Additional Tumor Suppressor Genes Research Articles

Combined Deficiency of Chromosome 7 Myeloid Tumor Suppressors Enhances Chemotherapy Resistance

Monosomy 7 and del(7q) [-7/del(7q)] are recurrent in myeloid neoplasms and associated with chemoresistance. -7/del(7q) is identified in up to half of therapy-related myeloid neoplasms (t-MN), high-risk secondary malignancies arising after prior exposure to chemotherapy or radiation. -7/del(7q) is also detected in clonal hematopoiesis, suggesting chromosome 7 aberrations can be early events in disease etiology. Despite this prevalence, the pathogenesis of -7/del(7q) in leukemogenesis remains unclear. We previously reported that deficiency of the transcription factor CUX1, a 7q-encoded tumor suppressor gene, promotes hematopoietic stem and progenitor cell drug resistance and t-MN transformation. Herein, we determined the combined impact of CUX1 loss with additional 7q tumor suppressor genes. To this end, we established a CRISPR/Cas9-based murine model of del(7q) clonal hematopoiesis and drug resistance. After targeting four 7q genes simultaneously, combined deficiency of CUX1 and the histone methyltransferase EZH2 uniquely promoted clonal outgrowth under genotoxic pressure in vivo and in vitro. Mechanistically, clonal selection is due, in part, to decreased apoptosis after chemotherapy exposure. RNA-seq in the absence of genotoxic insult revealed that Cux1 and Ezh2 loss has an additive transcriptional impact that is enriched for -7/del(7q) patient-derived gene signatures. Overall, we reveal a previously unknown genetic interaction between the 7q genes CUX1 and EZH2, supporting the concept of 7q as a contiguous gene syndrome region . A refined understanding of the molecular pathways driving del(7q) pathogenesis and drug resistance will enable development of therapies designed to counter or prevent these high-risk malignancies. In addition, we report a tractable approach for interrogating the pathogenesis of aneuploid events more broadly in cancer.

Fully accessible fitness landscape of oncogene-negative lung adenocarcinoma

Cancer genomes are almost invariably complex with genomic alterations cooperating during each step of carcinogenesis. In cancers that lack a single dominant oncogene mutation, cooperation between the inactivation of multiple tumor suppressor genes can drive tumor initiation and growth. Here, we shed light on how the sequential acquisition of genomic alterations generates oncogene-negative lung tumors. We couple tumor barcoding with combinatorial and multiplexed somatic genome editing to characterize the fitness landscapes of three tumor suppressor genes NF1, RASA1, and PTEN, the inactivation of which jointly drives oncogene-negative lung adenocarcinoma initiation and growth. The fitness landscape was surprisingly accessible, with each additional mutation leading to growth advantage. Furthermore, the fitness landscapes remained fully accessible across backgrounds with the inactivation of additional tumor suppressor genes. These results suggest that while predicting cancer evolution will be challenging, acquiring the multiple alterations that drive the growth of oncogene-negative tumors can be facilitated by the lack of constraints on mutational order.

Efficient mutation screening for cervical cancers from circulating tumor DNA in blood

BackgroundEarly diagnosis and continuous monitoring are necessary for an efficient management of cervical cancers (CC). Liquid biopsy, such as detecting circulating tumor DNA (ctDNA) from blood, is a simple, non-invasive method for testing and monitoring cancer markers. However, tumor-specific alterations in ctDNA have not been extensively investigated or compared to other circulating biomarkers in the diagnosis and monitoring of the CC. Therfore, Next-generation sequencing (NGS) analysis with blood samples can be a new approach for highly accurate diagnosis and monitoring of the CC.MethodUsing a bioinformatics approach, we designed a panel of 24 genes associated with CC to detect and characterize patterns of somatic single-nucleotide variations, indels, and copy number variations. Our NGS CC panel covers most of the genes in The Cancer Genome Atlas (TCGA) as well as additional cancer driver and tumor suppressor genes. We profiled the variants in ctDNA from 24 CC patients who were being treated with systemic chemotherapy and local radiotherapy at the Jeonbuk National University Hospital, Korea.ResultEighteen out of 24 genes in our NGS CC panel had mutations across the 24 CC patients, including somatic alterations of mutated genes (ZFHX3–83%, KMT2C-79%, KMT2D-79%, NSD1–67%, ATM-38% and RNF213–27%). We demonstrated that the RNF213 mutation could be used potentially used as a monitoring marker for response to chemo- and radiotherapy.ConclusionWe developed our NGS CC panel and demostrated that our NGS panel can be useful for the diagnosis and monitoring of the CC, since the panel detected the common somatic variations in CC patients and we observed how these genetic variations change according to the treatment pattern of the patient.

