Gregory Idos

Genome-wide association study of colorectal cancer identifies six new susceptibility loci

Genetic susceptibility to colorectal cancer is caused by rare pathogenic mutations and common genetic variants that contribute to familial risk. Here we report the results of a two-stage association study with 18,299 cases of colorectal cancer and 19,656 controls, with follow-up of the most statistically significant genetic loci in 4,725 cases and 9,969 controls from two Asian consortia. We describe six new susceptibility loci reaching a genome-wide threshold of P<5.0E-08. These findings provide additional insight into the underlying biological mechanisms of colorectal cancer and demonstrate the scientific value of large consortia-based genetic epidemiology studies.

Cloning of the cyclin A1 genomic structure and characterization of the promoter region. GC boxes are essential for cell cycle-regulated transcription of the cyclin A1 gene.

Cyclin A1 is a recently cloned cyclin with high level expression in meiotic cells in the testis. However, it is also frequently expressed at high levels in acute myeloid leukemia. To elucidate the regulation of cyclin A1 gene expression, we cloned and analyzed the genomic structure of cyclin A1. It consists of 9 exons within 13 kilobase pairs. The TATA-less promoter initiates transcription from several start sites with the majority of transcripts beginning within a 4-base pair stretch. A construct containing a fragment from −190 to +145 showed the highest transcriptional activity. Transfection of cyclin A1promoter constructs into S2 Drosophila cells demonstrated that Sp1 is essential for the activity of the promoter. Sp1, as well as Sp3, bound to four GC boxes between nucleotides −130 and −80 as observed by gel shift analysis. Mutations in two or more of the four GC boxes decreased promoter activity by >80%. The promoter was found to be cell cycle-regulated with highest activities found in late S and G2/M phase. Further analyses suggested that cell cycle regulation was accomplished by periodic repression of the GC boxes in G1 phase. Taken together, our data show that cyclin A1 promoter activity critically depends on four GC boxes, and members of the Sp1 family appear to be involved in directing expression of cyclin A1 in both a tissue- and cell cycle-specific manner.

RARβ2 is a candidate tumor suppressor gene in myelofibrosis with myeloid metaplasia

Myelofibrosis with myeloid metaplasia (MMM) is a clonal stem-cell disorder that leads to ineffective hematopoiesis, bone marrow fibrosis, and extramedullary hematopoiesis. The molecular mechanisms underlying the development of this myeloproliferative syndrome are currently unknown. In order to identify tumor suppressor genes that may be involved in the disease process, we performed an analysis for loss of heterozygosity (LOH) in CD34+ cells from 29 patients with MMM. We observed a frequency of allelic loss on chromosomal arm 3p in 24% of cases. Detailed mapping of 3p revealed a distinct region of deletion at 3p24. Among the genes known to map within this region is the retinoic acid receptor-β (RARβ2) gene. To determine whether RARβ2 gene activity is diminished in this disease, we analysed its expression in CD34+ cells from 17 patients with MMM using quantitative PCR. Our results indicate that expression of RARβ2 is significantly decreased in 100% of patient samples compared to that in CD34+ cells from 10 normal individuals. Since allelic loss at 3p24 occurs in <25% of patients, we investigated the contribution of epigenetic modifications to RARβ2 inactivity. Using methylation-specific PCR, we found hypermethylation of RARβ2 in 16 of 18 patients (89%), while the methylated form of the gene was absent in CD34+ cells from nine normal individuals. Our results suggest that RARβ2 acts as a tumor suppressor gene in MMM and that epigenetic changes are the most significant determinants of RARβ2 gene activity in these patients.

Genetic markers of malignant transformation in intraductal papillary mucinous neoplasm of the pancreas: a meta-analysis.

