuropathology review was performed and populations of infiltrating CD8+ T-cells were quantified histologically. Of 108 patients studied, 43 patients had poor survival (< 95 days) and 65 patients had extended long-term survival of > 403 days. Tumors from long-term survivors were more likely than short-term survivors to have intermediate or extensive T-cell infiltrates compared to focal or rare infiltrates, and this association appears to be most significant in Caucasian women (p < 0.006). Thus, CD8+ T-cell infiltrate is associated with prolonged survival. Our data provide the impetus for more sophisticated studies to further elucidate prospectively the specific T-cell subtypes associated with long-term survival.
Clinical outcome; Glioma; Glioblastoma; Immune infiltrate; T-cell infiltrate
Several genome-wide association studies identified the chr15q25.1 region, which includes three nicotinic cholinergic receptor genes (CHRNA5-B4) and the cell proliferation gene (PSMA4), for its association with lung cancer risk in Caucasians. A haplotype and its tagging single nucleotide polymorphisms (SNPs) encompassing six genes from IREB2 to CHRNB4 were most strongly associated with lung cancer risk (OR = 1.3; P < 10−20). In order to narrow the region of association and identify potential causal variations, we performed a fine-mapping study using 77 SNPs in a 194 kb segment of the 15q25.1 region in a sample of 448 African-American lung cancer cases and 611 controls. Four regions, two SNPs and two distinct haplotypes from sliding window analyses, were associated with lung cancer. CHRNA5 rs17486278 G had OR = 1.28, 95% CI 1.07–1.54 and P = 0.008, whereas CHRNB4 rs7178270 G had OR = 0.78, 95% CI 0.66–0.94 and P = 0.008 for lung cancer risk. Lung cancer associations remained significant after pack-year adjustment. Rs7178270 decreased lung cancer risk in women but not in men; gender interaction P = 0.009. For two SNPs (rs7168796 A/G and rs7164594 A/G) upstream of PSMA4, lung cancer risks for people with haplotypes GG and AA were reduced compared with those with AG (OR = 0.56, 95% CI 0.38–0.82; P = 0.003 and OR = 0.73, 95% CI 0.59–0.90, P = 0.004, respectively). A four-SNP haplotype spanning CHRNA5 (rs11637635 C, rs17408276 T, rs16969968 G) and CHRNA3 (rs578776 G) was associated with increased lung cancer risk (P = 0.002). The identified regions contain SNPs predicted to affect gene regulation. There are multiple lung cancer risk loci in the 15q25.1 region in African-Americans.
Prior studies of lung cancer and CYP1A1/2 in African American and Latino populations have shown inconsistent results and not yet investigated the haplotype block structure of CYP1A1/2 or addressed potential population stratification. To investigate haplotypes in the CYP1A1/2 region and lung cancer in African Americans and Latinos, we conducted a case-control study (1998-2003). African Americans (N = 535) and Latinos (N = 412) were frequency-matched on age, sex, and self-reported race/ethnicity. We used a custom genotyping panel containing 50 single nucleotide polymorphisms in the CYP1A1/2 region and 184 ancestry informative markers selected to have large allele frequency differences between Africans, Europeans, and Amerindians. Latinos exhibited significant haplotype main effects in two blocks, even after adjusting for admixture (odds ratio (OR) = 2.02, 95% confidence interval (CI): 1.28 – 3.19 and OR = 0.55, 95% CI: 0.36 – 0.83) but no main effects were found among African Americans. Adjustment for admixture revealed substantial confounding by population stratification among Latinos but not African Americans. Among Latinos and African Americans interactions between smoking level and haplotypes were not statistically significant. Evidence of population stratification among Latinos underscores the importance of adjusting for admixture in lung cancer association studies, particularly in Latino populations. These results suggest a variant occurring within the CYP1A2 region may be conferring an increased risk of lung cancer in Latinos.
lung cancer; haplotype; CYP1A1/2; population stratification; admixed
Base excision repair (BER) is the primary DNA damage repair mechanism for repairing small base lesions resulting from oxidation and alkylation damage. This study examines the association between 24 single-nucleotide polymorphisms (SNPs) belonging to five BER genes (XRCC1, APEX1, PARP1, MUTYH and OGG1) and lung cancer among Latinos (113 cases and 299 controls) and African-Americans (255 cases and 280 controls). The goal was to evaluate the differences in genetic contribution to lung cancer risk by ethnic groups. Analyses of individual SNPs and haplotypes were performed using unconditional logistic regressions adjusted for age, sex and genetic ancestry. Four SNPs among Latinos and one SNP among African-Americans were significantly (P < 0.05) associated with either risk of all lung cancer or non-small cell lung cancer (NSCLC). However, only the association between XRCC1 Arg399Gln (rs25487) and NSCLC among Latinos (odds ratio associated with every copy of Gln = 1.52; 95% confidence interval: 1.01–2.28) had a false-positive report probability of <0.5. Arg399Gln is a SNP with some functional evidence and has been shown previously to be an important SNP associated with lung cancer, mostly for Asians. Since the analyses were adjusted for genetic ancestry, the observed association between Arg399Gln and NSCLC among Latinos is unlikely to be confounded by population stratification; however, this result needs to be confirmed by additional studies among the Latino population. This study suggests that there are genetic differences in the association between BER pathway and lung cancer between Latinos and African-Americans.
