bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2228: Folate-related genes polymorphism and risk of pediatric acute lymphoplastic leukemia
Authors: Raafat Nermin , Gharib Amal , Elsafy Usama , Hassan Tamer
Published: May 2015

Background: Childhood acute lymphoblastic leukemia (ALL) is the most common malignancy affecting children, constituting about 30% of all cancers among children. It constitutes about 75% of pediatric acute leukemia with peak incidence between ages 3 and 4. The assertion that ALL may have a genetic basis has long been pursued through association studies based on candidate genes as folate metabolism. Polymorphisms in folate pathway genes may influence the susceptibility to acute lymphoblastic leukemia. Increased risk of ALL was observed in reduced folate carrier protein (RFC1) 80GA variant carriers. A 1.4-fold reduction in ALL risk was observed for carriers of the methyl tetrahydrofolate reductase (MTHFR) 677T allele. Objectives: this study was carried out to evaluate the contribution of MTHFR C677T and RFC1 80GA gene polymorphism and susciptability of childhood acute lymphoblastic leukemia. Materials &Methods: DNA was isolated from 200 children, divided into 2 groups, group I included 100 pediatric ALL patients and group II included 100 healthy donors as controls. Genotyping of MTHFR C677T and RFC1 G80A was performed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) using Hinf I and HhaI restriction endonuclease respectively. Results: MTHFR 677T allele carriers were significantly lower in ALL cases (46%) than healthy controls (56%). Reduction in ALL risk was observed for heterozygous (CT) or homozygous (TT) carriers of the MTHFR 677T allele (OR 0.8; 95% CI, 0.9-1.5; P < 0.002). RFC1 80A allelic carriers have an increased susceptibility to ALL. Risk was increased 2.0 times (OR 3.5; 95% CI, 1.5-4.8 ; P .02) for A-allelic carriers. Conclusion: our data suggested that the MTHFR gene variants are associated with decreased ALL rate and risk. the reduced risk associated with the MTHFR C677T polymorphisms may be the result of changed intracellular folate redistribution. the results of our study also supported the suggestion that RFC1 G80A RFC1 G80A single nucleotide polymorphism contributed to increase the susceptibility and the development of pediatric ALL.

Citation Format: Nermin Raafat, Amal Gharib, Usama Elsafy, Tamer Hassan. Folate-related genes polymorphism and risk of pediatric acute lymphoplastic leukemia. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2228. doi:10.1158/1538-7445.AM2014-2228
Abstract 223: High-throughput drug discovery against breast cancer stem cells
Authors: McDermott Sean P. , Ndaw Fatou , Fox Alexandra , Roest Steve R. Vander , Larsen Martha J. , Wicha Max S.
Published: May 2015

The cancer stem cell (CSC) hypothesis postulates that tumors are maintained by a self-renewing CSC population that is also capable of differentiating into non-self renewing cell populations that constitute the bulk of the tumor. Clinically, CSCs are predicted to mediate tumor recurrence after chemotherapy and radiotherapy due to the relative inability of these modalities to effectively target CSCs. Therefore, new therapies that target CSC must be developed and combined with standard therapy to achieve a true cure. Targeting breast CSC represents an attractive avenue for developing therapeutics; however, there are multiple pools of breast CSCs utilizing distinct signaling pathways, such as Wnt/β-catenin & NF-κB. These breast CSC pools are identified using flow cytometry as cells with high expression of aldehyde dehydrogenase (ALDH), CD44+CD24-, or CD90+. We have developed a high-throughput flow cytometry based screen against CD90+ breast CSCs. We screened ∼18,500 small molecules for inhibitors of CD90+ frequency and identified 19 compounds. Here, we will present data examining this compounds against ALDH+, and CD44+CD24- breast CSCs. By identifying compounds with that target multiple pools of breast CSCs, these compounds might be exploited in future drug development endeavors.

Citation Format: Sean P. McDermott, Fatou Ndaw, Alexandra Fox, Steve R. Vander Roest, Martha J. Larsen, Max S. Wicha. High-throughput drug discovery against breast cancer stem cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 223. doi:10.1158/1538-7445.AM2014-223
Abstract 2233: Rearrangement of ERG and CHD1 genes in prostate cancer as a marker of tumor heterogeneity
Authors: Tereshchenko Irina V. , Zhong Hua , Chekmareva Marina , Santanam Urmila , Petrosky Whitney , Kane-Goldsmith Noriko , Tischfield Jay A. , DiPaola Robert
Published: May 2015