Objectives The objective of this study was to determine the relationship between specific genetic alterations and malignant transformation in intraductal papillary mucinous neoplasm (IPMN) of the pancreas. Methods Quantitative meta-analysis was conducted of studies through October 2010 that adhered to the 1996 World Health Organization guidelines for distinguishing adenoma and borderline IPMN versus carcinoma in surgically resected specimens using a random-effects model. We developed a 6-point scoring system to assess study quality. Results Thirty-nine studies (1235 IPMN samples) satisfied the inclusion criteria, and we conducted pooled analysis of 8 genetic markers: MUC1, MUC2, MUC5AC, kRas, p53, hTERT (human telomerase reverse transcriptase), cyclooxygenase 2, and Shh (Sonic hedgehog). Markers having the strongest association with malignant IPMN were hTERT (odds ratio [OR], 11.4; 95% confidence interval [CI], 3.5–36.7) and Shh (OR, 6.9; 95% CI, 2.4–20.2), whereas MUC5AC (OR, 1.0; 95% CI, 0.1–13.9) and kRas (OR, 2.0; 95% CI, 1.0–4.3) showed weak association with IPMN histologic progression. Conclusions Expression of hTERT is strongly associated with malignant transformation in IPMN, consistent with up-regulation of hTERT as a key step in progression of IPMN to cancer. Expression of kRas and MUC5AC is common but not strongly associated with IPMN histologic progression. The quality criteria used here may guide future reporting of genetic markers related to malignant transformation of IPMN.

Increased yield of actionable mutations using multi-gene panels to assess hereditary cancer susceptibility in an ethnically diverse clinical cohort

This study aims to assess multi-gene panel testing in an ethnically diverse clinical cancer genetics practice. We conducted a retrospective study of individuals with a personal or family history of cancer undergoing clinically indicated multi-gene panel tests of 6-110 genes, from six commercial laboratories. The 475 patients in the study included 228 Hispanics (47.6%), 166 non-Hispanic Whites (35.4%), 55 Asians (11.6%), 19 Blacks (4.0%), and seven others (1.5%). Panel testing found that 15.6% (74/475) of patients carried deleterious mutations for a total of 79 mutations identified. This included 7.4% (35/475) of patients who had a mutation identified that would not have been tested with a gene-by-gene approach. The identification of a panel-added mutation impacted clinical management for most of cases (69%, 24/35), and genetic testing was recommended for the first degree relatives of nearly all of them (91%, 32/35). Variants of uncertain significance (VUSs) were identified in a higher proportion of tests performed in ethnic minorities. Multi-gene panel testing increases the yield of mutations detected and adds to the capability of providing individualized cancer risk assessment. VUSs represent an interpretive challenge due to less data available outside of White, non-Hispanic populations. Further studies are necessary to expand understanding of the implementation and utilization of panels across broad clinical settings and patient populations.

A novel BAP1 mutation is associated with melanocytic neoplasms and thyroid cancer

Germline mutations in the tumor suppressor gene, BRCA-1 associated protein (BAP1), underlie a tumor predisposition syndrome characterized by increased risk for numerous cancers including uveal melanoma, melanocytic tumors and mesothelioma, among others. In the present study we report the identification of a novel germline BAP1 mutation, c.1777C>T, which produces a truncated BAP1 protein product and segregates with cancer. Family members with this mutation demonstrated a primary clinical phenotype of autosomal dominant, early-onset melanocytic neoplasms with immunohistochemistry (IHC) of these tumors demonstrating lack of BAP1 protein expression. In addition, family members harboring the BAP1 c.1777C>T germline mutation developed other neoplastic disease including thyroid cancer. IHC analysis of the thyroid cancer, as well, demonstrated loss of BAP1 protein expression. Our investigation identifies a new BAP1 mutation, further highlights the relevance of BAP1 as a clinically important tumor suppressor gene, and broadens the range of cancers associated with BAP1 inactivation. Further study will be required to understand the full scope of BAP1-associated neoplastic disease.

Unexpected CDH1 Mutations Identified on Multigene Panels Pose Clinical Management Challenges

PurposeMutations in the CDH1 gene confer up to an 80% lifetime risk of diffuse gastric cancer and up to a 60% lifetime risk of lobular breast cancer. Testing for CDH1 mutations is recommended for individuals who meet the International Gastric Cancer Linkage Consortium (IGCLC) guidelines. However, the interpretation of unexpected CDH1 mutations identified in patients who do not meet IGCLC criteria or do not have phenotypes suggestive of hereditary diffuse gastric cancer is clinically challenging. This study aims to describe phenotypes of CDH1 mutation carriers identified through multigene panel testing (MGPT) and to offer informed recommendations for medical management.Patients and MethodsThis cross-sectional prevalence study included all patients who underwent MGPT between March 2012 and September 2014 from a commercial laboratory (n = 26,936) and an academic medical center cancer genetics clinic (n = 318) to estimate CDH1 mutation prevalence and associated clinical phenotypes. CDH1 mutation carriers were...