KRAS-driven miR-29b expression is required for tumor suppressor gene silencing.

KRAS activation drives DNA methylation and silencing of specific tumor suppressor genes (TSGs). We previously showed that the ERK pathway induces transcriptional repression of TET1, which results in conversion of TSG promoters from a hydroxymethylated, active state to a hypermethylated and silenced state. Here we identified miR-29b as a KRAS-induced molecule that represses TET1 expression. In KRAS-transformed cells, ectopic miR-29b inhibition restores expression of TET1, thereby reactivating TSGs by reducing methylation and restoring hydroxymethylation. Mining gene expression data of lung cancer cell lines identified additional TSGs suppressed by KRAS signaling whose expression was restored by inhibition of miR-29b and re-expression of TET1. Because KRAS changes TSG promoters from hydroxymethylated to hypermethylated with miR-29b-dependent silencing of TET1, we demonstrate a model in which DNMT1 is present on target promoters prior to KRAS transformation. In addition, we propose miR-29b as a potential circulating biomarker and target for rational treatment of specific malignancies.

Effects of long-term ethanol consumption and Aldh1b1 depletion on intestinal tumourigenesis in mice.

Ethanol and its metabolite acetaldehyde have been classified as carcinogens for the upper aerodigestive tract, liver, breast, and colorectum. Whereas mechanisms related to oxidative stress and Cyp2e1 induction seem to prevail in the liver, and acetaldehyde has been proposed to play a crucial role in the upper aerodigestive tract, pathological mechanisms in the colorectum have not yet been clarified. Moreover, all evidence for a pro-carcinogenic role of ethanol in colorectal cancer is derived from correlations observed in epidemiological studies or from rodent studies with additional carcinogen application or tumour suppressor gene inactivation. In the current study, wild-type mice and mice with depletion of aldehyde dehydrogenase 1b1 (Aldh1b1), an enzyme which has been proposed to play an important role in acetaldehyde detoxification in the intestines, received ethanol in drinking water for 1 year. Long-term ethanol consumption led to intestinal tumour development in wild-type and Aldh1b1-depleted mice, but no intestinal tumours were observed in water-treated controls. Moreover, a significant increase in DNA damage was detected in the large intestinal epithelium of ethanol-treated mice of both genotypes compared with the respective water-treated groups, along with increased proliferation of the small and large intestinal epithelium. Aldh1b1 depletion led to increased plasma acetaldehyde levels in ethanol-treated mice, to a significant aggravation of ethanol-induced intestinal hyperproliferation, and to more advanced features of intestinal tumours, but it did not affect intestinal tumour incidence. These data indicate that ethanol consumption can initiate intestinal tumourigenesis without any additional carcinogen treatment or tumour suppressor gene inactivation, and we provide evidence for a role of Aldh1b1 in protection of the intestines from ethanol-induced damage, as well as for both carcinogenic and tumour-promoting functions of acetaldehyde, including increased progression of ethanol-induced tumours. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Abstract S2-02: The landscape of somatic genetic alterations in BRCA1 and BRCA2 breast cancers