Efforts to develop targeted therapy based on genomic findings are complicated by the polygenic nature of drug resistance and genetic heterogeneity between separate biopsy cores. These findings have led to the conclusion that analysis of multiple biopsy cores and consideration of multiple targeted agents will be important. Because of our concern that these assessments may be missing an additional layer of heterogeneity within even a single biopsy core, which sequencing alone may not identify, we studied the genetic heterogeneity of cells in focal areas using TMA. Using confocal microscopy, we analyzed the most common and prostate cancer specific gene rearrangement (TMPRSS2-ERG fusion and CHD1 deletion) in neighboring single cells in a tumor focus without changing the stromal microenvironment. We evaluated 56 patients that underwent radical prostatectomy at CINJ with pathological assessment, grading, and ERG rearrangement status. Overall, ERG rearrangements occurred in 24/56 (42.9%) patients. The most frequent mechanism of rearrangements was deletion (58%), while a split event was confirmed in 25% of the ERG-rearranged cases. In 88% of cases the ERG rearrangement was associated with TMPRSS2-ERG gene fusion. A copy number gain of the non-rearranged ERG gene occurred in 10/56 (17.6%) cases. A heterogeneous mix of ERG deletions, translocations, and copy number changes occurred in 8/24 (33%) patients, with a combination of deletions and translocations alone occurring in 3/24 (12.5%) cases. 20/21 (95%) of analyzed cases with ERG-rearrangement were negative for CHD1 gene rearrangement (p< 0.01). Overall, 14/45 (31.1%) of patients had CHD1 deletion, and 13/14 (93%) CHD1- cases correlate with ERG-rearrangement negative status (p< 0.01). 10/45 (22.2%) cases represent CHD1 gene copy number gain. These results support the notion that additional heterogeneity may be present even in a focus of a single biopsy core. These data are extremely important and urgently require further study to understand heterogeneity of genetic changes, because clinical studies have already been launched using genetic changes such as TMPRSS2-ERG fusions found in a single core biopsy to direct a specific therapy, including a trial using ABT-888 only in patients with this fusion now ongoing at CINJ.

Citation Format: Irina V. Tereshchenko, Hua Zhong, Marina Chekmareva, Urmila Santanam, Whitney Petrosky, Noriko Kane-Goldsmith, Jay A. Tischfield, Robert DiPaola. Rearrangement of ERG and CHD1 genes in prostate cancer as a marker of tumor heterogeneity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2233. doi:10.1158/1538-7445.AM2014-2233
Abstract 2235: High frequency of fusion transcripts involving TCF7L2 in colorectal cancer: Novel fusion partner and splice variants
Authors: Hoff Andreas M. , Nome Torfinn , Bakken Anne Cathrine , Rognum Torleiv O. , Nesbakken Arild , Skotheim Rolf I.
Published: May 2015

VTI1A-TCF7L2 was reported as a recurrent fusion gene in colorectal cancer (CRC), found to be expressed in three out of 97 primary cancers, and one cell line, NCI-H508, where a genomic deletion joins the two genes (Bass et al. Nature Genetics, 2011). To investigate this fusion further, we analyzed high-throughput DNA and RNA sequencing data from seven CRC cell lines in addition to publicly available data from the NCI-H508 cell line. From both genome and transcriptome data, the gene RP11-57H14.3 (ENSG00000225292) was identified as a novel fusion partner for TCF7L2 in the HCT 116 cell line. The originally reported fusion VTI1A-TCF7L2 was identified as expected from the analysis of NCI-H508 data. By triplicate nested RT-PCR, we tested both the novel fusion transcript and VTI1A-TCF7L2 for expression in a series of 106 CRC tissues, 21 colon cancer cell lines, 14 normal colonic mucosa, and 20 normal tissues from miscellaneous anatomical sites. Altogether, 42% and 45% of the CRC samples expressed VTI1A-TCF7L2 and TCF7L2-RP11-57H14.3 fusion transcripts, respectively. The two fusion transcripts were both seen in 29% of the normal colonic mucosa samples, and in 25% and 75% of the tested normal tissues from other organs. Seven different splice variants were detected for the VTI1A-TCF7L2 fusion, of which three are novel. Four different splice variants were detected for the TCF7L2-RP11-57H14.3 fusion.

In conclusion, we have identified novel variants of VTI1A-TCF7L2 fusion transcripts, including a novel fusion partner gene, RP11-57H14.3. We demonstrated detectable levels of TCF7L2 involving fusion transcripts in a large fraction of CRC samples, as well as in normal colonic mucosa and other normal tissue types, revealing that these fusions, at transcript level, are neither specific to cancer nor to the colon and rectum.

Citation Format: Andreas M. Hoff, Torfinn Nome, Anne Cathrine Bakken, Torleiv O. Rognum, Arild Nesbakken, Rolf I. Skotheim. High frequency of fusion transcripts involving TCF7L2 in colorectal cancer: Novel fusion partner and splice variants. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2235. doi:10.1158/1538-7445.AM2014-2235
Abstract 2237: Acquisition of biospecimens to support the Genotype-Tissue Expression (GTEx) project
Authors: Carithers Latarsha , Rao Abhi , Moore Helen
Published: May 2015

The Genotype-Tissue Expression (GTEx) project is an NIH Common Fund study that is aimed at understanding how genetic variation influences gene expression in normal tissues. Here we describe a biospecimen collection platform that was developed to address the logistical challenges of acquiring a large collection of high-quality tissues from rapid autopsy and organ donors for GTEx. This complex infrastructure is utilized to procure specimens under a rigorous quality management program and a stringent ethical, legal, and regulatory framework. Ultimately, data from the GTEx study will be used to evaluate gene regulatory mechanisms and their contribution to human disease development.

