Heterozygosity In Human Cancers Research Articles

Epigenetic regulation of the Wnt signaling inhibitor DACT2 in human hepatocellular carcinoma

DACT2 (Dapper, Dishevelled-associated antagonist of β-catenin homolog 2) is a member of the DACT family involved in the regulation of embryonic development. Human DACT2 is localized on 6q27, a region of frequent loss of heterozygosity in human cancers. However, the regulation of DACT2 expression and function in hepatocellular carcinoma (HCC) remains unclear. In this study, genetic and epigenetic changes of DACT2 were analyzed in HCC cell lines and primary cancer. We found no single-nucleotide polymorphism (SNP) associated with HCC. Promoter region methylation was correlated with loss or reduction of DACT2 expression, and restoration of DACT2 expression was induced by 5-aza-2’-deoxycytidine (5-AZA) in HCC cell lines. Promoter region methylation was found in 54.84% of primary HCC. Reduction of DACT2 expression was associated with promoter hypermethylation, and expression of DACT2 was inversely related to β-catenin expression in primary HCC. DACT2 suppressed cell proliferation, induced G2-M arrest in cell lines and inhibited tumor growth in xenograft nude mice. The transcriptional activity of TCF-4 and the expression of Wnt signaling downstream genes were suppressed by DACT2 re-expression and reactivated by depletion of DACT2. In conclusion, DACT2 is frequently methylated in HCC and its expression is regulated by promoter hypermethylation. DACT2 suppresses HCC by inhibiting Wnt signaling in human HCC.

Low Expression of the Novel Tumor Suppressor Trim62 Is An Independent Adverse Prognostic Factor for Survival In Acute Myeloid

Abstract 563▪FN2▪This icon denotes a clinically relevant abstract Background:The role of TRIM (tripartite motif) proteins in human cancer is unknown except for TRIM19 (PML, promyelocytic leukemia gene), which acquires oncogenic activity when fused to RARα (retinoic acid receptor alpha) and causes acute promyelocytic leukemia. We have confirmed a putative role of TRIM62 as a tumor suppressor by genetically deleting the TRIM62 locus in a mouse model. TRIM62 maps to chromosome 1p35.1, a genomic region frequently associated with loss of heterozygosity in human cancer, including leukemia. Methodology:We assessed TRIM62 expression by high-throughput reverse phase protein array (RPPA) technology in 511 pts. Eleven CD34+ bone marrow (BM) and 10 normal peripheral blood (PB) lymphocyte samples were used as controls. Samples were printed as 5 serial 1:2 dilutions in duplicate using an Aushon 2470 Arrayer. Slides were probed with 195 antibodies against apoptosis, cell cycle, signaling, integrins, and phosphatases among other functional protein groups. Patients were segregated according to TRIM62 levels into TRIM62LOW (lower two tertiles) and TRIM62HIGH (highest tertile). Results:The level of TRIM62 protein in AML samples was significantly below (p=0.0048) that of CD34+ cells obtained from healthy volunteers., The level of expression was similar in the paired PB and BM derived AML samples (p=0.2433). Cases of AML-M4 and –M5 exhibited lower expression of TRIM62 than other FAB subtypes.,Stratification according to specific cytogenetic abnormalities (p=0.9286), standard cytogenetic risk criteria (favorable, intermediate, poor; p=0.6824), or mutational status (FLT3, NPM1, RAS, p53, or IDH1/2) failed to show statistically significant differences in TRIM62 expression.Overall survival (OS) was significantly different in TRIM62LOW and TRIM62HIGH for all patients (40.4 vs. 75.6 weeks, p=0.00002), and this difference was greater among patients with cytogenetically normal AML (CN-AML; 47 vs. 125.3 weeks, p=0.00001; Figure 1‘A). TRIM62HIGH patients had significantly longer OS compared with TRIM62LOW patients, regardless of whether they had received standard cytarabine-based chemotherapy (p=0.00003) or epigenetic therapy (i.e. hypomethylating agents with or without histone deacetylase inhibitors; p=0.02). TRIM62LOW was associated with a trend toward shorter remission duration compared with TRIM62HIGH for all cases (38 vs. 63 weeks, p=0.06) and in CN-AML patients (47.7 vs. 58.1 weeks, p=0.09). By multivariate analysis, TRIM62HIGH was identified as an independent favorable risk factor for survival both in the whole cohort (p=0.0085) and in patients with CN-AML (p=0.0014). Notably, TRIM62LOW discriminated a subset of patients with a distinct poor prognosis among those with FLT3MUT (36 vs. 128.5 weeks, p=0.006), FLT3WT (66 vs. 129 weeks, p=0.001), NPM1MUT (34.6 vs. 253.1 weeks, p=0.0026), NPM1WT (51.7 vs. 102.6 weeks, p=0.017), and RASWT (46.3 vs. 110, p=0.0002). Among patients with the most favorable prognosis (FLT3WTNPM1MUT CN-AML), TRIM62HIGH identified a subset of patients for whom stem cell transplant is not recommended: OS for FLT3WTNPM1MUTTRIM62HIGH vs. FLT3WTNPM1MUTTRIM62LOW patients was 100% vs. 35%, after 6 years of follow-up (Figure 1B). When patients with FLT3WTNPM1mutant were excluded from the analysis, TRIM62 still stratified two distinct groups of patients with AML regarding OS, both for the whole cohort (p=0.03) and for those with CN-AML (p=0.0006). By assessing the level of 195 other proteins on the same RPPA chip, we showed that TRIM62 levels correlated significantly and positively (R>0.2) with proteins involved in leukemia stem cell pathways (β-catenin and Notch), migration/homing (fibronectin, integrin β3), hypoxia (VHL, HIF1α), and the p53 pathway (MDM2, p21). [Display omitted] Conclusions:TRIM62 loss, consistent with a role as a tumor suppressor, represents a powerful independent adverse prognostic factor in AML. TRIM62HIGH identifies a population of CN-AML patients with superior outcomes compared with TRIM62LOW amongst those with CN-AML even after stratification by NPM1 and FLT3 mutational status. Disclosures:No relevant conflicts of interest to declare.