Abstract Introduction: Women carrying BRCA1 or BRCA2 germline mutations have a 45-80% lifetime risk of developing breast cancer (BC). BRCA1 and BRCA2 are perceived as bona fide tumor suppressor genes, whereby bi-allelic inactivation in tumor cells is required for tumorigenesis. Recent studies have indicated that loss of heterozygosity (LOH) of the wild-type allele of BRCA1 may be heterogeneous and constitute a late event. Therefore, additional somatic events prior to full BRCA1/2 inactivation may be required for tumorigenesis. Given that the somatic events that result in the development of BRCA1/2-BCs and their chronology are not understood, here we sought to define the genomic landscape of BRCA1/2-BCs and whether LOH of BRCA1/2 wild-type allele and/or mutations affecting additional tumor suppressor genes would be clonal or subclonal in these cancers. Methods: We retrieved 29 BRCA1-BCs and 10 BRCA2-BCs from the Pathology Departments of the authors' institutions. DNA extracted from microdissected tumor and normal breast samples was subjected to targeted capture massively parallel sequencing using either the MSK-IMPACT assay or an assay targeting all exons of 254 genes recurrently mutated in BC or related to DNA repair. Somatic single nucleotide variants, small insertions and deletions and copy number alterations affecting genes present in both sequencing assays (111 genes) were defined using state-of-the-art bioinformatics algorithms. ABSOLUTE and FACETS were employed to define clonal (i.e. present virtually in 100% of the cancer cells of a given case) and subclonal mutations and the presence of LOH of the BRCA1 and BRCA2 wild-type alleles. Results: Our analysis revealed bi-allelic inactivation of BRCA1 in 28 of 29 BRCA1-BCs (93% harbored LOH of the BRCA1 wild-type allele and 3% harbored a second somatic BRCA1 pathogenic mutation). The only BRCA1-BC lacking bi-allelic inactivation of BRCA1 was an estrogen receptor-positive lobular carcinoma, lacking genomic features consistent with homologous recombination DNA repair defects, diagnosed at 62 years of age. Bi-allelic inactivation of BRCA2 was found in all cases (100% of harbored LOH of the BRCA2 wild-type allele). A clonal somatic 'second hit' resulting in bi-allelic inactivation of BRCA1 or BRCA2 was detected in 76% and 100% of BRCA1-BCs and BRCA2-BCs, respectively. In BRCA1-BCs, TP53 mutations were detected in 76% of cases, and these mutations were found to be clonal in 58% of cases. The repertoire of somatic mutations affecting BRCA1-BCs included clonal somatic mutations or homozygous deletions of known tumor suppressor genes, such as PTEN, RB1, CDKN2A and NF1. In contrast, only 10% of the BRCA2-BCs harbored TP53 somatic mutations. Though clonal somatic mutations in several cancer genes were detected, 40% of BRCA2-BCs had no mutations affecting the cancer genes analyzed. Conclusions: Bi-allelic inactivation of BRCA1 and BRCA2 are frequent events in BRCA1-BCs and BRCA2-BCs, respectively. In a subset of BRCA1-BCs, however, the second 'hit' appeared to be subclonal, whereas mutations affecting TP53 and other tumor suppressor genes were clonal, supporting the notion that at least in a subset of these tumors, loss of the wild-type allele of BRCA1 may be preceded by inactivation of another tumor suppressor gene. Citation Format: Geyer FC, Burke KA, Macedo GS, Piscuoglio S, Ng CK, Martelotto LG, Papanastatiou AD, De Filippo MR, Schultheis AM, Brogi E, Robson M, Wen YH, Weigelt B, Schnitt SJ, Tung N, Reis-Filho JS. The landscape of somatic genetic alterations in BRCA1 and BRCA2 breast cancers [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr S2-02.

Early B-cell-specific inactivation of ATM synergizes with ectopic CyclinD1 expression to promote pre-germinal center B-cell lymphomas in mice.

Ataxia telangiectasia-mutated (ATM) kinase is a master regulator of the DNA damage response. ATM is frequently inactivated in human B-cell non-Hodgkin lymphomas, including ~50% of mantle cell lymphomas (MCLs) characterized by ectopic expression of CyclinD1. Here we report that early and robust deletion of ATM in precursor/progenitor B cells causes cell autonomous, clonal mature B-cell lymphomas of both pre- and post-germinal center (GC) origins. Unexpectedly, naive B-cell-specific deletion of ATM is not sufficient to induce lymphomas in mice, highlighting the important tumor suppressor function of ATM in immature B cells. Although EμCyclinD1 is not sufficient to induce lymphomas, EμCyclinD1 accelerates the kinetics and increases the incidence of clonal lymphomas in ATM-deficient B-cells and skews the lymphomas toward pre-GC-derived small lymphocytic neoplasms, sharing morphological features of human MCL. This is in part due to CyclinD1-driven expansion of ATM-deficient naive B cells with genomic instability, which promotes the deletions of additional tumor suppressor genes (i.e. Trp53, Mll2, Rb1 and Cdkn2a). Together these findings define a synergistic function of ATM and CyclinD1 in pre-GC B-cell proliferation and lymphomagenesis and provide a prototypic animal model to study the pathogenesis of human MCL.

Oncotargets in different renal cancer subtypes.