Citation Format: Latarsha Carithers, Abhi Rao, Helen Moore. Acquisition of biospecimens to support the Genotype-Tissue Expression (GTEx) project. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2237. doi:10.1158/1538-7445.AM2014-2237
Abstract 224: The role of Bmi-1 in benzyl isothiocyanate-mediated suppression of breast cancer stem cell self-renewal
Authors: Kim Su-Hyeong , Singh Shivendra V.
Published: May 2015

We have shown previously that benzyl isothiocyanate (BITC), a nontoxic small-molecule constituent of edible cruciferous vegetables (eg, garden cress) inhibits breast cancer stem cells (bCSC) in vitro and in vivo but the underlying mechanisms are not fully understood. Bmi-1 is a member of the polycomb group family of proteins that function in the epigenetic silencing of genes governing self-renewal, differentiation, and proliferation. The present study was undertaken to investigate the role of Bmi-1 in BITC-mediated inhibition of bCSC. BITC treatment downregulated protein level of Bmi-1 in cultured human breast cancer cells (MCF-7, SUM159 and MDA-MB-231), and in MDA-MB-231 xenografts in vivo. Ectopic expression of Bmi-1 alone was sufficient to drive stemness in MCF-7 cells. In addition, Bmi-1 overexpression in MCF-7 cells conferred marked protection against BITC-mediated inhibition of bCSC as evidenced by mammosphere frequency and aldehyde dehydrogenase 1 (ALDH1) activity. Consistent with these observations, silencing of Bmi-1 in SUM159 cells remarkably reduced the portion of bCSC. Apoptosis induced by BITC was also partially attenuated by overexpression of Bmi-1 in MCF-7 cells but its silencing had no effect on BITC-induced apoptosis in SUM159 cells. Previous studies have shown that miRNA128 inhibits cell proliferation and self-renewal by direct regulation of Bmi-1. BITC treatment modestly but statistically significantly increased the expression of miRNA128 in both MCF-7 and SUM159 cells. Inhibition of miRNA128 increased the mammosphere formation in MCF-7 and SUM159 cells and partially attenuated BITC-mediated inhibition of mammosphere formation. These observations led us to conclude that Bmi-1 is a novel target of BITC-mediated inhibition of bCSC. This study was supported by the grant RO1 CA129347 awarded by the National Cancer Institute.

Citation Format: Su-Hyeong Kim, Shivendra V. Singh. The role of Bmi-1 in benzyl isothiocyanate-mediated suppression of breast cancer stem cell self-renewal. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 224. doi:10.1158/1538-7445.AM2014-224
Abstract 2240: TERT promoter mutations are highly prevalent in bladder cancer and represent a potential new urinary biomarker
Authors: Hurst Carolyn D. , Platt Fiona M. , Knowles Margaret A.
Published: May 2015

Somatic mutation of the promoter region of the telomerase reverse transcriptase (TERT) gene was recently reported as a potential mechanism by which telomerase becomes activated in cancer cells. Here, we aimed to assess the frequency and spectrum of TERT promoter mutations in bladder cancer and detect mutations in voided urine. A SNaPshot assay designed to simultaneously interrogate two positions (-124bp and -146bp) previously reported as mutated in melanoma was used to screen 47 bladder cancer cell lines and 262 tumors. Forty cell lines and 201 tumors carried mutations at either -124bp or -146bp. Mutations at these positions were mutually exclusive. A germline mutation at -57bp (T>G) was previously reported in a melanoma-prone family. A second SNaPshot assay was designed to interrogate position -57bp and we screened all samples that were wildtype at positions -124bp and -146bp. Mutations at -57bp were detected in two cell lines and eight tumors. Matched blood samples were available for six of the tumors and one cell line, and somatic mutation status was confirmed. Sanger sequencing of the TERT core promoter in samples that were wildtype at positions -57bp, -124bp and -146bp revealed a further nine somatic mutations. Four mutations at positions -45bp, -72/-77bp, -141bp, and -54bp were novel, and all but one (-54bp) created new binding sites for ETS/TCF transcription factors. Overall, TERT promoter mutations were detected in 89% of cell lines and 83% of tumors. There was no association of TERT promoter mutation status with stage (p=0.1089, chi-square test) or grade (p=0.2438, chi-square test). As mutations are highly frequent in both non-muscle-invasive and invasive bladder cancer, these represent potential biomarkers for urine-based disease monitoring. For sixty-four tumors used in this study, matched urine samples were also collected prior to surgery. Fifty-three of the tumors had mutations at -124bp or -146bp and using the -124bp/-146bp SNaPshot assay we were able to detect 51 of these in the corresponding urine samples including all mutations (n=23) in 29 patients with low-grade noninvasive tumors. The use of TERT promoter mutation analysis in combination with other DNA-based biomarkers could markedly improve the sensitivity for urine-based detection of recurrent disease.