Patient communication of cancer genetic test results in a diverse population

Research on the communication of genetic test results has focused predominately on non-Hispanic White (NHW) mutation-positive families with high-risk hereditary cancer conditions. Little is known about this process for racially and ethnically diverse individuals or for those with mutations in moderate risk genes. The communication behaviors of study participants who carry a gene mutation were analyzed 3 months after disclosure of genetic test results. Participants were queried about communication of their results, as part of a prospective study of multi-gene panel genetic testing. The responses of particpants who tested positive were analyzed by race/ethnicity and by level of cancer risk (high vs. moderate). Of the 216 mutation-positive study participants, 136 (63%) responded. Self-reported race/ethnicity was 46% NHW, 41% Hispanic, 10% Asian, and 2% Black. The majority (99.0%, n = 135) had shared their results with someone and 96% had told a family member (n = 130). Hispanic respondents were less likely to have told a healthcare provider about their results than NHW (29% vs. 68%, p < .0001). Asian respondents were less likely than NHW to encourage family members to undergo testing (OR = 0.1, p = .03); but Asian family members were more likely to undergo testing (OR = 8.0, p = .03). There were no differences in communication between those with a mutation in a high- or moderate-risk gene. Three months post genetic testing, communication of results was very high; 30% reported a family member underwent genetic testing. Further studies are needed to better understand the communication process in individuals from diverse racial/ethnic backgrounds.

Erratum: Genome-wide association study of colorectal cancer identifies six new susceptibility loci (Nature Communications 6:7138)

Corrigendum: Genome-wide association study of colorectal cancer identifies six new susceptibility loci

Genetic Markers of Malignant Transformation in IPMN of the Pancreas: A Meta-Analysis

Background: Intraductal papillary mucinous neoplasm (IPMN) of the pancreas has been recognized as an important precursor to pancreatic cancer. Since the WHO established a consensus in 1996 for grading IPMNs, numerous studies have characterized genetic mutations associated with these various histologic grades. However, the molecular basis for progression into cancer remains poorly understood. We hypothesize that malignant transformation of IPMN reflects a progression in molecular changes. To determine the risk of malignant transformation associated with specific genetic abnormalities, we performed themost comprehensive meta-analysis to date of genetic associations with IPMN grade. Methods: We conducted a meta-analysis reviewing Pubmed, Cochrane Library, and Embase databases from January 1996 thru October 2010. We included only studies that adhered to 1996 WHO guidelines for distinguishing adenoma and borderline IPMNs (IPM-A, IPM-B) vs. carcinoma in situ and invasive carcinoma (IPM-C) based on surgically resected specimens. Quantitative meta-analysis was performed of gene candidates using both fixedand random-effects models. The I2 test was used to evaluate study heterogeneity. Results: We identified 178 studies that satisfied the inclusion criteria (1,343 IPMN samples). In pooled analysis there was an overall positive association for IPM-C with MUC-1 expression (OR 5.90; 95% C.I. 2.5413.70), k-Ras mutations (OR 1.90; 95% C.I. 1.04-3.47), and p53 mutations (OR 2.00; 95% C.I. 0.80-4.98). The strongest positive association for IPM-C vs. IPM-A/B was with telomerase mutations (OR 15.95; 95% C.I. 6.32-40.25), consistent with this genetic abnormality occurring relatively later in the histologic progression of IPMN into cancer. Expression of MUC5AC was not associated with IPM-C vs. IPM-A/B (OR 0.71; 95% C.I. 0.23-2.23). We further summarize associations with 91 additional genes including Shh, Cox2, Smad4, APC, and S100A4 as well as findings from cyst and pancreatic fluid assays. Conclusion: This metaanalysis identifies MUC-1 expression and mutations in telomerase, p53 and k-Ras genes as molecular changes associated with IPMN progression to cancer. These associations provide a basis for the use of molecular markers in diagnostic grading of IPMNs as well as a basis for therapeutic targets.