Renal cell cancer is a heterogeneous group of cancers with different histologic subtypes. The majority of renal tumors in adults are clear cell renal cell carcinomas, which are characterized by von Hippel- Lindau (VHL) gene alterations. Recent advances in defining the genetic landscape of renal cancer has shown the genetic heterogeneity of clear cell renal cell carcinomas (ccRCC) and the presence of at least 3 additional ccRCC tumor suppressor genes on chromosome 3p. Due to inactivation of VHL, renal cancer cells produce the HIF-responsive growth factor VEGF. The PI3K--mTORC1 signaling axis also represents a target for therapy. The new systemic therapies, including tyrosine kinase inhibitors, monoclonal antibodies, and mTOR inhibitors, aim to suppress angiogenesis with vascular endothelial growth factor as a target. Various VEGF-inhibitors are approved for the treatment of ccRCC and we discuss recent advancements in the treatment of metastatic ccRCC. Other gene alterations have been identified in hereditary cancer syndromes, e.g. FLCN, TSC1, TSC2, TFE3, TFEB, MITF, FH, SDHB, SDHD, MET, and PTEN and we review their role in renal tumor carcinogenesis, prognosis, and targeted therapy. By reviewing the associations between morphologic features and molecular genetics of renal cancer we provide insight into the basis for targeted renal cancer therapy.

Molecular genetics of clear-cell renal cell carcinoma.

Renal cell carcinoma of clear-cell type (ccRCC) is an enigmatic tumor type, characterized by frequent inactivation of the VHL gene (infrequently mutated in other tumor types), responsiveness to angiogenesis inhibitors, and resistance to both chemotherapy and conventional radiation therapy. ccRCC tumors exhibit substantial mutation heterogeneity. Recent studies using massively parallel sequencing technologies have implicated several novel driver genes. In VHL wild-type tumors, mutations were discovered in TCEB1, which encodes Elongin C, a protein that binds to VHL and is required for its function. Several additional tumor suppressor genes have been identified near the VHL gene, within a region that is frequently deleted in ccRCC on chromosome 3p: SETD2, BAP1, and PBRM1. Mutations in BAP1 and PBRM1 are largely mutually exclusive and are associated with different tumor biology and patient outcomes. In addition, the mTORC1 pathway is deregulated by mutations in MTOR, TSC1, PIK3CA, and PTEN in approximately 20% of ccRCCs. Mutations in TSC1, and possibly other genes, may predict for sensitivity to mTORC1 inhibitors. These discoveries provide insight into ccRCC development and set the foundation for the first molecular genetic classification of the disease, paving the way for subtype-specific therapies.

Tumor suppressive mechanisms in hepatocellular carcinoma through inhibition of IL-6 signaling

Hepatocellular carcinoma (HCC) is the most frequent malignant tumor in the liver and the third leading cause of cancer death worldwide (696,000 deaths, 9.2%). Although significant progress has been made in the HCC field, the molecular mechanisms and signaling pathways underlying HCC development and progression are still poorly understood. This is probably the case because HCC is very heterogeneous in terms of clinical presentation, genomic alterations and gene expression patterns. In a recent study, we applied an integrative approach of high-resolution array-based comparative genomic hybridization (arrayCGH) and gene expression profiling coupled with patient prognosis to identify tumor 'driver' genes which have genomic aberrations in HCC clinical specimens and functionally contribute to tumor progression. This analysis led to the identification of 10 potential cancer driver genes. Six of these genes were located on chromosome 8 p which is associated with poor outcome. Importantly, the six genes on chromosome 8 p included the tumor suppressor gene DLC1 that had been shown previously to be involved in HCC development. Next we performed clonogenicity, cell migration assays and xenograft mouse models to analyze the five additional genes' for tumor suppressive properties. These experiments confirmed the tumor suppressor function of two additional tumor suppressor genes on chromosome 8 p, SH2D4A and SORBS3. Furthermore, functional analysis revealed that SH2D4A and SORBS3 function in a convergent manner to inhibit the IL-6 pathway. Thereby, SORBS3 inhibits the IL-6 pathway via activation of the estrogen receptor alpha and SH2D4A via inhibition of STAT3 signaling. Taken together, in this study we identified two novel tumor suppressor genes on chromosome 8 p which functionally cooperate to inhibit IL-6 signaling.

Prospective study of tumor suppressor gene (TSG) methylation as a prognostic biomarker in surgically resected non-small cell lung cancer (NSCLC).