Citation Format: Carolyn D. Hurst, Fiona M. Platt, Margaret A. Knowles. TERT promoter mutations are highly prevalent in bladder cancer and represent a potential new urinary biomarker. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2240. doi:10.1158/1538-7445.AM2014-2240
Abstract 2242: Comprehensive genome methylation and whole genome expression analysis in penile carcinoma: Uncovering new molecular markers
Authors: Kuasne Hellen , Colus Ilce Mara de Syllos , Hernandez-Vargas Hector , Busso-Lopes Ariane Fernanda , Barros-Filho Mateus C. , Marchi Fabio A. , Scapulatempo-Neto Christovan , Faria Eliney F. , Lopes Ademar , Guimaraes Gustavo C. , Herceg Zdenko , Rogatto Silvia R.
Published: May 2015

Background: Penile carcinoma (PeCa) is frequently associated with high morbidity rates. Unlikely of the vast majority of tumors, there is no molecular markers described that are able to assist in diagnosis and prognosis or with potential to be therapeutic targets in PeCa.

Patients and methods: DNA methylation status (244K Human DNA Methylation Microarray platform, Agilent Technologies) and large-scale expression analysis (4x44K Whole Human Genome Microarray, Agilent Technologies) were performed in 35 and 37 PeCa, respectively. Quantitative bisulfite pyrosequencing (qBP) and RT-qPCR were used to validate the findings in 93 samples. HPV status was assessed using the Linear Array HPV Genotyping kit (Roche Molecular Diagnostics, CA, USA).

Results: Methylome analysis revealed 171 hypermethylated and 449 hypomethylated CpGs sites and the transcriptome profiling showed 2986 down- and 2817 over-expressed genes. HPV positivity was found in 32.7% of the cases, mainly the HPV16. The integrative analysis in 32 PeCa revealed a panel of 96 genes with inverse correlation between methylation and gene expression levels. The CpG hypermetlylation and gene downexpression, was confirmed for TWIST1, RSOP2, SOX3, SOX17, CD133, OTX2, HOXA3 and MEIS. In addition, BIRC5, DNMT1 and DNMT3B presented low levels of methylation and overexpression. The comparison of the results with clinical findings revealed that LIN28A, NKX2.2, NKX2.3, LHX5, BDNF, FOXA1 and CDX2 were associated with poor prognosis features.

Conclusion: Putative prognostic markers were detected revealing that DNA methylation modulates the expression of several genes in PeCa. These data may prove instrumental for biomarker discovery in clinics and molecular epidemiology of PeCa.

Citation Format: Hellen Kuasne, Ilce Mara de Syllos Colus, Hector Hernandez-Vargas, Ariane Fernanda Busso-Lopes, Mateus C. Barros-Filho, Fabio A. Marchi, Christovan Scapulatempo-Neto, Eliney F. Faria, Ademar Lopes, Gustavo C. Guimaraes, Zdenko Herceg, Silvia R. Rogatto. Comprehensive genome methylation and whole genome expression analysis in penile carcinoma: Uncovering new molecular markers. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2242. doi:10.1158/1538-7445.AM2014-2242
Abstract 2244A: Prion protein cross-talks with Notch1 to promote pancreatic ductal adenocarcinoma progression
Authors: Wang Yiwei , Huang Dan , Zhou Lan , Xin Wei
Published: May 2015

Human prion protein (PrP) is a glycosylphosphatidylino¬sitol (GPI)-anchored membrane protein and has been found to be up-regulated in some of the pancreatic ductal adenocarcinoma (PDAC). Current literature also suggests that over-expression of PrP induces inherent resistance to cancer therapeutics, while repression of PrP induces sensitization to cancer therapeutics. Our previous studies revealed the increased level of PrP expression associated with worse prognosis in PDAC patients. In PDACs, instead of forming normal GPI anchored protein, PrP exists as a pro-prion protein that binds Filamin A (FLNa) and exerts a cascade of activities subsequently augment the invasiveness of PDAC. In this study, we would like to investigate whether PrP also involves other signal transduction pathway in PDACs, especially Notch pathway.

Using immunoprecipitation and immunocytometry, we investigated the binding partners for PrP in several PDAC cell lines and found that PrP associates with Notch1 but not with Notch2 or Notch3 in cytoplasm and cell membrane. The association of PrP and Notch1 was further validated by confocal microscopy. Silencing PrP by siRNA not only decreased PrP expression, but also decreased Notch1 in the cytoplasm and the nuclear localized cleaved Notch1. Knocking down PrP in PDAC cells with high levels of PrP led to decreased cell proliferation and tumor invasion and increased apoptosis. PDAC cells showed Increased tumor necrosis after inhibition of PrP in xenografts, which accompanied by down-regulation of intracellular domain of Notch1 (Notch1NTM). In comparison, over-expression of PrP in PDAC cells with lower levels of PrP increased Notch1NTM expression, cell proliferation, tumor invasion, and xenograft tumor growth which could be blocked by the gamma secretase inhibitor. These findings suggest that PrP cross talking Notch1 in PDAC cells regulates pancreatic cancer cell survival and invasion. Elucidating the mechanisms for PrP mediated Notch activation in PDAC will further our long-term goal of understanding the role of oncogenic PrP in conferring pancreatic cancer aggressiveness, and potentially will lead to novel therapeutic options for treatment of pancreatic cancer.