A kinase shRNA screen links LATS2 and the pRB tumor suppressor

pRB-mediated inhibition of cell proliferation is a complex process that depends on the action of many proteins. However, little is known about the specific pathways that cooperate with the Retinoblastoma protein (pRB) and the variables that influence pRB's ability to arrest tumor cells. Here we describe two shRNA screens that identify kinases that are important for pRB to suppress cell proliferation and pRB-mediated induction of senescence markers. The results reveal an unexpected effect of LATS2, a component of the Hippo pathway, on pRB-induced phenotypes. Partial knockdown of LATS2 strongly suppresses some pRB-induced senescence markers. Further analysis shows that LATS2 cooperates with pRB to promote the silencing of E2F target genes, and that reduced levels of LATS2 lead to defects in the assembly of DREAM (DP, RB [retinoblastoma], E2F, and MuvB) repressor complexes at E2F-regulated promoters. Kinase assays show that LATS2 can phosphorylate DYRK1A, and that it enhances the ability of DYRK1A to phosphorylate the DREAM subunit LIN52. Intriguingly, the LATS2 locus is physically linked with RB1 on 13q, and this region frequently displays loss of heterozygosity in human cancers. Our results reveal a functional connection between the pRB and Hippo tumor suppressor pathways, and suggest that low levels of LATS2 may undermine the ability of pRB to induce a permanent cell cycle arrest in tumor cells.

The interferon-α responsive gene TMEM7 suppresses cell proliferation and is downregulated in human hepatocellular carcinoma

Multiple regions on the chromosome arm 3p are frequently affected by loss of heterozygosity in human cancers. A candidate tumor suppressor gene is TMEM7, at 3p21.3, which encodes a transmembrane protein. TMEM7 is expressed specifically in the liver, and the encoded protein shares substantial sequence homology with human and mouse 28-kDa interferon-α (IFN-α) responsive protein. In investigation of the possible role of TMEM7 in development of hepatocellular carcinoma (HCC), we examined TMEM7 expression in 20 primary HCC and 18 HCC cell lines and found recurrent functional alterations. Although TMEM7 mRNA was expressed in normal hepatic cells, downregulation or inactivation of the gene was detected in 85% of primary HCC and 33% of HCC cell lines. To identify the mechanisms responsible, we examined genomic deletion and mutation, and also the effect of inhibitors of DNA methyltransferase and histone deacetylase on cells with low or no endogenous TMEM7 expression. Homozygous deletion of TMEM7 was not detected in 17 pairs of human HCC and corresponding noncancerous liver tissues, nor in any of the 18 HCC cell lines. TMEM7 mutation was not detected in the 18 HCC cell lines (low or normal TMEM7 expression). Treatment of two of six cell lines exhibiting downregulation or loss of TMEM7 with 5-aza-2′-deoxycytidine and trichostatin A yielded additive increase in TMEM7 expression, implicating aberrant DNA methylation and histone deacetylation in transcriptional silencing of this gene. Ectopic expression of TMEM7 in two TMEM7-deficient HCC lines suppressed cell proliferation, colony formation, and cell migration in vitro and reduced tumor formation in nude mice. Treatment of two highly invasive HCC cell lines with IFN-α for 7 days significantly increased TMEM7 expression and inhibited cell migration. These findings implicate loss of TMEM7 expression in hepatocarcinogenesis and suggest that modification of TMEM7 expression by IFN-α may have therapeutic relevance in a subset of HCC.

LOSS OF CONSTITUTIONAL HETEROZYGOSITY IN HUMAN CANCER

INTRODUCTION 282 CANCER AS A GENETIC DISEASE 282 Visible Chromosomal Rearrangements in Tumor Cells. . . . . . . . . . . . . . .. . . . . .. . . . . . . . . . . . . . . 282 Inherited Syndromes Predisposing to Cancer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 283 Behavior of the Transformed Trait in Cell Hybrids . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 284

Loss Of Constitutional Heterozygosity In Human Cancer

Loss Of Constitutional Heterozygosity In Human Cancer