7066 Background: In the New England Journal of Medicine, Brock et al. (2008) published a nested case-control study which tested the association between early recurrence of NSCLC after surgical resection, and TSG promoter methylation in tumor and lymph nodes detected by methylation-specific PCR (MSP). They reported that promoter methylation of the TSGs p16, CDH13, RASSF1A, and APC was significantly associated with early recurrence in 51 patients with stage I NSCLC compared to matched patients without early recurrence. We attempted to confirm these findings. Methods: In a prospective study, fresh frozen tumor tissue was acquired after surgical resection in 107 patients with stage I-IIIA NSCLC between 2003-2008. The promoter methylation status of the same 4 genes examined by Brock, et al (p16, CDH13, RASSF1A, APC), as well as 6 additional TSGs (MGMT, WIF-1, METH-2, GSTP1, SOCS3, DAPK) were assessed in the tumor tissue using quantitative MSP (MethyLight assay), with any amount of methylation scored as positive. Methylation status was correlated with clinical features, pathologic stage, disease-free survival (DFS), and overall survival (OS). Results: No significant associations were observed between promoter methylation of individual TSGs and DFS. No significant associations were observed between the number of methylated TSGs and DFS, or OS. Increased RASSF1A methylation was observed in poorly-differentiated and undifferentiated tumors compared to tumors that were well- or moderately-differentiated (p=0.031). Increased WIF-1 methylation and GSTP1 methylation were associated with increasing T (p=0.01) and N stage (p=0.028), respectively. Squamous cell carcinomas (SQCCs) were characterized by increased p16 methylation (p=0.0314) and decreased APC methylation (p=0.0146) compared to tumors of non-SQCC histologies. Conclusions: In this prospective study, we did not confirm that the promoter methylation of p16, CDH13, RASSF1A, and APC, or 6 other TSGs, was prognostic for early recurrence in surgically resected NSCLC.

Abstract 230: Modeling tumor progression and identifying genotype-drug interactions by the sequential introduction of cancer mutations into the genome of human normal cells

Abstract The mutational landscape of cancer genomes displays a complex combination of genetic lesions affecting oncogenes and tumor suppressor genes. The construction of cellular models closely recapitulating cancer-linked genetic alterations is a prerequisite to understand their role in tumor progression and to identify genotype- specific pharmacological responses. To recapitulate point mutations affecting oncogenes we used homologous recombination to ‘knock-in’ specific nucleotide changes in the genome of human cells. We focused on the following alleles: EGFR (delE746-A750), KRAS (G13D), BRAF (V600E) and PIK3CA (E545K and H1047R), that are found in multiple human cancers. As recipient cells, a human non transformed epithelial cell line of breast origin (hTERT-HME1) has been employed. This cell line has been immortalized by hTERT and harbors a mutation on TP53 gene, a genetic event frequently observed in naturally occurring human tumors. To recapitulate the inactivation of additional tumor suppressor genes in our cellular models, we exploited shRNAs to permanently knock-down the expression of PTEN and RB1 genes, as they are frequently down regulated in human tumors. Knock-down of PTEN and RB1 was performed both in the parental hTERT-HME1 cells and in the ‘knock-in’ models, thus allowing us to combine tumor suppressor genes inactivation with mutational activation of specific oncogenes. The resulting cellular models are hereafter refereed to as combinatorial ‘matrix’. To assess how specific combinations of genetic lesions affect the tumorigenic properties of not transformed cells, we measured the ability of the ‘matrix’ to grow in soft agar. At the same time we injected our cellular models in the cleared, humanized mammary fat pad of immunocompromised mice. For the latter approach, we focused on genotypes recapitulating mutations found in breast tumors, such as PTEN or RB1 inactivation and PIK3CA or KRAS mutations, alone or in combination. In vitro and in vivo results showed that those combinations of genetic events are not sufficient to transform the normal hTERT-HME1 cells. However, preliminary data indicates that few genotypes of the ‘matrix’ acquired the ability to colonize the murine fat pads. With the rational of unveiling new pharmaco-genetic interactions involving one or multiple cancer mutations, the “matrix” was used to screen a panel of compounds used for cancer therapy, or for which an anti-proliferative activity was previously reported. This approach led to the identification of a subset of genotype-drug interactions thus underscoring how the multifactorial genomic milieu of cancer cells impact the response to therapies. Grant support: This work is part of the EU FP7 grant COLTHERES Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 230. doi:10.1158/1538-7445.AM2011-230

Histone methyltransferase gene SETD2, a novel tumor suppressor gene in clear cell renal cell carcinoma

Sporadic clear cell renal cell carcinoma (cRCC) is genetically characterized by the recurrent loss of 3p regions. Inactivation of the Von Hippel Lindau (VHL) gene, located on 3p26-p25, has been shown to play an important role in the development of cRCC. Inactivation of VHL results in accumulation of transcription factor HIF1-a, resulting in the activation of several pathways that promote tumour development. The recurrent loss of 3p21 suggests the presence of an additional important tumor suppressor gene in this region.