Citation Format: Yiwei Wang, Dan Huang, Lan Zhou, Wei Xin. Prion protein cross-talks with Notch1 to promote pancreatic ductal adenocarcinoma progression. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2244A. doi:10.1158/1538-7445.AM2014-2244A
Abstract 2245: Mechanism of replicative senescence induced by SNF5 loss
Authors: Wei Darmood , Chung HoYoon , Simpson Dennis A. , Kaufmann William K. , Weissman Bernard E.
Published: May 2015

Cellular senescence was initially described as the phenomenon by which normal diploid cells cease to divide after reaching the Hayflick limit. However, with the identification of replicative senescence, other inducers of this process have appeared in the literature including oncogene induced senescence (OIS). Of interest, different oncogenes can initiate OIS through alternative pathways e.g. HRAS (G12V) and BRAF (V600E) that effect senescence through p16 dependent and p16 independent pathways, respectively. Alternatively, loss of tumor suppressor gene function can also induce senescence. In malignant rhabdoid tumors (MRTs), SNF5 behaves as a tumor suppressor and its re-expression in MRT cell lines results in growth arrest via the RB pathway. Paradoxically, when we knocked out SNF5 in normal human fibroblasts (NHF-1), we also observed senescence. We therefore hypothesize that perturbation of the SWI/SNF complex (e.g. knocking down SNF5) results in cellular senescence in most normal tissues. This suggests an epigenetic mechanism of senescence occurring through a p16 independent pathway, as previous publications have demonstrated a critical role for SNF5 in p16 expression. Additionally, in BRAF induced senescence, changes in levels of CD44, phospho-MEK1/2, and p16 are observed. However, we do not find these changes during SNF5 loss. Our findings suggest a relationship between SWI/SNF's chromatin remodeling activity and the regulation of cellular senescence. Importantly, our studies will identify the key signaling pathways that allow MRTs to escape cellular senescence after SNF5 loss. These results will further elucidate the role of SNF5 loss in MRT development and provide new targets for clinical intervention.

Acknowledgement:

We thank the members of the Weissman lab for their help, advice, and support and R01CA91048 (B.E.W.) , T32ES007126 (D.W.), UNC Dissertation Completion Fellowship (D.W.) for financial support.

Citation Format: Darmood Wei, HoYoon Chung, Dennis A. Simpson, William K. Kaufmann, Bernard E. Weissman. Mechanism of replicative senescence induced by SNF5 loss. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2245. doi:10.1158/1538-7445.AM2014-2245
Abstract 2246: Senescence as a result of impaired ribosome biogenesis
Authors: Lessard Frédéric , Bourdeau Véronique , Deschênes-Simard Xavier , Igelmann Sebastian , Montero Marinieve , Ferbeyre Gerardo
Published: May 2015

Senescence is a stable arrest of cell proliferation in which the cells remain viable and metabolically active but display a constitutive activation of the DNA damage response and of the tumor suppressors p53 and RB. The senescent phenotype can be induced by multiple stresses including short telomeres and oncogenes. We have shown that senescence, involves the ERK-dependent degradation of selective proteins involved in cell cycle progression and tumorigenesis. We call this process senescence associated protein degradation (SAPD) and it involves many nucleolar proteins that play a role in ribosome biogenesis. Using tritium pulse labelling we found a strong decrease of rRNA synthesis in senescent cells indicating that the degradation of nucleolar proteins is functionally relevant. Because we know how exactly the human 47S precursor rRNA is processed, it was possible to design primers on both sides of some processing sites and study their maturation by QPCR. In this way we showed defects in the processing of rRNA in senescent cells. Knocking down some of the nucleolar proteins degraded in senescence was sufficient to trigger the process indicating that a decrease in ribosome biogenesis is causal to cellular senescence. Mechanistically, the degradation of nucleolar proteins during senescence involves the ubiquitin-proteasome system suggesting that E3 ligases link the oncogenic stress that trigger senescence to nucleolar proteins degradation. We will discuss ongoing efforts to identify these enzymes.

Citation Format: Frédéric Lessard, Véronique Bourdeau, Xavier Deschênes-Simard, Sebastian Igelmann, Marinieve Montero, Gerardo Ferbeyre. Senescence as a result of impaired ribosome biogenesis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2246. doi:10.1158/1538-7445.AM2014-2246
Abstract 2247: Histone H3 cleavage via Cathepsin L drives a cellular senescence program
Authors: Duarte Luis F. , Young Andrew R. , Wu Hsan-Au , Panda Taniya , Wang Zichen , Maia Alexandre G. , Hasson Dan , Kapoor Avnish , Narita Masashi , Bernstein Emily
Published: May 2015

