Paula K. Greer

BACKGROUND: Malignant cells must maintain their telomeres, but genetic mechanisms responsible for telomere maintenance in tumors have only recently been discovered. In particular, mutations of the telomere binding proteins alpha thalassemia/mental retardation syndrome X-linked (ATRX) or death-domain associated protein (DAXX) have been shown to underlie a telomere maintenance mechanism not involving telomerase (alternative lengthening of telomeres), and point mutations in the promoter of the telomerase reverse transcriptase (TERT) gene increase telomerase expression and have been shown to occur in melanomas. METHODS: To further define the tumor types in which TERT plays a role, we surveyed 1,230 tumors of 60 different types. We also analyzed the relationship between median overall survival (OS) of patients with IDH1/2 and TERT promoter mutations in a panel of 473 adult gliomas with the hypothesis that genetic signatures capable of distinguishing among several types of gliomas could be established providing clinically relevant information that can serve as an adjunct to histopathological diagnosis. RESULTS: We found that tumors could be divided into types with low and high frequencies of TERT promoter mutations. The nine TERT-high tumor types almost always originated in tissues with relatively low rates of self renewal, including melanomas, liposarcomas, hepatocellular carcinomas, urothelial carcinomas, squamous cell carcinomas of the tongue, medulloblastomas, and subtypes of gliomas. TERT and ATRX mutations were mutually exclusive, suggesting that these two genetic mechanisms confer equivalent selective growth advantages. We found that mutations in the TERT promoter occurred in 74.2% of glioblastomas (GBM), but occurred in a minority of Grade II-III astrocytomas (18.2%). In contrast, IDH1/2 mutations were observed in 78.4% of Grade II-III astrocytomas, but were uncommon in primary GBM. In oligodendrogliomas, TERT promoter and IDH1/2 mutations co-occurred in 79% of cases. Patients whose Grade III-IV gliomas exhibit TERT promoter mutations alone predominately have primary GBMs associated with poor median OS rates (11.5 months). Patients whose gliomas exhibit IDH1/2 mutations alone predominately have astrocytic morphologies and exhibit a median OS of 57 months while patients whose tumors exhibit both TERT promoter and IDH1/2 mutations predominately exhibit oligodendroglial morphologies and exhibit median OS of 125 months. CONCLUSIONS: In addition to their implications for understanding the relationship between telomeres and tumorigenesis, TERT mutations provide a biomarker that may aid in the classification and prognostication of brain tumors. SECONDARY CATEGORY: Tumor Biology.

// Patrick J. Killela 1,* , Christopher J. Pirozzi 1,* , Patrick Healy 2 , Zachary J. Reitman 1 , Eric Lipp 1 , B. Ahmed Rasheed 1 , Rui Yang 1 , Bill H. Diplas 1 , Zhaohui Wang 1 , Paula K. Greer 1 , Huishan Zhu 1 , Catherine Y. Wang 1 , Austin B. Carpenter 1 , Henry Friedman 1 , Allan H. Friedman 1 , Stephen T. Keir 1 , Jie He 3 , Yiping He 1 , Roger E. McLendon 1 , James E. Herndon II 2 , Hai Yan 1 and Darell D. Bigner 1 1 Department of Pathology, Duke University Medical Center, The Preston Robert Tisch Brain Tumor Center at Duke, and Pediatric Brain Tumor Foundation Institute at Duke, Durham, NC, USA 2 Department of Biostatistics and Bioinformatics, Duke University Medical Center, Durham, NC, USA 3 Department of Thoracic Surgery, Cancer Institute and Hospital Chinese Academy of Medical Sciences, Beijing, 100021, China * Denotes equal contribution Correspondence: Hai Yan, email: // Keywords : TERT promoter, IDH1, IDH2, Glioma Received : January 26, 2014 Accepted : January 28, 2014 Published : January 28, 2014 Abstract Frequent mutations in isocitrate dehydrogenase 1 and 2 ( IDH1 and IDH2 ) and the promoter of telomerase reverse transcriptase ( TERT ) represent two significant discoveries in glioma genomics. Understanding the degree to which these two mutations co-occur or occur exclusively of one another in glioma subtypes presents a unique opportunity to guide glioma classification and prognosis. We analyzed the relationship between overall survival (OS) and the presence of IDH1/2 and TERT promoter mutations in a panel of 473 adult gliomas. We hypothesized and show that genetic signatures capable of distinguishing among several types of gliomas could be established providing clinically relevant information that can serve as an adjunct to histopathological diagnosis. We found that mutations in the TERT promoter occurred in 74.2% of glioblastomas (GBM), but occurred in a minority of Grade II-III astrocytomas (18.2%). In contrast, IDH1/2 mutations were observed in 78.4% of Grade II-III astrocytomas, but were uncommon in primary GBM. In oligodendrogliomas, TERT promoter and IDH1/2 mutations co-occurred in 79% of cases. Patients whose Grade III-IV gliomas exhibit TERT promoter mutations alone predominately have primary GBMs associated with poor median OS (11.5 months). Patients whose Grade III-IV gliomas exhibit IDH1/2 mutations alone predominately have astrocytic morphologies and exhibit a median OS of 57 months while patients whose tumors exhibit both TERT promoter and IDH1/2 mutations predominately exhibit oligodendroglial morphologies and exhibit median OS of 125 months. Analyzing gliomas based on their genetic signatures allows for the stratification of these patients into distinct cohorts, with unique prognosis and survival.