Abstract CN02-02: Epigenetic targets in endometrial cancer

Abstract CN02-02: Epigenetic targets in endometrial cancer

Phenotype‐rescue of cyclin‐dependent kinase inhibitor p16/INK4A defects in a spontaneous canine cell model of breast cancer

Mammary cancer is among the most frequently observed canine tumors in unspayed female dogs resulting in death due to metastatic disease. These tumors are excellent models of human breast cancer but until recently there was only anecdotal evidence regarding underlying genetic defects. We recently reported expression defects in the cyclin-dependent kinase p21/Cip1 and p53 among three independent canine mammary tumor (CMT) cell lines derived from spontaneous canine mammary cancers. We investigated further defects in the same three cell lines focusing on additional tumor suppressor gene defects in cyclin-dependent kinase inhibitors. p27/KIP1 appeared normally expressed and did not appear to encode inactivating mutations. In contrast, expression of p16/INK4A was defective/absent in two cell lines and normal/slightly induced in the third cell line. To determine if defects were causative in maintaining the transformed phenotype, a p16/INK4A transgene was permanently transfected followed by selection and single cell cloning. CMT/p16 clones were characterized for transgene expression, p16 protein content and phenotype including proliferation rate, cell cycle phase distribution, contact inhibition, substrate dependent cell growth and cell morphology. All cell lines appeared unique yet clear indications of phenotype rescue due to p16/INK4A transgene complementation were observed suggesting that defects in p16 expression were present in all three. In some cases cellular senescence also appeared to be induced. These data provide evidence supporting p16/INK4A mutations as causative defects promoting transformation in canine mammary cancer and further characterizes tumor suppressor gene defects with functional consequences in these cells supporting their application as spontaneous animal models of human disease.

Mutational inactivation of PTPRD in glioblastoma multiforme and malignant melanoma.

An additional tumor suppressor gene on chromosome 9p telomeric to the CDKN2A/B locus has long been postulated to exist. Using Affymetrix 250K single nucleotide polymorphism arrays to screen for copy number changes in glioblastoma multiforme (GBM), we detected a high frequency of deletions of the PTPRD gene, which encodes a receptor protein tyrosine phosphatase at chromosome 9p23-24.1. Missense and nonsense mutations of PTPRD were identified in a subset of the samples lacking deletions, including an inherited mutation with somatic loss of the wild-type allele. We then sequenced the gene in melanoma and identified 10 somatic mutations in 7 of 57 tumors (12%). Reconstitution of PTPRD expression in GBM and melanoma cells harboring deletions or mutations led to growth suppression and apoptosis that was alleviated by both the somatic and constitutional mutations. These data implicate PTPRD in the pathogenesis of tumors of neuroectodermal origin and, when taken together with other recent reports of PTPRD mutations in adenocarcinoma of the colon and lung, suggest that PTPRD may be one of a select group of tumor suppressor genes that are inactivated in a wide range of common human tumor types.

S-adenosylhomocysteine hydrolase downregulation contributes to tumorigenesis

With the idea to discover novel genes involved in proliferation, we have performed a genome-wide loss-of-function genetic screen to identify additional putative tumor suppressor genes. We have previously identified five genes belonging to different biochemical families. In this report, we focused on the study of one of these genes designated S-adenosylhomocysteine hydrolase (SAHH), which has also been previously identified in an independent short hairpin RNA screening. SAHH inactivation confers resistance to both p53 and p16(INK4)-induced proliferation arrest. Interestingly, SAHH inactivation inhibits p53 transcriptional activity and impairs DNA damage-induced transcription of p21(Cip1). Given that SAHH downregulation modulates senescence in primary cells, we also studied SAHH expression in human tumors at the messenger RNA (mRNA) and protein levels. SAHH mRNA was lost in 50% of tumor tissues from 206 patients with different kinds of tumors in comparison with normal tissue counterparts. Moreover, SAHH protein was also affected in some colon cancers. Such findings may be of relevance to cancer research, suggesting that SAHH might be a largely unexplored tumor suppressor.