Oncogene-induced senescence (OIS) is characterized by irreversible loss of proliferative capacity. It has been proposed that OIS functions as a tumor suppressor mechanism, and several studies have demonstrated induction of senescence markers in pre-malignant lesions. In addition to profound morphological changes, significant alterations in chromatin structure occur during senescence including formation of senescence-associated heterochromatin foci (SAHF). We have used well-characterized senescence models in human primary lung fibroblasts and melanocytes to study how chromatin alterations contribute to the senescence phenotype. We discovered that the N'-terminal tail of histone H3 is proteolytically cleaved by the lysosomal protease Cathepsin L1 (CTSL1). We show that CTSL1 is induced during senescence both at the mRNA and protein levels, and that it is present in the chromatin fraction of senescent cells. Chemical and shRNA-mediated inhibition of CTSL1 activity effectively block H3 tail cleavage and leads to a SAHF formation defect. We identified two distinct cleaved forms of H3 in senescent cells, which are associated with chromatin and incorporated into nucleosomes. Using antibodies directed against specific post-translational modifications (PTMs) of the histone H3 tail, we have mapped the approximate locations of the cleavage sites. Furthermore, overexpression of flag-tagged cleaved versions of H3, but not the full-length histone, trigger robust senescence in IMR90 cells accompanied by a modest induction of SAHFs. Interestingly, we also detect cleaved H3 in chromatin extracted from benign nevi tissue samples, providing evidence of H3 tail cleavage in senescence models in vivo. Collectively, our results suggest that H3 tail cleavage may contribute significantly to the senescence phenotype, potentially through transcriptional regulation of key senescence target genes. This area of investigation is currently ongoing.

Citation Format: Luis F. Duarte, Andrew R. Young, Hsan-Au Wu, Taniya Panda, Zichen Wang, Alexandre G. Maia, Dan Hasson, Avnish Kapoor, Masashi Narita, Emily Bernstein. Histone H3 cleavage via Cathepsin L drives a cellular senescence program. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2247. doi:10.1158/1538-7445.AM2014-2247
Abstract 2248: The implication of STAT3 degradation and its mitochondrial functions in cellular senescence
Authors: Igelmann Sebastian , Deschênes-Simard Xavier , Lessard Frédéric , Bourdeau Véronique , Ferbeyre Gerardo
Published: May 2015

Cellular senescence is a stable growth arrest of the cell cycle considered to be a mechanism of tumor suppression. We have recently described a senescence associated protein degradation or SAPD process that targets many proteins required for tumor progression. The Signal transducer and activator of transcription 3 is one of the targets of SAPD and its downregualtion by RNAi is sufficient to trigger senescence. In the last decade several different functions of STAT3 have been described. Beyond its canonical role as a transcription factor, it has been shown that STAT3 is imported into mitochondria and regulate the activity of the respiratory chain. In this study we want to clarify which function of STAT3 is required to prevent senescence. To do that we have made several mutants of STAT3 affecting different domains of the protein and tested their ability to rescue the senescence response to an anti-Stat3 shRNA. We have found that phosphorylation on residue S727 of STAT3 is important to reestablish proliferation in shSTAT3 induced senescence. Further, transcriptional activity of STAT3 is not required to rescue senescence as a mutant unable to enter the nucleus or the transcriptionally death STAT3 K180A are both able to bypass shSTAT3-induced senescence. Moreover, phosphorylation on Y705 seems not to be important as Y705F STAT3 is also able to reestablish proliferation. All in all, it seems that mitochondrial function of STAT3 is required to prevent senescence since STAT3 S727 phosphorylation is linked to the mitochondrial import of STAT3.

Citation Format: Sebastian Igelmann, Xavier Deschênes-Simard, Frédéric Lessard, Véronique Bourdeau, Gerardo Ferbeyre. The implication of STAT3 degradation and its mitochondrial functions in cellular senescence. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2248. doi:10.1158/1538-7445.AM2014-2248
Abstract 225: Overexpression of truncated Ron augments benzyl isothiocyanate-induced apoptosis in human breast cancer cells
Authors: Sehrawat Anuradha , Singh Shivendra V.
Published: May 2015

Phytochemicals from edible plants remain attractive for prevention of breast cancer, which is a leading cause of cancer-related mortality in women worldwide. We have shown previously that benzyl isothiocyanate (BITC), a small-molecule found in edible cruciferous vegetables such as garden cress, confers significant protection against mammary cancer development in MMTV-neu transgenic mice. BITC-mediated prevention of mammary carcinogenesis in MMTV-neu mice in vivo is associated with tumor cell apoptosis induction and elimination of breast cancer stem cells (bCSC). In cultured human breast cancer cells, BITC-induced apoptosis is accompanied by activation of multidomain proapoptotic protein Bax whereas inhibition of bCSC self-renewal is significantly attenuated by overexpression of truncated Ron (recepteur d’origine nantais) tyrosine kinase. Ron expression is very low in normal human breast epithelium but its overexpression has been reported in about 50% of primary breast cancers. In addition, mammary-specific overexpression of wild-type Ron as well as its constitutively active form induces highly metastatic breast tumors in mice. The present study was undertaken to study the role of truncated Ron (sfRon) in BITC-induced apoptosis using MCF-7 and MDA-MB-361 human breast cancer cells. Overexpression of sfRon resulted in statistically significant augmentation of BITC-induced apoptotic cell death in both cell lines as judged by flow cytometry after staining with Annexin V and propidium iodide, analysis of histone associated DNA fragment release into the cytosol, and/or cell proliferation assay. On the other hand, sfRon overexpressing MCF-7 cells were resistant to BITC-mediated G2/M phase cell cycle arrest when compared with parental cells. Increased apoptosis following BITC treatment in sfRon overexpressing cells was due to enhanced activation of Bax and/or Bak. Taken together, these findings indicate that overexpression of sfRon sensitizes human breast cancer cells to BITC-induced apoptosis. This investigation was supported by the NCI grant CA129347-06.