A variety of methods are available for estimating genetic admixture proportions in populations; however, few investigators have conducted detailed comparisons using empirical data. The authors characterized admixture proportions among self-identified African Americans (n = 535) and Latinos (n = 412) living in the San Francisco Bay Area who participated in a lung cancer case-control study (1998–2003). Individual estimates of genetic ancestry based on 184 informative markers were obtained from a Bayesian approach and 2 maximum likelihood approaches and were compared using descriptive statistics, Pearson correlation coefficients, and Bland-Altman plots. Case-control differences in individual admixture proportions were assessed using 2-sample t tests and logistic regression analysis. Results indicated that Bayesian and frequentist approaches to estimating admixture provide similar estimates and inferences. No difference was observed in admixture proportions between African-American cases and controls, but Latino cases and controls significantly differed according to Amerindian and European genetic ancestry. Differences in admixture proportions between Latino cases and controls were not unexpected, since cases were more likely to have been born in the United States. Genetic admixture proportions provide a quantitative measure of ancestry differences among Latinos that can be used in analyses of genetic risk factors.
African Americans; case-control studies; epidemiologic methods; genetics, population; Hispanic Americans; linkage disequilibrium; lung neoplasms; statistics
Few studies on the association between nucleotide excision repair (NER) variants and lung cancer risk have included Latinos and African Americans. We examine variants in six NER genes (ERCC2, ERCC4, ERCC5, LIG1, RAD23B and XPC) in association with primary lung cancer risk among 113 Latino and 255 African American subjects newly diagnosed with primary lung cancer from 1998 to 2003 in the San Francisco Bay Area, and 579 healthy controls (299 Latinos and 280 African Americans). Individual single nucleotide polymorphism and haplotype analyses, multifactor dimensionality reduction, and principal components analysis were performed to assess the association between six genes in the NER pathway and lung cancer risk. Among Latinos, ERCC2 haplotype CGA (rs238406, rs11878644, rs6966) was associated with reduced lung cancer risk [odds ratio (OR) of 0.65 and 95% confidence interval (CI): 0.44-0.97], especially among non-smokers (OR=0.29; 95% CI: 0.12-0.67). From multifactor dimensionality reduction analysis, in Latinos, smoking and three SNPs (ERCC2 rs171140, ERCC5 rs17655, and LIG1 rs20581) together had a prediction accuracy of 67.4% (p=0.001) for lung cancer. Among African Americans, His/His genotype of ERCC5 His1104Asp (rs17655) was associated with increased lung cancer risk (OR=1.78; 95% CI: 1.09-2.91), and LIG1 haplotype GGGAA (rs20581, rs156641, rs3730931, rs20579, and rs439132) was associated with reduced lung cancer risk (OR=0.61; 95% CI: 0.42-0.88). Our study suggests different elements of the NER pathway may be important in the different ethnic groups resulting either from different linkage relationship, genetic backgrounds, and/or exposure histories.
nucleotide excision repair; DNA repair; lung cancer; African Americans; Latinos
Glioblastoma multiforme (GBM) can present as either de novo or secondary tumors arising from previously diagnosed low-grade gliomas. Although these tumor types are phenotypically indistinguishable, de novo and secondary GBMs are associated with distinct genetic characteristics. PTEN mutations, which result in activation of the phosphoinositide 3-kinase (PI3K) signal transduction pathway, are frequent in de novo but not in secondary GBMs or their antecedent low-grade tumors. Results we present here show that grade II astrocytomas, oligodendrogliomas, and oligoastrocytomas commonly display methylation of the PTEN promoter, a finding that is absent in nontumor brain specimens and rare in de novo GBMs. Methylation of the PTEN promoter correlates with protein kinase B (PKB/Akt) phosphorylation, reflecting functional activation of the PI3K pathway. Our results also demonstrate frequent methylation of the PTEN promoter in grade III astrocytomas and secondary GBMs, consistent with the hypothesis that these tumors arise from lower grade precursors. PTEN methylation is rare in de novo GBMs and is mutually exclusive with PTEN mutations. We conclude that methylation of the PTEN promoter may represent an alternate mechanism by which PI3K signaling is increased in grade II and III gliomas as well as secondary GBMs, a finding that offers new therapeutic approaches in these patients.
low-grade glioma; methylation; PKB/Akt; PTEN; secondary glioblastoma