BCR expression is decreased in meningiomas showing loss of heterozygosity of 22q within a new minimal deletion region

Neurofibromin 2 ( NF2), located on chromosome arm 22q, has been established as a tumor suppressor gene involved in meningioma pathogenesis. In our study, we investigated 149 meningiomas to determine whether there are additional tumor suppressor genes localized on chromosome 22q, apart from NF2, that might be involved in meningioma pathogenesis. The LOH analysis on chromosome 22q identified two regions of deletion: the first one, which is limited to the NF2 gene locus, and the second one, which is outside this location. The new minimal deletion region (MDR) included the following genes: BCR (breakpoint cluster region), RAB36 (a member of RAS oncogene family), GNAZ [guanine nucleotide binding protein (G protein), α-z polypeptide], and RTDR1 (rhabdoid tumor deletion region gene 1). The expression levels of all these genes, including NF2, were subsequently analyzed by quantitative real-time polymerase chain reaction. We observed a significantly lowered expression level of NF2 in meningiomas with 22q loss of heterozygosity (LOH) within NF2 region compared to the one in meningiomas with 22q retention of heterozygosity (ROH, P<0.05). Similarly, BCR showed a significantly lowered expression in meningiomas with 22q LOH within the new MDR compared to cases with 22q ROH ( P<0.05). Our data, together with the already published information considering BCR function suggest that BCR can be considered as a candidate tumor suppressor gene localized on chromosome 22q which may be involved in meningioma pathogenesis.

Analysis of Locus‐Specific DNA Methylation in Response to Chronic Folic Acid Supplementation and Withdrawal in Chinese Women

The objective of this study was to evaluate the effect of chronic folic acid supplementation and withdrawal on DNA methylation at specific genetic loci in a Chinese population. Women of reproductive age were assigned a daily folic acid supplement of 400 μg for 6 months, and whole blood samples were drawn at baseline, after 6 months of daily folic acid intake, and 3 months after folic acid supplementation was withdrawn. Blood DNA from 10 subjects (5 MTHFR CC and 5 TT) was analyzed by methylation‐specific PCR (MSP) for changes in methylation status at the promoters of 6 tumor suppressor genes: DAPK, ESR1, HIC1, MGMT, p16, and TIMP, genes known to be aberrantly silenced by hypermethylation of their promoter regions in a variety of human cancers. Overall, no widespread changes in normal DNA methylation patterns (i.e., hypermethylation) were detected for any of the genes tested in all 10 subjects in response to 6 months of folic acid supplementation and 3 months of folic acid withdrawal. To further assess potential changes in DNA methylation in response to folic acid fortification, additional tumor suppressor genes are being analyzed by MSP, and high resolution DNA methylation analyses by sodium bisulfite genomic sequencing are currently in progress. Supported by a collaborative CDC agreement.

Detailed mapping of chromosome 17p deletions reveals HIC1 as a novel tumor suppressor gene candidate telomeric to TP53 in diffuse large B-cell lymphoma

Deletions in the short arm of chromosome 17 (17p) involving the tumor suppressor TP53 occur in up to 20% of diffuse large B-cell lymphomas (DLBCLs). Although inactivation of both alleles of a tumor suppressor gene is usually required for tumor development, the overlap between TP53 deletions and mutations is poorly understood in DLBCLs, suggesting the possible existence of additional tumor suppressor genes in 17p. Using a bacterial artificial chromosome (BAC) and Phage 1 artificial chromosome (PAC) contig, we here define a minimally deleted region in DLBCLs encompassing approximately 0.8 MB telomeric to the TP53 locus. This genomic region harbors the tumor suppressor Hypermethylated in Cancer 1 (HIC1). Methylation-specific PCR demonstrated hypermethylation of HIC1 exon 1a in a substantial subset of DLBCLs, which is accompanied by simultaneous HIC1 deletion of the second allele in 90% of cases. In contrast, HIC1 inactivation by hypermethylation was rarely encountered in DLBCLs without concomitant loss of the second allele. DLBCL patients with complete inactivation of both HIC1 and TP53 may be characterized by an even inferior clinical course than patients with inactivation of TP53 alone, suggesting a functional cooperation between these two proteins. These findings strongly imply HIC1 as a novel tumor suppressor in a subset of DLBCLs.