Citation Format: Anuradha Sehrawat, Shivendra V. Singh. Overexpression of truncated Ron augments benzyl isothiocyanate-induced apoptosis in human breast cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 225. doi:10.1158/1538-7445.AM2014-225
Abstract 2250: Role of single strand binding protein 1 in TERT recruitment to telomeres and in maintaining telomere G-overhangs
Authors: Pandita Tej K. , Pandita Raj
Published: May 2015

Telomeres cap the ends of all eukaryotic chromosomes from degradation or end-to-end fusions. Mammalian telomeres are maintained by telomerase which is a ribonucleoprotein complex that includes a reverse transcriptase catalytic unit called TERT and a functional telomerase RNA (TR or TERC), which acts as a template to maintain telomere length. Telomerase maintains telomere length, by overcoming incomplete lagging strand synthesis known as the “end replication problem” during DNA replication. Mammalian stem and cancer cells utilize telomerase to extend telomeric G-overhangs to partially (stem cells) or completely (tumor cells) maintain telomere ends. Cells expressing telomerase also have higher levels of single strand binding 1 (SSB1) protein, which has a known critical role in the DNA damage responses and double strand break repair specifically by homologous recombination. We have established that SSB1 binds specifically to G-strand telomeric DNA in vitro, and associates in vivo with telomeres both in human and mouse cell lines. Interestingly, SSB1 interacts with the TERT protein, and determines the amount of TERT interaction with telomeres. Deletion/depletion of SSB1 reduces TERT interaction with telomeres and leads to G-overhang loss, but has no effect on in vitro telomerase activity. We will discuss, how SSB1 is required for telomerase recruitment to telomeres to facilitate G-strand DNA extension and to maintain telomeres.

Citation Format: Tej K. Pandita, Raj Pandita. Role of single strand binding protein 1 in TERT recruitment to telomeres and in maintaining telomere G-overhangs. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2250. doi:10.1158/1538-7445.AM2014-2250
Abstract 2256: p53 gene expression is epigenetically regulated during replicative senescence in keratinocytes
Authors: Kim Reuben H. , Kang Mo K. , Kim Terresa , Yang Paul , Hong Christine , Shin Ki-Hyuk , Park No-Hee
Published: May 2015

Cancer is one of the most prevalent age-associated diseases in elderly populations and is largely attributed to accumulation of genetic mutations secondary to the loss of tumor suppressive functions. p53 is a potent tumor suppressor that is implicated in organismal aging in vivo and replicative senescence in vitro; however, its transcriptional regulation during replicative senescence in cells remain unclear. The current study was undertaken to examine the epigenetic regulation of p53 transcription during replicative senescence in normal human keratinocytes. Normal human oral keratinocytes (NHOKs) undergoing senescence upon serial subcultures exhibited a decrease in p53 gene expression. Methylation status of two CpG islands in the p53 promoter was evaluated by treating with 5-aza-CdR, a demethylating agent, and by using bisulfite methylation sequencing analysis. Status of histone modification was evaluated using Chromatin Immunoprecipitation (ChIP) assay with anti-H3K27me3, a repressive mark, and anti-H3Ac, an active mark at the 5 different sites of the p53 promoter regions. In the presence of 5-aza-CdR, there was no increase in p53 expression in senescent NHOK at both mRNA and protein level. Bisulfite-sequencing analysis revealed no methylation changes at the CpG islands in the p53 promoters from young and senescent NHOK. However, during replicative senescence in NHOK, there were notable enrichment of anti-H3K27me3 occupancy and significant decrease in anti-H3Ac occupancy in the p53 promoter regions, indicating that chromatin at the p53 locus undergo compaction during replicative senescence. Our study demonstrates that the diminution of p53 during replicative senescence is epigenetically regulated in human keratinocytes.

Citation Format: Reuben H. Kim, Mo K. Kang, Terresa Kim, Paul Yang, Christine Hong, Ki-Hyuk Shin, No-Hee Park. p53 gene expression is epigenetically regulated during replicative senescence in keratinocytes. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2256. doi:10.1158/1538-7445.AM2014-2256
Abstract 226: Chemopreventive potential of curcumin in prostate cancer
Authors: Ide Hisamitsu , Lu Yan , Yamaguchi Raizo , Muto Satoru , Horie Shigeo
Published: May 2015

Curcumin contains multiple active compounds that have potent anti-inflammatory and anti-cancer properties. The present study investigates the molecular mechanisms of curcumin in the prostate cancer models. Treatment of LNCaP and 22Rv-1 prostate cancer cells with curcumin inhibited the cell proliferation. Genome-wide expression profiling, using oligonucleotide microarrays, revealed genes differentially expressed with curcumin treatment in prostate cancer cells. We identified one of the up-regulated genes, Aldo-Keto reductase 1C-2(AKR1C-2) which is involved in testosterone metabolism. Quantitative PCR confirmed significant up-regulation of AKR-1C2. The suppression of tumor growth is associated with the inhibition of expression of androgen receptor and androgen metabolism through AKR1C2 leading the induction of cell death by apoptosis. These results suggest that curcumin natural bioactive compounds could have potent anti-cancer property and the regulation of both androgen and androgen receptor. These pathways could exert pleiotrophic effects in prevention and treatment of prostate cancer.