Abstract The majority of glioblastomas can be classified into molecular subgroups based on mutations in the TERT promoter ( TERTp ) and isocitrate dehydrogenase 1 or 2 ( IDH ). These molecular subgroups utilize distinct genetic mechanisms of telomere maintenance, either TERTp mutation leading to telomerase activation or ATRX- mutation leading to an alternative lengthening of telomeres phenotype (ALT). However, about 20% of glioblastomas lack alterations in TERTp and IDH . These tumors, designated TERTp WT - IDH WT glioblastomas, do not have well-established genetic biomarkers or defined mechanisms of telomere maintenance. Here we report the genetic landscape of TERTp WT - IDH WT glioblastoma and identify SMARCAL1 inactivating mutations as a novel genetic mechanism of ALT. Furthermore, we identify a novel mechanism of telomerase activation in glioblastomas that occurs via chromosomal rearrangements upstream of TERT . Collectively, our findings define novel molecular subgroups of glioblastoma, including a telomerase-positive subgroup driven by TERT -structural rearrangements ( IDH WT - TERT SV ), and an ALT-positive subgroup ( IDH WT -ALT) with mutations in ATRX or SMARCAL1 .

// Changcun Guo 1,2 , Lee H. Chen 1,2 , Yafen Huang 1,2 , Chun-Chi Chang 1,2 , Ping Wang 1,2 , Christopher J. Pirozzi 1,2 , Xiaoxia Qin 4 , Xuhui Bao 1,2 , Paula K. Greer 1,2 , Roger E. McLendon 1,2 , Hai Yan 1,2 , Stephen T. Keir 1,3 , Darell D. Bigner 1,2 , Yiping He 1,2 1 The Preston Robert Tisch Brain Tumor Center at Duke and Pediatric Brain Tumor Foundation Institute, Duke University, Durham, NC 2 Department of Pathology, Duke University, Durham, NC 3 Department of Surgery, Duke University, Durham, NC 4 Institute for Genome Sciences and Policy, Duke University, Durham, NC Correspondence: Yiping He, email: // Keywords : MLL2, isogenic cell line, gene knockout, enhancer Received : October 30, 2013 Accepted : November 1, 2013 Published : November 3, 2013 Abstract KMT2D (lysine (K)-specific methyltransferase 2D), formerly named MLL2 (myeloid/lymphoid or mixed-lineage leukemia 2, also known as ALR/MLL4), is a histone methyltransferase that plays an important role in regulating gene transcription. In particular, it targets histone H3 lysine 4 (H3K4), whose methylations serve as a gene activation mark. Recently, KMT2D has emerged as one of the most frequently mutated genes in a variety of cancers and in other human diseases, including lymphoma, medulloblastoma, gastric cancer, and Kabuki syndrome. Mutations in KMT2D identified thus far point to its loss-of-function in pathogenesis and suggest its role as a tumor suppressor in various tissues. To determine the effect of a KMT2D deficiency on neoplastic cells, we used homologous recombination- and nuclease-mediated gene editing approaches to generate a panel of isogenic colorectal and medulloblastoma cancer cell lines that differ with respect to their endogenous KMT2D status. We found that a KMT2D deficiency resulted in attenuated cancer cell proliferation and defective cell migration. Analysis of histone H3 modifications revealed that KMT2D was essential for maintaining the level of global H3K4 monomethylation and that its enzymatic SET domain was directly responsible for this function. Furthermore, we found that a majority of KMT2D binding sites are located in regions of potential enhancer elements. Together, these findings revealed the role of KMT2D in regulating enhancer elements in human cells and shed light on the tumorigenic role of its deficiency. Our study supports that KMT2D has distinct roles in neoplastic cells, as opposed to normal cells, and that inhibiting KMT2D may be a viable strategy for cancer therapeutics.