Citation Format: Hisamitsu Ide, Yan Lu, Raizo Yamaguchi, Satoru Muto, Shigeo Horie. Chemopreventive potential of curcumin in prostate cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 226. doi:10.1158/1538-7445.AM2014-226
Abstract 2260: c-Myc induced senescence is dependent on transactivation of actin depolymerizing factor (ADF)/cofilin to mediate potent bystander effect
Authors: Lin Shih-Ting , Tsai Cheng-Han , Lee Yi-Jang , Lo Chia-Chien , Chou Yen-Ting
Published: May 2015

Oncogene-induced senescence may prevent unlimited proliferation and lead to cell death. This phenomenon is likely to be associated with up-regulation of p53 and p16INK4 tumor suppressor. Morphological transformation is also an important feature of senescence that exhibits enlarged cell volume and flat morphology. In this study, we found that c-Myc was co-upregulated with cofilin-1, an actin-binding protein essential for actin dynamics and morphological maintenance in replicative senescence. Additionally, oxidative stress induced senescent phenotypes also accompanied by up-regulation of both c-myc and cofilin-1. Knockdown of cofilin-1 was sufficient to minimize the cell shapes and senescence associated β-galactosidase marker that was induced by over-expressed c-myc or oxidative stress. Over-expression of cofilin-1 was also sufficient to induce senescence in the absence of p53 and p16INK4, but was unable to influence c-Myc level. On the other hand, over-expression of c-Myc could induce cofilin-1 expression. We next found that c-myc was able to induce the transactivation of cofilin-1 gene according to results of quantitative polymerase chain reaction (qPCR) and luciferase reporter gene assay. Furthermore, the results of ChIP assay provided evidence that c-myc can bind to two proximal E-box elements nearby the transcriptional initiation on cofilin-1 gene promoter. Although c-myc promoted cellular senescence in vitro, the conditional medium from c-myc over-expressing A549 cells accelerated migration and proliferation in separate cultured cells without c-myc over-expression. Knockdown of cofilin-1 also alleviated this bystander effect caused by over-expression of c-myc. Therefore, c-myc oncogene may require cofilin-1 for cell senescence that influences the growth of other tumor cells. Taken together, current data provide evidence that c-myc/cofilin-1 signaling pathway is a novel mechanism on promoting oncogene-induced cellular senescence and tumor progression.

Citation Format: Shih-Ting Lin, Cheng-Han Tsai, Yi-Jang Lee, Chia-Chien Lo, Yen-Ting Chou. c-Myc induced senescence is dependent on transactivation of actin depolymerizing factor (ADF)/cofilin to mediate potent bystander effect. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2260. doi:10.1158/1538-7445.AM2014-2260
Abstract 2265: Sub-fractions of conditioned medium, from hypoxia-induced cells with multipotent potential, exhibit a significant anti-oncogenic activity
Authors: Pinney Emmett , Montes-Camacho Mayra , Brintle Kayler , Posch Christian , Menen Rhiana , Easter David , Bouvet Michael , Hoffman Robert , Naughton Gail
Published: May 2015

We have previously shown that a human soluble cell conditioned media (CCM) from hypoxia-induced, potentially multipotent cells, grown in a perfusion bioreactor system inhibits tumor cell growth in vitro and in two animal models. Here we present data that indicate that a low molecular weight sub-fraction from the CCM has an increased anti-oncogenic activity. This activity was demonstrated in orthotopic pancreatic models, nRAS melanoma assays and over 10 additional human cancer cell lines in vitro. In an orthotopic BxPC3 pancreatic cancer model, there was a statistically significant improvement in survival following weekly intravenous injections of 0.1ml of sub fractioned CCM, as well as a significant improvement with 0.5ml intraperitoneal injections post-resection, as measured by Kaplan Meier analysis (p<0.05) (figures below). Uveal and nRAS melanoma cancer cell growth was inhibited by greater than 90% in in-vitro proliferation assays. The mechanism of action of the CCM is the induction of apoptosis through the upregulation of Caspase 3 and 9, as demonstrated by immunostaining of Annexin V and immunoblot ayalysis. In a Miapaca nude mouse daily dose study, tumor mass and metastatic extent was substantially reduced in treated mouse versus the control. Further, in a BXPC3 nude mouse daily dose study, 60% (p<0.05) of treated mice were tumor free after four weeks of treatment, while only 10% were tumor free in the control group . Little or no apparent drug toxicity was observed. These results indicate that the CCM sub-fraction could be a useful raw material for a treatment for a large range of neoplastic diseases.

Citation Format: Emmett Pinney, Mayra Montes-Camacho, Kayler Brintle, Christian Posch, Rhiana Menen, David Easter, Michael Bouvet, Robert Hoffman, Gail Naughton. Sub-fractions of conditioned medium, from hypoxia-induced cells with multipotent potential, exhibit a significant anti-oncogenic activity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2265. doi:10.1158/1538-7445.AM2014-2265